Sample records for maculatus fab coleoptera

  1. Activités insecticides de Striga hermonthica (Del.) Benth (Scrophulariaceae) sur Callosobrichus maculatus (Fab.) (Coleoptera : Bruchidae)


    Nacoulma OG.; Ouedraogo AP.; Kiendrebeogo M.


    Insecticidal activities of Striga hermonthica (Del.) Benth (Scrophulariacecae) on Callobruchus maculatus (Fab.) (Coleptera Bruchidae). This paper deals with insecticidal potentialities of Striga hermonthica (Del.) (Scrophulariaceae) in protection of cowpea Vigna unguculata (L.) Walp against Callosobruchus maculatus (Fab.) (Coleoptera: Bruchidae) during storage. Crude acetone extract at 0,5% w/w (100 mg of extract for 20 g of grain) exhibits 48% of ovicidal effect and then reduces by half emer...

  2. Activités insecticides de Striga hermonthica (Del. Benth (Scrophulariaceae sur Callosobrichus maculatus (Fab. (Coleoptera : Bruchidae

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    Nacoulma OG.


    Full Text Available Insecticidal activities of Striga hermonthica (Del. Benth (Scrophulariacecae on Callobruchus maculatus (Fab. (Coleptera Bruchidae. This paper deals with insecticidal potentialities of Striga hermonthica (Del. (Scrophulariaceae in protection of cowpea Vigna unguculata (L. Walp against Callosobruchus maculatus (Fab. (Coleoptera: Bruchidae during storage. Crude acetone extract at 0,5% w/w (100 mg of extract for 20 g of grain exhibits 48% of ovicidal effect and then reduces by half emergence rate of adult beetles at the first generation. This extract shows a weak insecticide activity against adults of C. maculatus. Petroleum ether fraction (0,4% w/w of the crude extract reveals ovicidal (51% and larvicidal (72% effects which reduce the emergence rate of adults to only 9%. LD50 and LD90 are monitored during crude extract fractionation to follow ovicidal and larvicidal compounds and to evaluate their efficacy during the isolation procedure. One fraction, mainly composed of two triterpenoid compounds has been identified as responsible of the ovicidal activity of S. hermonthica while the origin of the larvicidal activity hasn’t been identified.

  3. Impact of the egg parasitoid Uscana lariophaga and the larval-pupal parasitoid Dinarmus basalis on Callosobruchus maculatus populations and cowpea losses

    NARCIS (Netherlands)

    Huis, van A.; Alebeek, van F.A.N.; Es, van M.; Sagnia, S.B.


    In West Africa, Uscana lariophaga Steffan (Hymenoptera: Trichogrammatidae) and Dinarmus basalis (Rondani) (Hymenoptera: Pteromalidae) parasitize the eggs and larvae, respectively, of Callosobruchus maculatus Fab. (Coleoptera: Bruchidae), an important pest of stored cowpea. The impact of the

  4. Attraction of Callosobruchus maculatus (F.) (Coleoptera: Bruchidae) to four varieties of Lathyrus sativus L. seed volatiles. (United States)

    Adhikary, P; Mukherjee, A; Barik, A


    Callosobruchus maculatus (F.) (Coleoptera: Bruchidae) is an important stored grain pest of Lathyrus sativus L. (Leguminosae), commonly known as khesari, in India, Bangladesh and Ethiopia. Volatiles were collected from four varieties, i.e., Bio L 212 Ratan, Nirmal B-1, WBK-14-7 and WBK-13-1 of uninfested khesari seeds, and subsequently identified and quantified by gas chromatography mass spectrometry and gas chromatography flame ionization detector analyses, respectively. A total of 23 volatiles were identified in the four varieties of khesari seeds. In Bio L 212 Ratan and WBK-13-1 seeds, nonanal was the most abundant followed by farnesyl acetone; whereas farnesyl acetone was predominant followed by nonanal in Nirmal B-1 and WBK-14-7 khesari seeds. The olfactory responses of female C. maculatus toward volatile blends from four varieties of khesari seeds, and individual synthetic compounds and their combinations were examined through Y-shaped glass tube olfactometer bioassays. Callosobruchus maculatus showed significant preference for the whole volatile blends from Bio L 212 Ratan seeds compared to whole volatile blends from other three varieties. The insect exhibited attraction to five individual synthetic compounds, 3-octanone, 3-octanol, linalool oxide, 1-octanol and nonanal. A synthetic blend of 448, 390, 1182, 659 and 8114 ng/20 μl methylene chloride of 3-octanone, 3-octanol, linalool oxide, 1-octanol and nonanal, respectively, was most attractive to C. maculatus, and this combination might be used for insect pest management program such as baited traps.

  5. Determination of fluoxetine in Dermestes maculatus (Coleoptera: Dermestidae) by a spectrophotometric method. (United States)

    Zanetti, Noelia I; Ferrero, Adriana A; Centeno, Néstor D


    The aims of this study were to detect and quantify fluoxetine, an antidepressant, from entomological samples. Larvae, pupae and adults of Dermestes maculatus (Coleoptera, Dermestidae) were reared on pig muscle previously treated with fluoxetine. The concentration selected, 2000mg/kg, emulates a fluoxetine overdose lethal to humans and laboratory animals. Thirty larvae on the fourth and fifth stages, 50 adults and several exuviae were analyzed for fluoxetine content. Detection of fluoxetine was performed by UV spectrophotometry at 270 and 277nm. All developmental stages of D. maculatus and exuviae were positive for fluoxetine. We also quantified the drug and no significant differences were found either between the days or the stages in the general model, but at 277nm a tendency of the concentration to decrease with time was observed. Concentrations of fluoxetine at 277nm were almost equal or greater than those at 270nm. This is the first study to detect and quantify fluoxetine from entomological samples and, in particular, from D. maculatus beetles. Copyright © 2016 The Chartered Society of Forensic Sciences. Published by Elsevier Ireland Ltd. All rights reserved.


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    Antonia Mirian Nogueira de Moura Guerra


    Full Text Available This study aimed to evaluate the insecticidal activity of eight medicinal plants on Callosobruchus maculatus (F. (Coleoptera: Bruchidae. Plant powder from Rosmarinus officinalis (L. leaves, Peumus boldus (Mol leaves, Matricaria chamomilla (L. flowers, Baccharis trimera (Less. leaves, Camellia sinensis (L. leaves, Thea sinensis (L. leaves, Ilex paraguariensis (St. Hil. leaves, and fruits of Pimpinella anisum (L. were used in the experiment. Bioassays were carried out under constant conditions of temperature (28±2 °C, relative humidity (70±5% and scotophase of 24 hours. Experimental units consisted of 20 grams of cowpea (Vigna unguiculata (L. Walp. grains with 13% wet basis moisture content, 2 g plant powder and ten adult insects, both placed in plastic containers (60 mm in diameter x 60 mm high. The analyzed variables included: insect mortality, oviposition (egg number and instantaneous rate of population growth (ri. The number of dead insects was recorded 5 days after bioassays were started, the egg number was recorded after 12 days, and ri was estimated on the 60th day. All the tested plants had toxicity to C. maculatus. The highest mortality was found in insects treated with P. boldus and I. paraguariensis. Number of eggs and ri also varied significantly among the powders. The lowest mean values for these parameters were recorded in the plots treated with P. boldus, P. anisum and R. officinalis.

  7. Biological Strategies of Dermestes maculatus DeGeer (Coleoptera: Dermestidae) at Larval Stages in Different Temperatures. (United States)

    Zanetti, N I; Visciarelli, E C; Centeno, N D


    The intraspecific variation in larval instars is a widely distributed phenomenon amongst holometabolous insects. Several factors can affect the number of instars, such as temperature, humidity, and density. Only a few references could be found in the literature because the invariability in the number of larval instars is considered normal, and the issue has raised little to no interest. Despite this, no study to date has intended to assess or focus on the larval development. Here, we analyzed the effect of different rearing temperature on the larval stage of Dermestes maculatus DeGeer (Coleoptera: Dermestidae). The results indicated that at all temperatures, L5 represented a decisive point for individuals as well as the other later larval instars, because the next step to follow was to pupate or molt to the next larval instar. Furthermore, there were mainly two populations, L5 and L6, although in different proportions according to temperature. We also found that at a greater number of instars, the larval development at all temperatures lasted longer. Moreover, the exponential model was the best adjustment in the developmental time of all populations as well as for the accumulated developmental time of L1-L4. Thus, we conclude that random factors such as genetics could probably cause interspecific variability in D. maculatus larval development.

  8. Cowpea Vicilins: Fractionation of Urea Denatured Sub-Units and Effects on Callosobruchus maculatus F. (Coleoptera: Bruchidae Development

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    Antônio Chagas Mota


    Full Text Available Vicilins (7S storage globulins isolated from cowpea (Vigna unguiculata L. seeds which were susceptible (S and resistant (R to the cowpea weevil (Callosobruchus maculatus F., Coleoptera: Bruchidae were denatured by urea and fractionated by ion-exchange chromatography. Isolated fractions were incorporated in artificial seeds for assessment of their toxicity to C. maculatus. The most acidic fractions of both susceptible (CE-31 cultivar and resistant (IT81D-1045 line seeds were shown to affect development and survival of the bruchid. Results indicated that vicilin polypeptides of toxic nature were expressed in both types of storage globulins although at different levels.Vicilinas (globulinas de reserva 7S isoladas de sementes de feijão-de-corda (Vigna unguiculata L., susceptíveis (S e resistentes (R ao caruncho/gorgulho (Callosobruchus maculatus F., Coleoptera: Bruchidae foram desnaturadas por uréia e fracionadas por cromatografia de troca iônica. As frações isoladas foram incorporadas em sementes artificiais para avaliação de sua toxicidade a C. maculatus. As fracões mais ácidas de ambas vicilinas afetaram o desenvolvimento e a sobrevivência do bruquídeo. Sugerimos que polipeptídeos de vicilinas de natureza tóxica são expressos em ambos tipos de globulinas de reserva, embora em níveis diferentes.

  9. Jatropha curcas (L. and Jatropha gossypifolia (L., Botanical Entomocides For Poor Resource Farmers As Protectants Of Cowpea Seeds Against Infestation by Callosobruchus maculatus (Fab. (Coleoptera: Bruchidae - See more at:

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    Ileke Kayode David


    Full Text Available The powder, wood ash and latex of Jatropha carcus and Jatropha gossypifolia were bioassayed for their insecticidal potential against cowpea bruchid, Callosobruchus maculatus in the laboratory at ambient temperature and humidity. The plant wood ash, powder and latex were tested at rate 2g/20g of cowpea seeds. Adult mortality and adult emergence of the insect were investigated. Result showed that wood ash of J. carcus evoked 100% mortality of adult cowpea bruchid after 4 days of application. This is followed by powder of J. carcus which caused 92.5% mortality while powder of J. gossypifolia was the least effective that caused 90% mortality of adult bruchids. There was no adult emergence in seeds treated with wood ash and latex of both J. carcus and J. gossypifolia compared with untreated that had 81.1% adult emergence. The result obtained from this study revealed that J. carcus and J. gossypifolia wood ash and latex were effective in controlling C. maculatus and could serve as an alternative to synthetic insecticides for the protection of stored cowpea against cowpea bruchid for the poor resource farmers in developing countries. -

  10. Inbreeding depression in two seed-feeding beetles, Callosobruchus maculatus and Stator limbatus (Coleoptera: Chrysomelidae). (United States)

    Fox, C W; Scheibly, K L; Smith, B P; Wallin, W G


    Inbreeding depression is well documented in insects but the degree to which inbreeding depression varies among populations within species, and among traits within populations, is poorly studied in insects other than Drosophila. Inbreeding depression was examined in two long-term laboratory colonies of the seed beetle, Callosobruchus maculatus (Fabricius), which are used frequently as models for experiments in ecology, evolution and behaviour. Inbreeding depression in these laboratory colonies are compared with one recently field-collected population of a different seed beetle, Stator limbatus Horn. Inbreeding reduced embryogenesis, egg hatch and larval survival in both species, such that eggs produced by sib matings were >17% less likely to produce an adult offspring. Inbred larvae also took 4-6% longer to develop to emergence in both species. Inbreeding depression varied among the measured traits but did not differ between the two populations of C. maculatus for any trait, despite the large geographic distance between source populations (western Africa vs. southern India). Inbreeding depression was similar in magnitude between C. maculatus and S. limbatus. This study demonstrates that these laboratory populations of C. maculatus harbour substantial genetic loads, similar to the genetic load of populations of S. limbatus recently collected from the field.

  11. Alternation of cowpea genotypes affects the biology of Callosobruchus maculatus (fabr. (Coleoptera: Bruchidae

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    Lima Marcileyne Pessôa Leite de


    Full Text Available Callosobruchus maculatus (Fabr. is an important pest in stored cowpea, Vigna unguiculata (L. Walp., with ample distribution in tropical and subtropical regions. The effect of alternation of cowpea genotypes, susceptible (S and resistant (R, on the biology of (C. maculatus was studied after four generations. A no-choice test was carried out in a completely randomized design, factorial scheme, with five treatments, four host combinations (RR, RS, SR and SS and five replications. Each replication consisted of 30 grains of each genotype infested by two insect couples. The number of eggs per female was not different within or between combinations, evidencing that the genotypes and their alternation did not affect C. maculatus fecundity. Egg viability, however, varied between genotypes and between combinations. In combination RR, the longest duration of the immature stage was verified for genotype IT89KD-245; in addition, all genotypes presented the smallest survival for the same stage, resulting in a higher mortality of the pest. The resistance index categorized combination RR as moderately resistant for genotypes IT89KD-245, BR14-Mulato and BR17-Gurguéia, and as susceptible (S only for IT89KD-260, demonstrating that these combinations were not very adequate for the development of C. maculatus, a fact that was confirmed by the better performance of the pest on the genotype from combination SS, and because of a reduction in its performance when it returned to resistant genotypes.

  12. Modification of postmortem wounds by Dermestes maculatus (Coleoptera: Dermestidae) activity: A preliminary study. (United States)

    Zanetti, Noelia I; Ferrero, Adriana A; Centeno, Néstor D


    The purpose of this analysis was to evaluate the effects of insect activity on hacking trauma which was inflicted with a small cleaver and a razor blade under controlled conditions. Three pig hooves were each subjected to a blow with a small cleaver and a cut with a razor blade prior to insect exposure. We used Dermestes maculatus DeGeer 1774 species. These beetles made principally depressions and destruction on both wounds, and bites were observed on the edges of the wounds. As time passed and insects fed and refuge, chop marks were deformed and disappeared, taking this less than a month. Thus, D. maculatus could mask postmortem wounds and probably premortem wounds, and so the cause of death. Copyright © 2015 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.

  13. Control of Cowpea Weevil, Callosobruchus Maculatus (F.) (Coleoptera: Bruchidae), Using Natural Plant Products


    Bamphitlhi Tiroesele; Kesegofetse Thomas; Seipati Seketeme


    A laboratory study was conducted to investigate the effects of natural products on the reproduction and damage of Callosobruchus maculatus, the cowpea weevil, on cowpea seeds at Botswana College of Agriculture in Gaborone, Botswana. The cowpea variety Blackeye was used in the study. Fifty grams of each plant product (garlic, peppermint and chilies) was added to 500 g of the cowpea seeds. Findings of this experiment revealed that chilies and garlic had negative effects on cowpea weevils for al...

  14. BIOATIVIDADE DE Solanum melongena L. E Capsicum annuum L. SOBRE Callosobruchus maculatus (COLEOPTERA: BRUCHIDAE

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    Glauciene Ferreira Freire


    Full Text Available RESUMOO uso contínuo e indiscriminado de produtos químicos na agricultura pode trazer sérios prejuízos à saúde humana e ao meio ambiente. Uma opção é o emprego de plantas com ação inseticida. Diante do exposto, o objetivo desse trabalho foi avaliar a atividade inseticida do pó de folhas de Solanum melongena L. e Capsicum annuum L. contra Callosobruchus maculatus. O experimento foi conduzido no Laboratório de Entomologia da Universidade Federal de Campina Grande (UFCG, Campus de Pombal, Paraíba, Brasil. Os grãos de feijão-caupi foram tratados com os pós nas concentrações 0,0; 2,5; 5,0 e 10,0 % [100*(massa do pó/massa de grãos] e realizados testes de sobrevivência e repelência contra C. maculatus. Os dados da sobrevivência foram analisados utilizando o teste de Log-rank (p ≤ 0,05, pelo método de D-collet e para a repelência utilizou-se o teste do Qui-Quadrado (p ≤ 0,05. Todos os pós e concentrações avaliadas foram repelentes contra C. maculatus, com exceção do pó de C. annuum na concentração de 2,5 %. No que se refere à sobrevivência, ambas as espécies vegetais causaram elevada mortalidade em C. maculatus, com morte total dos insetos em até 120 h.ABSTRACTThe continuous and indiscriminate use of chemicals in agriculture can bring serious problems to human health and the environment. One option is the use of plants with insecticidal action. Given the above, the aim of this work was to evaluate the insecticide activity powder of leaves of Solanum melongena L. and Capsicum annuum L. against Callosobruchus maculatus in three concentrations. The experiment was conducted at the Laboratory of Entomology, Federal University of Campina Grande (UFCG, Campus de Pombal, Paraiba, Brazil. The grains of cowpea were treated with the powders in concentrations 0.0, 2.5, 5.0 and 10.0 % [100*(mass of powder/ mass of grains] and performed tests of survival and repellency against C. maculatus. Survival data were analyzed using

  15. Condition dependence of male and female genital structures in the seed beetle Callosobruchus maculatus (Coleoptera: Bruchidae). (United States)

    Cayetano, L; Bonduriansky, R


    Theory predicts that costly secondary sexual traits will evolve heightened condition dependence, and many studies have reported strong condition dependence of signal and weapon traits in a variety of species. However, although genital structures often play key roles in intersexual interactions and appear to be subject to sexual or sexually antagonistic selection, few studies have examined the condition dependence of genital structures, especially in both sexes simultaneously. We investigated the responses of male and female genital structures to manipulation of larval diet quality (new versus once-used mung beans) in the bruchid seed beetle Callosobruchus maculatus. We quantified effects on mean relative size and static allometry of the male aedeagus, aedeagal spines, flap and paramere and the female reproductive tract and bursal spines. None of the male traits showed a significant effect of diet quality. In females, we found that longer bursal spines (relative to body size) were expressed on low-quality diet. Although the function of bursal spines is poorly understood, we suggest that greater bursal spine length in low-condition females may represent a sexually antagonistic adaptation. Overall, we found no evidence that genital traits in C. maculatus are expressed to a greater extent when nutrients are more abundant. This suggests that, even though some genital traits appear to function as secondary sexual traits, genital traits do not exhibit heightened condition dependence in this species. We discuss possible reasons for this finding.

  16. The Effect of Temperature and Laboratory Rearing Conditions on the Development of Dermestes maculatus (Coleoptera: Dermestidae). (United States)

    Zanetti, Noelia I; Visciarelli, Elena C; Centeno, Néstor D


    Experiments were conducted to study the life cycle of Dermestes maculatus and to establish the total developmental time and the developmental time of immature stages, in relation with six different temperatures. We also analyzed the variations in size, morphology, and other indicators of temporal variation during life cycle of D. maculatus, in relation with temperature. One hundred larvae were selected per experiment, reared individually. The remaining larvae were reared to evaluate and establish temporal variations among the instars (length, cephalic width, and dry weight). In all trials, survivorship was greater than 50% and seven larval instars were found. Data of the average developmental time of immature stages and of the total cycle, at different temperatures, are provided. This is of relevance when estimating particularly, a minimum PMI. No relation between morphometric parameters and temperature was found, suggesting that other random factors may have been involved. Thus, this indicates that the method of isomegalen diagrams could not be used for calculating PMI. © 2015 American Academy of Forensic Sciences.

  17. Insecticidal activity of 2-tridecanone against the cowpea weevil Callosobruchus maculatus (Coleoptera: Bruchidae

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    Yussef F.B. Braga


    Full Text Available The effect of 2-tridecanone vapor on the cowpea weevil (Callosobruchus maculatus development was determined. Seeds of cowpea were infested with adults and exposed to different doses of 2-tridecanone isolated from Pilocarpus microphyllus Stapf ex Holm, a plant species native from northeastern Brazil. The pure monoterpene was evaluated both undiluted as well as in the dilutions 1:10, 1:100 and 1:1,000 (v/v. The following parameters of the cowpea weevil life cycle were analyzed in response to decreasing doses of 2-tridecanone: number of eggs laid, percentage of egg hatching on seeds, percentage of adult emergence, adult weight at emergence, mean developmental time and number of adults emerged. Vapor of 2-tridecanone caused a significant (P O efeito dos vapores da 2-tridecanona sobre o caruncho do feijão-de-corda (Callosobruchus maculatus foi avaliado. Sementes de feijão-de-corda infestados com insetos adultos foram expostas a diferentes doses de 2-tridecanona isolada de Pilocarpus microphyllus, uma espécie nativa do Nordeste do Brasil. O monoterpeno puro foi utilizado nas diluições 1:10, 1:100 e 1:1000 (v/v. Os parâmetros da biologia do inseto foram analisados em função da resposta a doses decrescentes de 2-tridecanona: número de ovos postos por fêmea, percentagem de eclosão de ovos, percentagem de emergência de adultos, peso dos adultos recém-emergidos, tempo médio de desenvolvimento e número total de ovos emergidos. Diferenças significativas (P < 0.05 entre as doses de 2-tridecanona testadas foram observadas, para quatro dos seis parâmetros biológicos analisados. Os resultados obtidos indicaram que a 2-tridecanona é tóxica para C. maculatus, reduzindo significativamente (P < 0.05 o número de insetos emergidos após a infestação. Esse efeito foi causado principalmente pela significativa redução observada na eclosão dos ovos expostos ao vapor da substância.

  18. Effet comparé des huiles essentielles de quatre espèces végétales contre la bruche du niébé (Callosobruchus maculatus Fab. et le charançon du riz (Sitophilus oryzae L

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    Gakuru, S.


    Full Text Available Compared Effect of Four Plant Essential Oils against Cowpea Weevil Callosobruchus maculatus Fab. and Rice Weevil Sitophilus oryzae L. The aim of this paper is to compare the essential oil effects of Eucalyptus tereticornis Sm., Eucalyptus citriodora Hook, Citrus sinensis (L. Obs. and Ocimum basilicum L. against the cowpea weevil (Callosobruchus maculatus Fab. and the rice weevil (Sitophilus oryzae L. The experiments were conducted in laboratory, in Petri dishes. As nutritive support, we used cowpea and maize grains for respectively C. maculatus and S. oryzae. The Petri dish bottom had a filter paper impregnated with a desired essential oil quantity. The obtained results showed that the tested essential oils were not efficient against S. oryzae. On the other hand, the essential oils of E. citriodora and O. basilicum were very efficient against C. maculatus. The respective LD 50 was 1.26 ml and 1.49 ml in acetone dilution of 2 %.

  19. Fumigant Toxicity of Crushed Bulbs of Two Allium Species to Callosobruchus maculatus (Fabricius (Coleoptera: Bruchidae Toxicidad Fumigante para Callosobruchus maculatus (Fabriciu (Coleoptera: Bruchidae de Bulbos Trozados de Especies Allium

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    T.I Ofuya


    Full Text Available Fumigant toxicity of crushed fresh bulbs of Allium sativum L. and A. cepa L. to the Callosobruchus maculatus (Fabricius, a major pest of stored cowpea (Vigna unguiculata (L. Walp. seeds was assessed under laboratory conditions in Akure, Nigeria. In the tests, 20 g of infested cowpea seeds were suspended in a piece of muslin cloth, over an amount of crushed bulb in a container with a tightly fitted lid. Adult emergence was completely prevented from freshly laid eggs of C. maculatus on cowpea seeds that was fumigated with 6.0 g or more of crushed bulbs of A. sativum. Such fumigated seeds were not holed at all. Other amounts of A. sativum tested (1.0, 2.0, 3.0, 4.0 and 5.0 g significantly reduced C. maculatus adult emergence from fumigated eggs and seed holing in comparison with the control. Crushed A. sativum was ineffective in preventing adult emergence from fumigated C. maculatus larvae in seeds. The fumigant effect of crushed A. cepa did not kill all C. maculatus eggs. An amount of 7.0 g significantly reduced C. maculatus adult emergence from fumigated eggs and seed holing in comparison with the control. There is good prospect in using crushed bulbs of A. sativum> as fumigant in C. maculatus control in stored cowpea seeds.Se evaluó la toxicidad fumigante de bulbos frescos trozados de Allium sativum L. y A. cepa L. sobre Callosobruchus maculatus (Fabricius, una importante plaga de semilla almacenada de caupí Vigna unguiculata (L. Walp. bajo condiciones de laboratorio en Akure, Nigeria. En las pruebas, 20 g de semillas infestadas se suspendieron en un trozo de tela sobre cierta cantidad de bulbos trozados en un contenedor con una tapa ajustada. Se previno completamente la emergencia de adultos desde huevos recién puestos de C. maculatus en semillas de caupí que se fumigaron con 6,0 g o más de bulbos de A. sativum. Estas semillas fumigadas no estaban ahuecadas. Otras cantidades de A. sativum probadas (1.0, 2.0, 3.0, 4.0 y 5.0 g redujeron

  20. Sex pheromones of Callosobruchus subinnotatus and C. maculatus (Coleoptera: Bruchidae): congeneric responses and role of air movement. (United States)

    Mbata, G N; Shu, S; Ramaswamy, S B


    Females of Callosobruchus spp. are known to produce sex pheromones that attract males. These sex pheromones cannot be adopted for use in pest management without first investigating the responses of the males in the windless conditions of storage environments. Consequently, behavioural bioassays of Callosobruchus subinnotatus Pic males were conducted in an olfactometer in the absence of air-flow. Under these conditions males were found to be able to follow odour trails to the source. However, the latency period was longer in diffusional bioassays than for insects in a Y-tube olfactometer that provided directional wind cues. The highest percentage of males reached the pheromone source when components of the pheromones, (E)-3-methyl-2-heptenoic acid (E32A) and (Z)-3-methyl-2-heptenoic acid (Z32A), were formulated in a 50:50 or 25:75 ratio. Males of C. maculatus (Fabricius) responded to sex pheromone of C. subinnotatus, but males of C. subinnotatus did not respond to that of C. maculatus. The two sex pheromone components of C. subinnotatus are also constituents of C. maculatus sex pheromone. These two components may be potentially useful in monitoring the populations of both species in stored beans. It is postulated that (Z)-3-methyl-3-heptenoic acid (Z33A), the major component of the sex pheromone of C. maculatus, must have acted as an antagonist inhibiting response of C. subinnotatus to the sex pheromone of C. maculatus.

  1. Bioactivity of Powder and Extracts from Garlic, Allium sativum L. (Alliaceae and Spring Onion, Allium fistulosum L. (Alliaceae against Callosobruchus maculatus F. (Coleoptera: Bruchidae on Cowpea, Vigna unguiculata (L. Walp (Leguminosae Seeds

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    Abiodun A. Denloye


    Full Text Available Laboratory bioassays were conducted to investigate the bioactivity of powders, extracts, and essential oils from Allium sativum L. (Alliaceae and A. fistulosum L. (Liliaceae against adults, eggs, and larvae of Callosobruchus maculatus F. (Coleoptera: Bruchidae. On the basis of 48 hr median lethal toxicity (LC50, test plant powders and extracts from A. sativum were more toxic to C. maculatus adults than those from A. fistulosum. The 48 hr LC50 values for the powder against the test insect species were 9.66 g/kg and 26.29 g/kg for A. sativum and A. fistulosum, respectively. Also the 48 hr LC50 values obtained show that aqueous extracts of the test plant species, 0.11 g/L (A. sativum and 0.411 g/L (A. fistulosum were more toxic to C. maculatus than the corresponding ethanol extracts. There was no significant difference in the toxicity of vapours from the two test plant species against C. maculatus, although A. sativum gave lower values. The study shows that A. sativum and A. fistulosum have potentials for protecting stored cowpea from damage by C. maculatus.

  2. Effect of plant volatile oils in protecting stored cowpea Vigna unguiculata (L.) Walpers against Callosobruchus maculatus (F.) (Coleoptera: Bruchidae) infestation. (United States)

    Raja; Albert; Ignacimuthu; Dorn


    Adult Callosobruchus maculatus (F.) were introduced into cowpea seeds which were stored in containers with volatile oils derived from Mentha arvensis, M. piperata, M. spicata and Cymbopogon nardus. The numbers of eggs laid, adult mortality, adult emergence and subsequent seed damage were studied for four months. All oils significantly influenced all parameters (PM. piperata>M. arvensis>C. nardus.

  3. Efficacy of nanostructured silica as a stored pulse protector against the infestation of bruchid beetle, Callosobruchus maculatus (Coleoptera: Bruchidae) (United States)

    Arumugam, Ganesh; Velayutham, Veeramani; Shanmugavel, Sakthivelkumar; Sundaram, Janarthanan


    The treatment of hydrophobic silica nanoparticles (SNPs) with the pulse seeds of Cajanus cajan, Macrotyloma uniflorum, Vigna mungo, Vigna radiata, Cicer arietinum and Vigna unguiculata against the infestation of stored pulse beetle, Callosobruchus maculatus revealed a significant reduction in oviposition, adult emergence and seed damage potential. There was a complete retardation of growth of this beetle in the treated seeds of C. cajan. SNP-treated seeds of these six varieties of pulses revealed no effect on the growth of seeds as revealed by seed germination, growth rate of root and shoot. Similarly, the soil microflora measured in terms of colony forming units was not affected by silica nanoparticles upon its treatment with pulse seeds. The results of this study thus clearly demonstrated the useful nature of silica nanoparticles as seed protecting agent for the control of C. maculatus.

  4. Susceptibility of the bruchid Callosobruchus maculatus (Coleoptera: Bruchidae) and its parasitoid Dinarmus basalis (Hymenoptera: Pteromalidae) to three essential oils. (United States)

    Ketoh, Guillaume K; Glitho, Adole I; Huignard, Jacques


    The bruchid Callosobruchus maculatus (F.) causes major losses during the storage of seeds of Vigna unguiculata (Walp.) in West Africa. An endemic parasitoid, the pteromalid Dinarmus basalis (Rond.) reduces the increase in bruchid populations in stores and could be used for biological control. African farmers often introduce essential oils into granaries at harvest time. In Togo, essential oils were extracted from two Gramineae, Cymbopogon nardus (L.) and Cymbopogon schoenanthus (L.) and from a Lamiaceae, Ocimum basilicum (L.). The major components of these essential oils were citronellal in C. nardus, carene-2 and piperitone in C. schoenanthus and estragol in O. basilicum. Cymbopogon schoenanthus was the most toxic oil for C. maculatus adults. D. basalis adults were more susceptible to the three essential oils than the adults of their hosts C. maculatus. In the presence of cowpea seeds, the LC50s of the three essential oils were lower than in their absence, suggesting that the seeds may absorb a part of the volatiles. High doses of three essential oils slightly affected the survival of the fourth instar or the pupae of C. maculatus. This high survival was due to protection of larvae from volatiles by the surrounding seeds. The D. basalis were more affected by the oil volatiles than their hosts. Sub-lethal doses of essential oils reduced the duration of the adult life of both insect species and fecundity of the females. The differences in sensitivity of the host and its parasitoid could influence their population dynamics. The introduction of the essential oils into storage systems potentially could reduce density of parasitoid populations and increase seed losses.

  5. The seed coat of Phaseolus vulgaris interferes with the development of the cowpea weevil [Callosobruchus maculatus (F. (Coleoptera: Bruchidae

    Directory of Open Access Journals (Sweden)

    Luciana B. Silva


    Full Text Available We have confirmed here that the seeds of the common bean (Phaseolus vulgaris, L. do not support development of the bruchid Callosobruchus maculatus (F., a pest of cowpea [Vigna unguiculata (L. Walp] seeds. Analysis of the testa (seed coat of the bean suggested that neither thickness nor the levels of compounds such as tannic acid, tannins, or HCN are important for the resistance. On the other hand, we have found that phaseolin (vicilin-like 7S storage globulin, detected in the testa by Western blotting and N-terminal amino acid sequencing, is detrimental to the development of C. maculatus. As for the case of other previously studied legume seeds (Canavalia ensiformis and Phaseolus lunatus we suggest that the presence of vicilin-like proteins in the testa of P. vulgaris may have had a significant role in the evolutionary adaptation of bruchids to the seeds of leguminous plants.Neste trabalho confirmamos a resistência do feijão comum ( Phaseolus vulgaris ao bruquídeo Callosobruchus maculatus (F., inseto que preda sementes de feijão-de-corda (Vigna unguiculata (L.. A resistência desta semente não está relacionada com a espessura do tegumento nem com os níveis de diversos compostos como ácido taníco, fenóis e ácido cianídrico conforme demonstram nossos resultados. No entanto, faseolina, detectada no tegumento por ‘‘Western blotting’’ e seqüenciamento N-terminal, é tóxica a C. maculatus. Esses dados estão de acordo com estudos anteriores feitos com duas outras sementes de leguminosas (Canavalia ensiformis e Phaseolus lunatus e nos levam a sugerir que a presença de proteínas do tipo vicilina no tegumento de sementes de leguminosas tiveram papel importante nos mecanismo de adaptação de bruquídeos a sementes de leguminosas.

  6. Efficacy of Plectranthus glandulosus (Lamiaceae) and Callistemon rigidus (Myrtaceae) Leaf Extract Fractions to Callosobruchus maculatus (Coleoptera: Bruchidae). (United States)

    Danga, Simon Pierre Yinyang; Nukenine, Elias Nchiwan; Younoussa, Lame; Adler, Cornel; Esimone, Charles Okechukwu


    As part of on-going efforts to use eco-friendly alternatives to chemical pesticides, methanol crude extracts of Plectranthus glandulosus and Callistemon rigidus leaves were sequentially fractionated in hexane, chloroform, ethyl acetate, and methanol to establish the most active fraction(s) against Callosobruchus maculatus in cowpea. Cowpea seeds (25 g) were treated with 0.5, 1, 2, and 4 g/kg of extract to evaluate the contact toxicity and F1 progeny production of the beetles in the laboratory. Mortality was recorded 1, 3, and 7 d postexposure. P. glandulosus hexane fraction was more toxic than the other fractions recording 100% mortality at 4 g/kg, within 7 d with LC50 of 0.39 g/kg. Hexane fraction of C. rigidus showed superior toxicity, causing 100% mortality at 4 g/kg within only 1 d of exposure with LC50 of 1.02 g/kg. All the fractions greatly reduced progeny emergence, with C. rigidus hexane fraction being the best progeny inhibitor. Fractions of P. glandulosus and C. rigidus leaves had sufficient efficacy to be a component of storage pest management package for C. maculatus.

  7. Global proteome changes in larvae of Callosobruchus maculatus Coleoptera:Chrysomelidae:Bruchinae) following ingestion of a cysteine proteinase inhibitor

    DEFF Research Database (Denmark)

    Nogueira, Fábio C S; Silva, Carlos P; Alexandre, Daniel


    proteomic changes induced in the intestinal tract of larval C. maculatus challenged by the ingestion of cystatin, a cysteine peptidase inhibitor, was investigated by a nanoLC-MS/MS approach. The ingestion of cystatin caused a delay in the development of the larvae, but the mortality was not high, indicating....... Ingestion of cystatin led to significant changes in the proteome of both the midgut epithelia and midgut contents. We have observed that proteins related to plant cell wall degradation, particularly the key glycoside hydrolases of the families GH5 (endo-β-1,4-mannanase) and GH 28 (polygalacturonase) were...... overexpressed. Conversely, α-amylases were downexpressed, indicating that an increase in hemicelluloses digestion helps the larvae to cope with the challenge of cystatin ingestion. Furthermore, a number of proteins associated with transcription/translation and antistress reactions were among the cystatin...

  8. Laboratory evaluation of the toxic properties of forest anchomanes, Anchomanes difformis against pulse beetle Callosobruchus maculatus (Coleoptera: Bruchidae)

    Institute of Scientific and Technical Information of China (English)



    Laboratory experiments were carried out to investigate the efficacy of crude stem extracts of forest anchomanes, Anchomanes difformis (P. Beauv.) a plant occurring in West African forests, against the pulse beetle Callosobruchus maculatus (Fabricius). Crude stem extracts at 3% concentration showed high contact toxicity to adult beetles within 24 h after application, while it was moderately toxic to the beetles at the lowest (1%) concentration. At the highest application rate, the plant extract provided good protection to grains stored for 90 days. Grain viability and water absorption capacity were not affected by treatments with ethanol extracts ofA. difformis. The significance of these findings is discussed in relation to biopesticide-means of controlling cowpea bruchids.

  9. Comparaison de quelques caractéristiques biologiques entre Dinarmus basalis Rond. (Hymenoptera: Pteromalidae élevé soit sur son hôte habituel Callosobruchus maculatus F. (Coleoptera: Bruchidae soit sur Acanthoscelides macrophthalmus Schaef. ou Bruchidius lineatopygus Pic. identifiés comme hôtes de substitution

    Directory of Open Access Journals (Sweden)

    Nuto, Y.


    Full Text Available Comparison of some Biological Characteristics between Dinarmus basalis Rond. (Hymenoptera: Pteromalidae Reared either on his Habitual Host Callosobruchus maculatus F. (Coleoptera: Bruchidae or on Acanthoscelides macrophthalmus Schaef. and Bruchidius lineatopygus Pic. Identified as Substitution Hosts. In this study, we have measured some biological parameters of Dinarmus basalis Rond. (Hymenoptera: Pteromalidae when this parasitoid grows on three hosts: Callosobruchus maculatus F. (usual host, Acanthoscelides macrophthalma Schaef. and Bruchidius lineatopygus Pic. (substitution hosts. The results of this comparative study in no choice situation show that, life duration and development duration of this parasitoid are identical in laboratory conditions. By contrast, the female fecundity, the sexual rate and the parasitism rate of D. basalis are different according to the hosts presented. Although such differences appear to be small between C. maculatus and A. macrophthalmus. In presence of these two species, the number of eggs laid by the female of D. basalis is respectively 61 ± 5.24 and 54 ± 5.60, the sexual rate 37.63 ± 2.13% and 41.73 ± 3.69% and the parasitism rate 85.46 ± 2.61% and 76.48 ± 5.90%. This indicates that these two Bruchids can be used as hosts to D. basalis for a mass production. In presence of B. lineatopygus, the fecundity and the parasitism rate of D. basalis female are very low and the sexual rate more favourable of the males. This situation is unfavourable for a parasitoid production. When D. basalis females are placed in choice situation with equal number of the hosts nowely C. maculatus and A. macrophthalmus, the parasitism rate obtained, is higher for C. maculatus that of A. macrophthalmus. This parasitism rate remains high for C. maculatus even when the other host A. macrophthalmus is twice the number of C. maculatus. Thus the parasitoid seems to have a preference for his natural host. However, A. macrophthalmus can

  10. First report of toxicity of Xylopiaparviflora (A. Rich.) Benth (Annonaceae) root bark's essential oil against cowpea seed bruchid, Callososbruchus maculatus Fabricius (Coleoptera: Chrysomelidae: Bruchinae). (United States)

    Babarinde, Samuel Adelani; Pitan, Olufemi Olutoyin Richard; Olatunde, Ganiyu Olatunji; Ajala, Michael Oluwole


    The fumigant toxicity of Xylopia parviflora (A. Rich.) Benth (Annonaceae) root bark's essential oil (EO) against cowpea seed bruchid, Callosobruchus maculatus, was investigated in the laboratory. Dose had significant (P < 0.0001) effect on mortality at 6 hours after treatment (HAT) at a concentration of 6.25 μL/mL air which exerted 81.70% mortality, while there was no mortality in all other lower doses. At 12 HAT, 75.05% and 90.00% mortality were observed at doses of 3.15 and 6.25 μL/mL air, respectively. It was significantly (P < 0.05) higher than the mortality (50.58%) observed when 0.78 μL/mL air was applied. The lethal time for 50% of assayed adults (LT50) obtained when the bruchid was exposed to X. parviflora EO at a dose of 6.25 μL/mL air (2.71 h) was significantly lower than LT50 obtained at exposure of bruchid to other lower doses of 0.78-3.15 μL/mL air.

  11. Influência do período de armazenamento do caupi [Vigna unguiculata (L. Walp.], tratado com óleos essenciais e fixos, no controle de Callosobruchus maculatus (Fabricius, 1775 (Coleoptera, Chrysomelidae, Bruchinae Influence of the storage period of cowpea [Vigna unguiculata (L. Walp.] treated with essential and fixed oils, for the control of Callosobruchus maculatus (Fabricius, 1775 (Coleoptera, Chrysomelidae, Bruchinae

    Directory of Open Access Journals (Sweden)

    Adriana Carla Ribeiro Lopes Pereira


    Full Text Available Compostos secundários obtidos de plantas podem ser utilizados no controle de Callosobruchus maculatus, como uma tática alternativa potencial aos inseticidas sintéticos. Foram testados óleos essenciais (Cymbopogon martini Roxb., Piper aduncum L., Piper hispidinervum C.DC., Melaleuca sp., Lippia gracillis Shau e fixos (Helianthus annus L., Sesamum indicum L., Gossypium hirsutum L., Glycine max L. e Caryocar brasiliense Camb., na concentração de 50µl/20g, de acordo com estudos anteriores. Grãos de caupi, cv. Sempre Verde, foram impregnados com os óleos, em recipientes de vidro e submetidos à agitação manual por dois minutos. Cada parcela de 20g foi infestada com oito fêmeas de C. maculatus com 0 a 48h de idade, durante quatro dias. Os óleos foram avaliados logo após a impregnação e aos 30, 60, 90 e 120 dias de armazenamento. Na primeira avaliação, todos os óleos essenciais provocaram 100% de mortalidade e para os óleos fixos, a mortalidade variou entre 35% (G. hirsutum e 67,5% (G. max. Com o prolongamento do período de armazenamento, houve um aumento do número de ovos viáveis e de insetos emergidos, exceto para P. aduncum. Em relação aos óleos fixos, S. indicum, G. max, G. hirsutum e C. brasiliense foram os mais eficientes até os 30 dias de armazenamento. Os resultados indicam que os óleos testados na concentração de 50µl/20g apresentam baixo efeito residual, com exceção de P. aduncum, que foi efetivo durante todo o período de armazenamento.The secondary compounds extracted from plants are considered potential alternative to synthetic insecticides in the control of agricultural pests. Essential oils (Cymbopogon martini Roxb., Piper aduncum L., P. hispidinervum C.DC., Melaleuca sp. and Lippia gracillis Shau and fixed oils (Helianthus annus L., Sesamum indicum L., Gossypium hirsutum L., Glycine max L. and Caryocar brasiliense Camb. at the concentration of 50µl/20g were tested according to previous studies. Samples

  12. 三种植物粉剂对四纹豆象(鞘翅目:豆象科)行为、死亡率和繁殖的影响%Effects of Three Plant Powders on Behaviour, Mortality and Reproductive Fitness of Callosobruchus maculatus (Fabricius) (Coleoptera: Bruchidae)

    Institute of Scientific and Technical Information of China (English)



    The effects of powders from dry flower buds of Eugenia aromatica Baill., seeds of Piper guineense Schum and Thonn and fruits of Capsicum frutescens L. on adult behaviour, mortality and reproductive fitness of the cowpea seed beetle, Callosobruchus maculatus (Fab.) were investigated under ambient laboratory conditions. All experiments were carried out in glass Petri plates. All powders elicited aversion in adult beetles. The contact toxicity symptoms included restlessness, loss of coordination, knock down and eventual death of adult beetles. These behaviours were more pronounced with E. aromatica in which adult beetles died within 16 hours. P. guineense and C. frutescens powders did not cause 100% mortality of adult beetles, even after 24 hours. Each of the three powders significantly (P<0.05) reduced the mating competition of adult males after sub-lethal exposure for one, two, and three hours, respectively. E. aromatica powder caused more reduction in male mating competition for females than any of the other two powders after each period of exposure. Receptiveness of treated females to courting males was also decreased by exposure to any of the three powders. Similarly, E. aromatica powder caused greater decreases in female receptiveness to males than any of the other two powders after each period of exposure. Exposure of either adult male or female C. maculatus to the powders for sub-lethal times of three, six and nine hours significantly reduced the fecundity of the beetles. E. aromatica powder caused the most reduction of eggs laid and also significantly reduced fertility of the eggs.

  13. Atividade inseticida de óleos essenciais e fixos sobre Callosobruchus maculatus (FABR., 1775 (Coleoptera: Bruchidae em grãos de caupi [Vigna unguiculata (L. WALP.] Insecticide activity of essential and fixed oils in Callosobruchus maculatus (FABR., 1775 (Coleoptera: Bruchidae in cowpea grains [Vigna unguiculata (L. WALP.

    Directory of Open Access Journals (Sweden)

    Adriana Carla Ribeiro Lopes Pereira


    Full Text Available O caruncho, Callosobruchus maculatus (Fabr., é considerado a praga mais importante do caupi, Vigna unguiculata (L. Walp., armazenado em regiões tropicais e subtropicais. Visando minimizar os efeitos indesejáveis dos inseticidas químicos sintéticos, o controle dessa praga com óleos de origem vegetal vem se constituindo numa alternativa promissora, de baixo custo e segura para os aplicadores e consumidores. Foram testados os óleos essenciais [(Cymbopogon martini (Roxb. J.F. Watson], Piper aduncum L., Piper hispidinervum C.DC., Melaleuca sp., Lippia gracillis Shauer e fixos (Helianthus annuus L, Sesamum indicum L, Gossypium hirsutum L., Glycine max (L. Merr. e Caryocar brasiliense Camb., em grãos de caupi, cv. Sempre Verde. Os óleos foram utilizados nas concentrações 10, 20, 30, 40 e 50mL/20g, correspondendo a 0,5, 1,0, 1,5, 2,0 e 2,5 L/t e impregnados aos grãos no interior de recipientes de plástico, com auxílio de pipetador automático e agitados manualmente durante dois minutos. Parcelas de 20g de caupi foram infestadas com oito fêmeas de C. maculatus, com 0 a 48 h de idade. Cada óleo foi testado, separadamente, em delineamento inteiramente casualisado com seis repetições. Os óleos essenciais de C. martini, P. aduncum e L. gracillis causaram 100% de mortalidade em todas as concentrações, P. hispidinervum a partir de 1,5 L/t e Melaleuca sp. nas concentrações de 2,0 e 2,5 L/t. A redução do número de ovos viáveis e de insetos emergidos foi de 100% para todos os óleos essenciais, exceto Melaleuca sp. Por outro lado, os óleos fixos, apesar de apresentarem baixa mortalidade em todas as concentrações testadas, reduziram em praticamente 100% o número de ovos viáveis e de insetos emergidos.The weevil Callosobruchus maculatus (Fabr. is considered the most important pest of cowpea, Vigna unguiculata (L. Walp., stored in tropical and subtropical regions. Focusing to minimize the undesired effects of synthetic chemical

  14. Repellent activity of some essential oils against two stored product beetles Callosobruchus chinensis L. and C. maculatus F. (Coleoptera: Bruchidae) with reference to Chenopodium ambrosioides L. oil for the safety of pigeon pea seeds. (United States)

    Pandey, Abhay K; Palni, Uma T; Tripathi, N N


    Essential oils from 35 aromatic and medicinal plant species of Gorakhpur Division (U. P., India) were evaluated for their repellent activity against pulse bruchids Callosobruchus chinensis L. and C. maculatus F. of stored pigeon pea seeds. The oil concentration was at 0.36 μl/ml. Out of 35 essential oils, Adhatoda vasica Ness and Chenopodium ambrosioides L. oils showed absolute (100 %) insect repellency. Chenopodium oil exhibited 100 % mortality for both the test insects at 10 μl concentration (LD50 = 2.8 μl for C. chinensis & 2.5 μl for C. maculatus) and more toxic than Adhatoda oil (LD50 = 6.8 μl for C. chinensis & 8.4 μl for C. maculatus). During in vivo evaluation, 0.29 and 0.58 μl/ml of Chenopodium oil significantly enhanced feeding deterrence in insects and reduced the seed damage as well as weight loss of fumigated pigeon pea seeds up to 6 months of storage as compared to control set. Thus, Chenopodium oil can be used as an effective option of commercial fumigants for the storage of pigeon pea seeds against pulse bruchids.

  15. Efficacy of entomopathogenic fungi in combination with diatomaceous earth against Callosobruchus maculatus(Coleoptera: Bruchidae)%昆虫病原真菌与硅藻土混用对四纹豆象的毒力效果

    Institute of Scientific and Technical Information of China (English)

    Neda NABAEI; Ali MEHRVAR; Moosa SABER; Mohammad BAGHERI


    在室内条件下检测了白僵菌Beauveria bassiana (Balsamo) Vuillemin菌株IRAN-187C和金龟子绿僵菌Metarhizium anisopliae (Metschnikow) Sorokin菌株A-396分别与硅藻土(DE)制剂InsectoSec(R)混用对四纹豆象Callosobruchus maculatus成虫的毒力效果.结果表明:硅藻土与致死剂量和亚致死剂量的真菌分离物混用导致了四纹豆象成虫的高死亡率,且这些处理均缩短了真菌的致死中时(LT50).LC50的白僵菌B.bassiana菌株IRAN-187C和LC50的DE混用对四纹豆象的LT50 (122.2 h)最低,在95%置信限上与LC50的金龟子绿僵菌M.anisopliae菌株A-396与LC50的DE混用得到的LT50(128.2 h)相当,说明两种真菌分离物与DE混用对四纹豆象成虫都具有较好的毒力效果.用四纹豆象成虫检测3个温度(22,27和30℃)和2个相对湿度(40%和55%)组合对DE活性的影响,结果显示30℃和40%相对湿度条件下得到的LT50(110.11 h)最低,说明高温低湿条件下DE活性较高.%Experiments were conducted to enhance the virulence of two isolates of entomopathogenic fungi,IRAN-187C of Beauveria bassiana (Balsamo) Vuillemin, and A-396 isolate of Metarhizium anisopliae (Metschnikow) Sorokin,by combination with InsectoSec(R) formulation of diatomaceous earth (DE) against the adults of Callosobruchus maculatus F.under laboratory conditions.The results demonstrated that the combinations between diatomaceous earth and the fungal isolates at lethal and sub-lethal doses achieved high mortality rates in the adult populations ; moreover,the treatments improved LT50 values of the fungi.The lowest value of LT50 (122.2 h) was achieved by LC50 (B.bassiana,IRAN-187C) + LC50 (DE) treatment which was on par with that of LC50 (M.anisopliae,A-396) + LC50 (DE) combination (128.2 h) based on their 95% fiducial limit values,resulting in better efficacy of each fungal isolate in combination with diatomaceous earth against C.maculatus adults.Also,effects of the three temperatures (22,27,and 30

  16. Beyond Fab Four (United States)

    Babichev, E.; Charmousis, C.; Langlois, D.; Saito, R.


    We show that the two additional Lagrangians that appear in theories beyond Horndeski can be reexpressed in terms of simple generalizations of the 'John' and 'Paul' terms of the Fab Four theories. We find that these extended Fab Four satisfy the same properties of self-tuning as the original Fab Four.

  17. Beyond Fab Four

    CERN Document Server

    Babichev, E; Langlois, D; Saito, R


    We show that the two additional Lagrangians that appear in theories beyond Horndeski can be reexpressed in terms of simple generalizations of the "John" and "Paul" terms of the Fab Four theories. We find that these extended Fab Four satisfy the same properties of self-tuning as the original Fab Four.

  18. Preliminary results on evaluation of chickpea, Cicer arietinum, genotypes for resistance to the pulse beetle, Callosobruchus maculatus. (United States)

    Erle, F; Ceylan, F; Erdemir, T; Toker, C


    Abstract The chickpea, Cicer arietinum L. (Fabales: Fabaceae), seeds are vulnerable, both in the field and in storage, to attack by seed-beetles. Beetles of the genus Callosobruchus are major storage pests of chickpea crops and cause considerable economic losses. In the present study, a total of 11 chickpea genotypes including five 'kabuli' (Mexican white, Diyar, CA 2969, ILC 8617 and ACC 245) and six 'desi' chickpeas (ICC 1069, ICC 12422, ICC 14336, ICC 4957, ICC 4969 and ICC 7509) were evaluated for resistance to the pulse beetle Callosobruchus maculatus F. (Coleoptera: Bruchidae). Resistance was evaluated by measuring percent damage to seeds. Damage to seeds by C. maculatus was manifested by the round exit holes with the 'flap' of seed coat made by emerging adults. Of the 11 genotypes tested, only one (ICC 4969) exhibited a complete resistance to C. maculatus in both free-choice and no-choice tests; no seed damage was found over the test period. In general, the 'desi' chickpeas were more resistant to C. maculatus than the 'kabuli' chickpeas. Among the tested chickpea genotypes, only ICC 4969 can be used as a source of C. maculatus resistance in breeding programmes that could then be grown in organic cultivation free from pesticides.

  19. Single chain Fab (scFab fragment

    Directory of Open Access Journals (Sweden)

    Brenneis Mariam


    Full Text Available Abstract Background The connection of the variable part of the heavy chain (VH and and the variable part of the light chain (VL by a peptide linker to form a consecutive polypeptide chain (single chain antibody, scFv was a breakthrough for the functional production of antibody fragments in Escherichia coli. Being double the size of fragment variable (Fv fragments and requiring assembly of two independent polypeptide chains, functional Fab fragments are usually produced with significantly lower yields in E. coli. An antibody design combining stability and assay compatibility of the fragment antigen binding (Fab with high level bacterial expression of single chain Fv fragments would be desirable. The desired antibody fragment should be both suitable for expression as soluble antibody in E. coli and antibody phage display. Results Here, we demonstrate that the introduction of a polypeptide linker between the fragment difficult (Fd and the light chain (LC, resulting in the formation of a single chain Fab fragment (scFab, can lead to improved production of functional molecules. We tested the impact of various linker designs and modifications of the constant regions on both phage display efficiency and the yield of soluble antibody fragments. A scFab variant without cysteins (scFabΔC connecting the constant part 1 of the heavy chain (CH1 and the constant part of the light chain (CL were best suited for phage display and production of soluble antibody fragments. Beside the expression system E. coli, the new antibody format was also expressed in Pichia pastoris. Monovalent and divalent fragments (DiFabodies as well as multimers were characterised. Conclusion A new antibody design offers the generation of bivalent Fab derivates for antibody phage display and production of soluble antibody fragments. This antibody format is of particular value for high throughput proteome binder generation projects, due to the avidity effect and the possible use of

  20. FabIO

    DEFF Research Database (Denmark)

    Bergbäck Knudsen, Erik; Sørensen, Henning O.; Wright, Jonathan P.


    FabIO is a Python module written for easy and transparent reading of raw two-dimensional data from various X-ray detectors. The module provides a function for reading any image and returning a fabioimage object which contains both metadata (header information) and the raw data. All fabioimage...

  1. Fab Four Neutron Stars

    CERN Document Server

    Maselli, Andrea; Minamitsuji, Masato; Berti, Emanuele


    Horndeski's theory of gravity is the most general scalar-tensor theory with a single scalar whose equations of motion contain at most second-order derivatives. A subsector of Horndeski's theory known as "Fab Four" gravity allows for dynamical self-tuning of the quantum vacuum energy, and therefore it has received particular attention in cosmology as a possible alternative to the $\\Lambda$CDM model. Here we study compact stars in Fab Four gravity, which includes as special cases general relativity ("George"), Einstein-dilaton-Gauss-Bonnet gravity ("Ringo"), theories with a nonminimal coupling with the Einstein tensor ("John") and theories involving the double-dual of the Riemann tensor ("Paul"). We generalize and extend previous results in theories of the John class and we show that there are no viable compact star solutions in theories of the Paul class.

  2. Larval competition reduces body condition in the female seed beetle, Callosobruchus maculatus. (United States)

    Schade, Daynika J; Vamosi, Steven M


    Early body condition may be important for adult behavior and fitness, and is impacted by a number of environmental conditions and biotic interactions. Reduced fecundity of adult females exposed to larval competition may be caused by reduced body condition or shifts in relative body composition, yet these mechanisms have not been well researched. Here, body mass, body size, scaled body mass index, and two body components (water content and lean dry mass) of adult Callosobruchus maculatus (Fabricius) (Coleoptera: Chrysomelidae: Bruchinae) females exposed to larval competition or reared alone were examined. Experimental females emerged at significantly smaller body mass and body size than control females. Additionally, scaled body mass index and water content, but not lean dry mass, were significantly reduced in experimental females. To our knowledge, these are the first results that demonstrate a potential mechanism for previously documented direct effects of competition on fecundity in female bruchine beetles.

  3. Chemical Composition and Insecticidal Activity of Essential Oil from Coriandrum sativum Seeds against Tribolium confusum and Callosobruchus maculatus. (United States)

    Khani, Abbas; Rahdari, Tahere


    The biological activity of essential oil extracted from coriander, Coriandrum sativum L. (Apiaceae), seeds against adults of Tribolium confusum Duval (Coleoptera: Tenebrionidae) and Callosobruchus maculatus F. (Coleoptera: Bruchidae) was investigated in a series of laboratory experiments. Fumigant toxicity was assessed at 27 ± 1°C and 65 ± 5% R.H., in dark condition. Dry seeds of the plant were subject to hydrodistillation using a Clevenger-type apparatus. The composition of essential oil was analyzed by gas chromatography mass spectrometry. The predominant components in the oil were linalool (57.57%) and geranyl acetate (15.09%). The mortality of 1-7-day-old adults of the insect pests increased with concentration from 43 to 357 μL/L air and with exposure time from 3 to 24 h. In the probit analysis, LC(50) values (lethal concentration for 50% mortality) showed that C. maculatus (LC(50) = 1.34 μL/L air) was more susceptible than T. confusum (LC(50) = 318.02 μL/L air) to seed essential oil of this plant. The essential oil of C. sativum can play an important role in stored grain protection and reduce the risks associated with the use of synthetic insecticides.

  4. The De Novo Transcriptome and Its Functional Annotation in the Seed Beetle Callosobruchus maculatus.

    Directory of Open Access Journals (Sweden)

    Ahmed Sayadi

    Full Text Available Despite their unparalleled biodiversity, the genomic resources available for beetles (Coleoptera remain relatively scarce. We present an integrative and high quality annotated transcriptome of the beetle Callosobruchus maculatus, an important and cosmopolitan agricultural pest as well as an emerging model species in ecology and evolutionary biology. Using Illumina sequencing technology, we sequenced 492 million read pairs generated from 51 samples of different developmental stages (larvae, pupae and adults of C. maculatus. Reads were de novo assembled using the Trinity software, into a single combined assembly as well as into three separate assemblies based on data from the different developmental stages. The combined assembly generated 218,192 transcripts and 145,883 putative genes. Putative genes were annotated with the Blast2GO software and the Trinotate pipeline. In total, 33,216 putative genes were successfully annotated using Blastx against the Nr (non-redundant database and 13,382 were assigned to 34,100 Gene Ontology (GO terms. We classified 5,475 putative genes into Clusters of Orthologous Groups (COG and 116 metabolic pathways maps were predicted based on the annotation. Our analyses suggested that the transcriptional specificity increases with ontogeny. For example, out of 33,216 annotated putative genes, 51 were only expressed in larvae, 63 only in pupae and 171 only in adults. Our study illustrates the importance of including samples from several developmental stages when the aim is to provide an integrative and high quality annotated transcriptome. Our results will represent an invaluable resource for those working with the ecology, evolution and pest control of C. maculatus, as well for comparative studies of the transcriptomics and genomics of beetles more generally.

  5. Deep, Staged Transcriptomic Resources for the Novel Coleopteran Models Atrachya menetriesi and Callosobruchus maculatus. (United States)

    Benton, Matthew A; Kenny, Nathan J; Conrads, Kai H; Roth, Siegfried; Lynch, Jeremy A


    Despite recent efforts to sample broadly across metazoan and insect diversity, current sequence resources in the Coleoptera do not adequately describe the diversity of the clade. Here we present deep, staged transcriptomic data for two coleopteran species, Atrachya menetriesi (Faldermann 1835) and Callosobruchus maculatus (Fabricius 1775). Our sampling covered key stages in ovary and early embryonic development in each species. We utilized this data to build combined assemblies for each species which were then analysed in detail. The combined A. menetriesi assembly consists of 228,096 contigs with an N50 of 1,598 bp, while the combined C. maculatus assembly consists of 128,837 contigs with an N50 of 2,263 bp. For these assemblies, 34.6% and 32.4% of contigs were identified using Blast2GO, and 97% and 98.3% of the BUSCO set of metazoan orthologs were present, respectively. We also carried out manual annotation of developmental signalling pathways and found that nearly all expected genes were present in each transcriptome. Our analyses show that both transcriptomes are of high quality. Lastly, we performed read mapping utilising our timed, stage specific RNA samples to identify differentially expressed contigs. The resources presented here will provide a firm basis for a variety of experimentation, both in developmental biology and in comparative genomic studies.

  6. Fab Chaperone-Assisted RNA Crystallography (Fab CARC). (United States)

    Sherman, Eileen; Archer, Jennifer; Ye, Jing-Dong


    Recent discovery of structured RNAs such as ribozymes and riboswitches shows that there is still much to learn about the structure and function of RNAs. Knowledge learned can be employed in both biochemical research and clinical applications. X-ray crystallography gives unparalleled atomic-level structural detail from which functional inferences can be deduced. However, the difficulty in obtaining high-quality crystals and their phasing information make it a very challenging task. RNA crystallography is particularly arduous due to several factors such as RNA's paucity of surface chemical diversity, lability, repetitive anionic backbone, and flexibility, all of which are counterproductive to crystal packing. Here we describe Fab chaperone assisted RNA crystallography (CARC), a systematic technique to increase RNA crystallography success by facilitating crystal packing as well as expediting phase determination through molecular replacement of conserved Fab domains. Major steps described in this chapter include selection of a synthetic Fab library displayed on M13 phage against a structured RNA crystallization target, ELISA for initial choice of binding Fabs, Fab expression followed by protein A affinity then cation exchange chromatography purification, final choice of Fab by binding specificity and affinity as determined by a dot blot assay, and lastly gel filtration purification of a large quantity of chosen Fabs for crystallization.

  7. EFFECT OF INFESTATION OF Dermestes maculatus on the ...

    African Journals Online (AJOL)

    Olusola Fasunwon


    Dec 7, 2011 ... assessment was based on weight and proximate analysis differences ... populations of matured larvae have damaged wooden structural .... The Dermestes maculatus (hide beetles) were picked with a soft (moist) camel hair.

  8. Evaluation of the effectiveness of conventional insecticides against the cowpea weevil, Callosobruchus maculatus (Coleoptera: Bruchidae), on four different substrata surfaces%四种基质表面上常规杀虫剂对四纹豆象的药效评价

    Institute of Scientific and Technical Information of China (English)

    Vahid MAHDAVI; Moosa SABER; Samad VOJOUDI


    In pest management programs,the control of pests in structural facilities or sites,where processed food products are consumed or stored,requires the application of insecticides upon a variety of substrate surfaces. In order to determine the insecticidal effectiveness on different substrate surfaces,adults of the cowpea weevil,Callosobruchus maculatus ( F.),were exposed to field recommended concentrations of chlorpyrifos, abamectin and deltamethrin insecticides. The results showed that abamectin caused mortality rates of 63.33%,22.41%,12.9% and 11.9% while deltamethrin caused mortality rates of 55%,44.2%,41.3% and 37.4% on glass,ceramic tile,plastic and paper disc surfaces,respectively.Exposures to chlorpyrifos led to 100% mortality in all surfaces.Probit analysis of data showed that LC50 values were 8.66,13.6,29.16 and 56.5 μg/mL for chlorpyrifos,119.4,446.2,774.2 and 836.4 μg/mL for abamectin,and 1008,1131,1210 and 1336 μg/mL for deltamethrin on the glass,ceramic tile,plastic and paper disc surfaces,respectively,based on formulated materials.It is concluded that chlorpyrifos is the most toxic insecticide to the cowpea weevil,but that its toxicity was reduced in the glass,ceramic tile,plastic and paper disc surfaces in sequence.%在害虫治理中,在消费或贮藏粮食加工产品的建筑设施或场所进行害虫防治需要将杀虫剂施用在各种基质表面上.为了测定不同基质表面上杀虫剂的药效,将四纹豆象Callosobruchus maculatus(F.)成虫接触田间推荐剂量的阿维菌素、溴氰菊酯和毒死蜱.结果表明:施用在玻璃、瓷砖、塑料和纸盘表面上,阿维菌素对四纹豆象成虫的致死率分别为63.33%,22.41%,12.9%和11.9%,而溴氰菊酯在这4种基质表面上对四纹豆象成虫的致死率分别为55%,44.2%,41.3%和37.4%.在所有基质表面上接触毒死蜱,四纹豆象成虫的死亡率均为100%.对数据进行的Probit分析表明,毒死

  9. "Fab 13": The Learning Factory. (United States)

    Crooks, Steven M.; Eucker, Tom R.


    Describes how situated learning theory was employed in the design of Fab 13, a four-day simulation-based learning experience for manufacturing professionals at Intel Corporation. Presents a conceptual framework for understanding situated learning and discusses context, content, anchored instruction, facilitation, scaffolding, collaborating,…

  10. "Fab 13": The Learning Factory. (United States)

    Crooks, Steven M.; Eucker, Tom R.


    Describes how situated learning theory was employed in the design of Fab 13, a four-day simulation-based learning experience for manufacturing professionals at Intel Corporation. Presents a conceptual framework for understanding situated learning and discusses context, content, anchored instruction, facilitation, scaffolding, collaborating,…


    Directory of Open Access Journals (Sweden)

    Umi Widyastuti


    Full Text Available Abstrak Malaria masih merupakan masalah kesehatan di Kabupaten Kulonprogo, khususnya di Kecamatan Kokap. Dalam dua tahun terakhir menunjukkan bahwa pada tahun 2009, 1 dari 5 desa di Kecamatan Kokap sebagai daerah malaria dengan statifikasi Insidensi Kasus Rendah (Low Case Incidence/LCI. Pada tahun 2010 meningkat menjadi 3 desa, peningkatan kasus malaria di Kecamatan Kokap berkaitan dengan keberadaan nyamuk Anopheles yang berpotensi sebagai vektor. Beberapa spesies seperti An. maculatus, An. aconitus, dan An. balabacensis diduga sebagai vektor malaria potensial di daerah ini. Penelitian ini bertujuan untuk mengetahui kompetensi vektorial An. maculatus yang terdiri dari: kerentanan nyamuk An. maculatus terhadap Plasmodium, sifat antropofilik, angka paritas, dan kepadatan nyamuk. Kerentanan nyamuk terhadap Plasmodium diukur dengan deteksi antigen protein sporozoit (Circum Sporozoite Protein/CSP dari P. falciparum atau P. vivax pada dada-kepala dari semua nyamuk parous. Karakteristik antropofilik diukur dengan mendeteksi darah manusia pada perut nyamuk dengan kondisi penuh darah dan setengah gravid. Kedua deteksi tersebut dilakukan dengan teknik ELISA. Penelitian dilakukan pada bulan Mei sampai Oktober 2011 di dua desa yaitu: Tegiri dan Gunungrego, Kecamatan Kokap. Nyamuk ditangkap dengan menggunakan metode sesuai dengan pedoman WHO. Penangkapan dilakukan pada malam hari (18.00-06.00 terhadap nyamuk yang hinggap pada manusia di dalam rumah, di luar rumah, nyamuk yang istirahat baik di dalam rumah (dinding maupun luar rumah (sekitar kandang ternak dan penangkapan pada pagi hari (06.00-08.00. Kepadatan An. maculatus dihitung dan paritas nyamuk ditentukan dengan pembedahan ovarium secara mikroskopis. Hasil penelitian menunjukkan bahwa An. maculatus di Gunungrego rentan terhadap P. vivax dengan angka sporozoit 3,57%. Namun, CSP antigen P. vivax tidak terdeteksi pada dada-kepala nyamuk An. maculatus dari Tegiri. Antigen CSP P. falciparum negatif pada

  12. Nuclear DNA content in Galaxias maculatus (Teleostei: Osmeriformes: Galaxiidae Contenido de ADN nuclear en Galaxias maculatus (Teleostei: Osmeriformes: Galaxiidae

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    Pedro Jara-Seguel


    Full Text Available The nuclear DNA content (2C value was determined in the commercial fish Galaxias maculatus (Galaxiidae was determined by microdensitometry of erythrocyte nuclei after Feulgen staining; rainbow trout erythrocytes with a known 2C value were used as a standard. The 2C value of G. maculatus was 2.21 ± 0.12 pg and its C value was equivalent to 1.105 pg (1,082.9 Mbp. This C value is within the range recorded for other osmeriform species (0.62-3.2 pg. The average sperm head diameter of G. maculatus is lower than the average sperm head diameter of rainbow trout (used as a standard, which coincides with the differences observed in the nuclear DNA content of both species. This information increases the genome data available for G. maculatus and might be useful in future programs dealing with its genetic manipulation.El contenido de ADN nuclear (valor 2C fue determinado en el pez comercial Galaxias maculatus (Galaxiidae usando microdensitometría de núcleos de eritrocitos sometidos a tinción de Feulgen, utilizando como estándar eritrocitos de trucha arco iris con un valor 2C conocido. El valor 2C de G. maculatus fue 2,21 ± 0,12 pg y su valor C es equivalente a 1,105 pg (1.082,9 pMb. Este valor C está dentro del rango registrado para otras especies de osmeriformes (0,62-3,2 pg. El diámetro promedio de la cabeza del espermatozoide de G. maculatus es menor al promedio descrito para la trucha arco iris utilizado como estándar, lo que coincide con las diferencias observadas en el contenido de ADN nuclear entre ambas especies. Estos datos contribuyen a ampliar los antecedentes genómicos disponibles para G. maculatus y podrían ser útiles en futuros programas tendientes a su manipulación genética.

  13. Efficacy of plant extracts against the cowpea beetle, Callosobruchus maculatus

    NARCIS (Netherlands)

    Boeke, S.J.; Barnaud, B.; Loon, van J.J.A.; Kossou, D.K.; Huis, van A.; Dicke, M.


    Traditionally used African plant powders, with a known effect against the cowpea beetle Callosobruchus maculatus in stored cowpea, were extracted with water. The extracts, 13 volatile oils, 2 non-volatile oils and 8 slurries, were evaluated for their toxic and repellent effects against the beetle. A

  14. Baculovirus display of functional antibody Fab fragments. (United States)

    Takada, Shinya; Ogawa, Takafumi; Matsui, Kazusa; Suzuki, Tasuku; Katsuda, Tomohisa; Yamaji, Hideki


    The generation of a recombinant baculovirus that displays antibody Fab fragments on the surface was investigated. A recombinant baculovirus was engineered so that the heavy chain (Hc; Fd fragment) of a mouse Fab fragment was expressed as a fusion to the N-terminus of baculovirus gp64, while the light chain of the Fab fragment was simultaneously expressed as a secretory protein. Following infection of Sf9 insect cells with the recombinant baculovirus, the culture supernatant was analyzed by enzyme-linked immunosorbent assay using antigen-coated microplates and either an anti-mouse IgG or an anti-gp64 antibody. A relatively strong signal was obtained in each case, showing antigen-binding activity in the culture supernatant. In western blot analysis of the culture supernatant using the anti-gp64 antibody, specific protein bands were detected at an electrophoretic mobility that coincided with the molecular weight of the Hc-gp64 fusion protein as well as that of gp64. Flow cytometry using a fluorescein isothiocyanate-conjugated antibody specific to mouse IgG successfully detected the Fab fragments on the surface of the Sf9 cells. These results suggest that immunologically functional antibody Fab fragments can be displayed on the surface of baculovirus particles, and that a fluorescence-activated cell sorter with a fluorescence-labeled antigen can isolate baculoviruses displaying specific Fab fragments. This successful baculovirus display of antibody Fab fragments may offer a novel approach for the efficient selection of specific antibodies.

  15. Insecticide activity of essential oils of Mentha longifolia, Pulicaria gnaphalodes and Achillea wilhelmsii against two stored product pests, the flour beetle, Tribolium castaneum, and the cowpea weevil, Callosobruchus maculatus. (United States)

    Khani, Abbas; Asghari, Javad


    Essential oils extracted from the foliage of Mentha longifolia (L.) (Lamiales: Lamiaceae) and Pulicaria gnaphalodes Ventenat (Asterales: Asteraceae), and flowers of Achillea wilhelmsii C. Koch (Asterales: Asteraceae) were tested in the laboratory for volatile toxicity against two storedproduct insects, the flour beetle, Tribolium castaneum Herbst (Coleoptera: Tenebrionidae) and the cowpea weevil, Callosobruchus maculatus F. (Coleoptera: Bruchidae). The chemical composition of the isolated oils was examined by gas chromatography-mass spectrometry. InM longifolia, the major compounds were piperitenon (43.9%), tripal (14.3%), oxathiane (9.3%), piperiton oxide (5.9%), and d-limonene (4.3%). In P. gnaphalodes, the major compounds were chrysanthenyl acetate (22.38%), 2L -4L-dihydroxy eicosane (18.5%), verbenol (16.59%), dehydroaromadendrene (12.54%), β-pinen (6.43%), and 1,8 cineol (5.6%). In A. wilhelmsii, the major compounds were 1,8 cineole (13.03%), caranol (8.26%), alpha pinene (6%), farnesyl acetate (6%), and p-cymene (6%). C maculatus was more susceptible to the tested plant products than T castaneum. The oils of the three plants displayed the same insecticidal activity against C. maculatus based on LC(50) values (between 1.54µl/L air in P. gnaphalodes, and 2.65 µl/L air in A. wilhelmsii). While the oils of A. wilhelmsii and M. longifolia showed the same strong insecticidal activity against T. castaneum (LC(50) = 10.02 and 13.05 µl/L air, respectively), the oil of P. gnaphalodes revealed poor activity against the insect (LC(50) = 297.9 µl/L air). These results suggested that essential oils from the tested plants could be used as potential control agents for stored-product insects.

  16. Structure of the omalizumab Fab. (United States)

    Jensen, Rasmus K; Plum, Melanie; Tjerrild, Luna; Jakob, Thilo; Spillner, Edzard; Andersen, Gregers Rom


    Omalizumab is a humanized anti-IgE antibody that inhibits the binding of IgE to its receptors on mast cells and basophils, thus blocking the IgE-mediated release of inflammatory mediators from these cells. Omalizumab binds to the Fc domains of IgE in proximity to the binding site of the high-affinity IgE receptor FcℇRI, but the epitope and the mechanisms and conformations governing the recognition remain unknown. In order to elucidate the molecular mechanism of its anti-IgE activity, the aim was to analyse the interaction of omalizumab with human IgE. Therefore, IgE Fc Cℇ2-4 was recombinantly produced in mammalian HEK-293 cells. Functionality of the IgE Fc was proven by ELISA and mediator-release assays. Omalizumab IgG was cleaved with papain and the resulting Fab was purified by ion-exchange chromatography. The complex of IgE Fc with omalizumab was prepared by size-exclusion chromatography. However, crystals containing the complex were not obtained, suggesting that the process of crystallization favoured the dissociation of the two proteins. Instead, two structures of the omalizumab Fab with maximum resolutions of 1.9 and 3.0 Å were obtained. The structures reveal the arrangement of the CDRs and the position of omalizumab residues known from prior functional studies to be involved in IgE binding. Thus, the structure of omalizumab provides the structural basis for understanding the function of omalizumab, allows optimization of the procedure for complex crystallization and poses questions about the conformational requirements for anti-IgE activity.

  17. Characterization of cowpea genotype resistance to Callosobruchus maculatus

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    Maria de Jesus Passos de Castro


    Full Text Available The objective of this work was to characterize the resistance of 50 cowpea (Vigna unguiculata genotypes to Callosobruchus maculatus. A completely randomized design with five replicates per treatment (genotype was used. No-choice tests were performed using the 50 cowpea genotypes to evaluate the preference for oviposition and the development of the weevil. The genotypes IT85 F-2687, MN05-841 B-49, MNC99-508-1, MNC99-510-8, TVu 1593, Canapuzinho-1-2, and Sanzi Sambili show non-preference-type resistance (oviposition and feeding. IT81 D-1045 Ereto and IT81 D-1045 Enramador exhibit antibiosis against C. maculatus and descend from resistant genitors, which grants them potential to be used in future crossings to obtain cowpea varieties with higher levels of resistance.

  18. [Alpha-glucosidase inhibitory activity of Aeschynanthus maculatus]. (United States)

    Tian, Pu-yu; Kang, Wen-yi


    To study the inhibitory activity of Aeschynanthus maculatus on alpha-glucosidase. The inhibilitory model of in vitro alpha-glucosidase was established. Active extracts of A. maculatus were isolated and identified bymultiple chromatographic methods, and their molecular structures were identifiied by spectral techniques. Seven coumpounts were isolated from A. maculatus and isolated as lupeol(1), stigmasterol(2), ursolic acid(3), stigmast-5,22(E)-diene-3beta-ol(4), beta-daucosterol(5), 3-hydroxy-12-taraxasten-28-oic-acid(6) and oleanic acid(7). Compounds 1 (IC50 25.41 mg x L(-1)),3(IC0 4.42 mg L(-1)),4(IC50 11.50 mg x L(-1)),6(IC50 14.17 mg x L(-1)) and 7(IC50 2.88 mg x L(-1)) had higher inhibitory activities than that of acarbose (IC50 1103.01 mg x L(-1)) as the positive control drug. Compound 1-7 were isolated from this plant for the first time. Compound 6 was isolated from Gesneriaceae family for the first time. Compound 7 was isolated from Aeschynanthus genus for the first time.

  19. Peel and leaf powders of three fruits and a vegetable as promising botanicals against Callosobruchus maculatus (F.

    Directory of Open Access Journals (Sweden)

    Musa Abdulrasak K.


    Full Text Available The aim of this study was to examine the insecticidal activities of powders of bitter leaf, cashew leaf, orange peel and pawpaw leaf against Callosobruchus maculatus (F. (Coleoptera: Bruchidae. These were tested at 2.5%, 5.0% and 7.5% (w/w for their insecticidal activities against the insect. Results showed that the ovicidal and adulticidal activities of the plant powders depended on dosages and exposure time. Mortalities of 16.28% and 18.75% were observed in bitter leaf powder applied at 7.5% (w/w dosage within 24 and 48 hours post infestation (HPI, respectively. In grains treated with 7.5% orange peel powder, 12.50% and 16.28% mortalities were observed within 24 and 48 HPI, respectively. Each of bitter leaf and orange peel powders was significantly different (p orange peel > cashew leaf > pawpaw leaf. It is recommended that incorporating these plant parts in pest management of stored products will guarantee user safety, reduce environmental pollution and suppress insect infestation.

  20. A screen for bacterial endosymbionts in the model organisms Tribolium castaneum, T. confusum, Callosobruchus maculatus, and related species. (United States)

    Goodacre, Sara L; Fricke, Claudia; Martin, Oliver Y


    Reproductive parasites such as Wolbachia are extremely widespread amongst the arthropods and can have a large influence over the reproduction and fitness of their hosts. Undetected infections could thus confound the results of a wide range of studies that focus on aspects of host behavior, reproduction, fitness, and degrees of reproductive isolation. This potential problem has already been underlined by work investigating the incidence of Wolbachia infections in stocks of the model system Drosophila melanogaster. Here we survey a range of lab stocks of further commonly used model arthropods, focusing especially on the flour beetles Tribolium castaneum and Tribolium confusum, the cowpea weevil Callosobruchus maculatus and related species (Coleoptera: Tenebrionidae and Bruchidae). These species are widespread stored product pests so knowledge of infections with symbionts further has potential use in informing biocontrol measures. Beetles were assessed for infection with 3 known microbial reproductive parasites: Wolbachia, Rickettsia, Spiroplasma. Infections with some of these microbes were found in some of the lab stocks studied, although overall infections were relatively rare. The consequences of finding infections in these or other species and the type of previous studies likely to be affected most are discussed. © 2013 Institute of Zoology, Chinese Academy of Sciences.

  1. FAB (Functionally Alert Behavior Strategies) to Improve Self-Control (United States)

    Pagano, John


    This paper describes the FAB (Functionally Alert Behavior) Strategies approach to improve behavior in children and adolescents with complex behavioral challenges. FAB Strategies include evidence-based environmental adaptations, sensory modulation, positive behavioral support, and physical self-regulation strategies. FAB Strategies can be used by…

  2. Rapportage LTO FAB II 2008 : Functionele Agro Biodiversiteit

    NARCIS (Netherlands)

    Scheele, J.; Gurp, van H.; Alebeek, van F.A.N.; Belder, den E.; Elderson, J.


    Doel van het LTO FAB II project is een gebruiksklaar FAB concept te ontwikkelen voor een aantal ziekten en plagen in een aantal gewassen die op eenvoudige wijze door telers benut kan worden en voor de toepasser kostenneutraal zijn. Uitgangspunt in het FAB project is een evenwichtige balans tussen de

  3. FabH Mutations Confer Resistance to FabF-Directed Antibiotics in Staphylococcus aureus


    Parsons, Joshua B.; Yao, Jiangwei; Frank, Matthew W.; Rock, Charles O.


    Delineating the mechanisms for genetically acquired antibiotic resistance is a robust approach to target validation and anticipates the evolution of clinical drug resistance. This study defines a spectrum of mutations in fabH that render Staphylococcus aureus resistant to multiple natural products known to inhibit the elongation condensing enzyme (FabF) of bacterial type II fatty acid synthesis. Twenty independently isolated clones resistant to platensimycin, platencin, or thiolactomycin were...

  4. Production of single chain Fab (scFab fragments in Bacillus megaterium

    Directory of Open Access Journals (Sweden)

    Dübel Stefan


    Full Text Available Abstract Background The demand on antigen binding reagents in research, diagnostics and therapy raises questions for novel antibody formats as well as appropriate production systems. Recently, the novel single chain Fab (scFab antibody format combining properties of single chain Fv (scFv and Fab fragments was produced in the Gram-negative bacterium Escherichia coli. In this study we evaluated the Gram-positive bacterium Bacillus megaterium for the recombinant production of scFab and scFvs in comparison to E. coli. Results The lysozyme specific D1.3 scFab was produced in B. megaterium and E. coli. The total yield of the scFab after purification obtained from the periplasmic fraction and culture supernatant of E. coli was slightly higher than that obtained from culture supernatant of B. megaterium. However, the yield of functional scFab determined by analyzing the antigen binding activity was equally in both production systems. Furthermore, a scFv fragment with specificity for the human C reactive protein was produced in B. megaterium. The total yield of the anti-CRP scFv produced in B. megaterium was slightly lower compared to E. coli, whereas the specific activity of the purified scFvs produced in B. megaterium was higher compared to E. coli. Conclusion B. megaterium allows the secretory production of antibody fragments including the novel scFab antibody format. The yield and quality of functional antibody fragment is comparable to the periplasmic production in E. coli.

  5. [New metacercariae of the genus Tylodelphys (Trematoda, Diplostomatidae) in lacustrine populations of Galaxias maculatus (Teleostei, Galaxiidae)]. (United States)

    Quaggiotto, E A; Valverde, F


    Metacercariae of three new species of Tylodelphys (Trematoda, Diplostomatidae) were found in the brain of Galaxias maculatus (Teleostei, Galaxiidae), at the Meliquina Lake, Neuquen Province, Argentina. These metacercariae are morphologically described as Tylodelphys argentinus n. sp., Tylodelphys barilochensis n. sp. and Tylodelphys crubensis n. sp. This is the first time that a stage of genus Tylodelphys is mentioned parasitizing Galaxias maculatus. Metacercariae of T. barilochen sis n. sp. were also found in G. maculatus brain at the Perito Moreno Lake, Rio Negro Province, Argentina.

  6. Estabilidade da resistência de genótipos de caupi a Callosobruchus maculatus (Fabr. em gerações sucessivas

    Directory of Open Access Journals (Sweden)

    Lima Marcileyne Pessôa Leite de


    Full Text Available Callosobruchus maculatus (Fabr. (Coleoptera: Bruchidae é a principal praga do caupi (Vigna unguiculata L. armazenado em condições tropicais e subtropicais. Avaliaram-se a estabilidade da resistência e a capacidade de adaptação de C. maculatus a genótipos de caupi, durante seis gerações. Utilizou-se o teste sem chance de escolha, em delineamento experimental inteiramente casualizado, em esquema fatorial, com dez tratamentos, seis gerações do inseto e cinco repetições. Cada repetição constou de 30 grãos infestados com dois casais da praga. O número de ovos/fêmea diferiu entre os genótipos de caupi apenas na sexta geração, e entre as gerações em BR14-Mulato, Bico de Pato, TE90-180-3E e TE87-98-8G. A viabilidade de ovos diferiu entre as gerações em BR17-Gurguéia, BR14-Mulato, IT89KD-260 e IT89KD-245, e entre os genótipos nas terceira, quinta e sexta gerações. Os genótipos diferiram entre e dentro das gerações, em relação à duração e a viabilidade da fase imatura. Observou-se redução na emergência, especialmente em IT89KD-245, IT89KD-260, CNC 0434, Bico de Pato, TE90-180-10F e BR14-Mulato, provavelmente devido a substâncias químicas presentes nos grãos que afetaram a sobrevivência dos insetos, ao longo das gerações. Os genótipos IT89KD-245 e IT89KD-260 comportaram-se como moderadamente resistentes. Não se observou adaptação de C. maculatus aos genótipos, mantendo-se a resistência estável através das gerações.

  7. Erstnachweise von Paratrachelas maculatus in Österreich und Deutschland (Araneae, Corinnidae)


    Bauer, Tobias; Grabolle, Arno


    Three adult females of Paratrachelas maculatus (Thorell, 1875) were found inside a house in the south of Vienna, in a cellar in Cologne and in a house in Rüsselsheim. Additional notes on diet in captivity are presented.

  8. Monitoring strategy to match the advanced fabs (United States)

    Ackmann, Paul W.


    The reduction in feature size below the exposure wavelength, the requirement for high yields, the expectation for consistent cycletime and shipment to mix, all mean that the reticle industry must be like advanced wafer fabrication centers. Due to the lower output of write tools versus steppers, and the fact that a reticle is a lot of one instead of 25 or 50 wafers as well as the need to match ship data to Fab ramp, the reticle line monitoring strategy must be optimized for small sample size. The use of tool time and alternative inspection strategies can lead to the early detection of problems. Because every reticle is a customer specific design, the monitoring strategy takes on a new look compared to the Fab. We have organized the AMTC to resemble a wafer fab. We have a dedicated Integration group that works with the customers and technologists, to monitor the needs of the customers and then drive the development programs that improve reticle capability. We have dedicated yield team to identify and classify the yield loss mechanisms and define probable causes. The teams then work with the Process owners to fix the source of yield loss and track the corrective actions. All sources of variations must be modeled and then sources of errors reduced to levels below the tool specification. The manufacturing organization has all the process and tool experts to focus on Pilot Line and Development tasks to meet the advance needs of our customers. With the organization in place we can then develop the methods based on Reticle and Fab manufacturing to best control the line and provide development with manufacturing cycle times.

  9. Mask qualification strategies in a wafer fab (United States)

    Jaehnert, Carmen; Kunowski, Angela


    Having consistent high quality photo masks is one of the key factors in lithography in the wafer fab. Combined with stable exposure- and resist processes, it ensures yield increases in production and fast learning cycles for technology development and design evaluation. Preventive controlling of incoming masks and quality monitoring while using the mask in production is essential for the fab to avoid yield loss or technical problems caused by mask issues, which eventually result in delivery problems to the customer. In this paper an overview of the procedures used for mask qualification and production release, for both logic and DRAM, at Infineon Dresden is presented. Incoming qualification procedures, such as specification checks, incoming inspection, and inline litho process window evaluation, are described here. Pinching and electrical tests, including compatibility tests for mask copies for high volume products on optimized litho processes, are also explained. To avoid mask degradation over lifetime, re-inspection checks are done for re-qualification while using the mask in production. The necessity of mask incoming inspection and re-qualification, due to the repeater printing from either the processing defects of the original mask or degrading defects of being used in the fab (i.e. haze, ESD, and moving particles, etc.), is demonstrated. The need and impact of tight mask specifications, such as CD uniformity signatures and corresponding electrical results, are shown with examples of mask-wafer CD correlation.

  10. Fatty acid biosynthesis in Pseudomonas aeruginosa: cloning and characterization of the fabAB operon encoding beta-hydroxyacyl-acyl carrier protein dehydratase (FabA) and beta-ketoacyl-acyl carrier protein synthase I (FabB).


    Hoang, T.T.; Schweizer, H P


    The Pseudomonas aeruginosa fabA and fabB genes, encoding beta-hydroxyacyl-acyl carrier protein dehydratase and beta-ketoacyl-acyl carrier protein synthase I, respectively, were cloned, sequenced, and expressed in Escherichia coli. Northern analysis demonstrated that fabA and fabB are cotranscribed and most probably form a fabAB operon. The FabA and FabB proteins were similar in size and amino acid composition to their counterparts from Escherichia coli and to the putative homologs from Haemop...

  11. Beetles (Insecta, Coleoptera) associated with pig carcasses exposed in a Caatinga area, Northeastern Brazil. (United States)

    Santos, W E; Alves, A C F; Creão-Duarte, A J


    The species richness, abundance and seasonality of Coleoptera fauna associated with pig carcasses exposed in a Caatinga area were examined. Tray, pitfall and modified Shannon traps were settled together to collect these insects during two seasons (dry and rainy). 4,851 beetles were collected, belonging to 19 families and 88 species. Staphylinidae (2,184) and Histeridae (1,264) were the most abundant families and accounted for 71.1% of the specimens collected. Scarabaeidae (15) showed the highest species richness. The most abundant species were Atheta iheringi Bernhauer, 1908 (Staphylinidae) (1,685), Euspilotus sp. (Histeridae) (461), Stelidota geminata (Say, 1825) (Nitidulidae) (394), Xerosaprinus diptychus (Marseul, 1855) (Histeridae) (331) and Dermestes maculatus De Geer, 1774 (Dermestidae). Amongst these species, X. diptychus showed to be strongly influenced by seasonality, since 96.1% of the specimens were collected during the dry season.

  12. Beetles (Insecta, Coleoptera associated with pig carcasses exposed in a Caatinga area, Northeastern Brazil

    Directory of Open Access Journals (Sweden)

    WE Santos

    Full Text Available The species richness, abundance and seasonality of Coleoptera fauna associated with pig carcasses exposed in a Caatinga area were examined. Tray, pitfall and modified Shannon traps were settled together to collect these insects during two seasons (dry and rainy. 4,851 beetles were collected, belonging to 19 families and 88 species. Staphylinidae (2,184 and Histeridae (1,264 were the most abundant families and accounted for 71.1% of the specimens collected. Scarabaeidae (15 showed the highest species richness. The most abundant species were Atheta iheringi Bernhauer, 1908 (Staphylinidae (1,685, Euspilotus sp. (Histeridae (461, Stelidota geminata (Say, 1825 (Nitidulidae (394, Xerosaprinus diptychus(Marseul, 1855 (Histeridae (331 and Dermestes maculatus De Geer, 1774 (Dermestidae. Amongst these species, X. diptychus showed to be strongly influenced by seasonality, since 96.1% of the specimens were collected during the dry season.

  13. Fungi associated to Platypus cylindrus Fab. (Coleoptera: Platypodidae in cork oak Fungos associados ao insecto Platypus cylindrus Fab. (Coleoptera: Platypodidae em sobreiro

    Directory of Open Access Journals (Sweden)

    Joana Henriques


    Full Text Available Platypus cylindrus is a pest that since the 80’s of the last century has been considered a cork oak mortality agent in Portugal. It is an ambrosia beetle that establishes complex symbioses with fungi whose role in the insect-fungus-host interaction has not been completely clarified. In order to characterize P. cylindrus associated micoflora in Portugal, fungi were isolated from different beetle organs and from its galleries in cork oak trees. Fungi of the genera Acremonium, Aspergillus, Beauveria, Botrytis, Chaetomium, Fusarium, Geotrichum, Gliocladium, Nodulisporium, Paecilomyces, Penicillium, Raffaelea, Scytalidium, Trichoderma and of the order Mucorales were identified. An actinomycete of the genus Streptomyces was also identified. Some of these genera were related for the first time to this interaction. In the present work the isolated fungi are characterized and their contribution for beetle population establishment and tree weakness is discussed.Platypus cylindrus é uma praga que desde os anos 80 do século passado tem sido referida como agente de mortalidade do sobreiro em Portugal. É um insecto ambrósia que estabelece simbioses complexas com fungos cujo papel não está completamente esclarecido na interacção insecto-fungo-sobreiro. Com o objectivo de caracterizar a micoflora associada a P. cylindrus em Portugal foram efectuados isolamentos a partir de diferentes órgãos do insecto e suas galerias em sobreiro. Identificaram-se fungos dos géneros Acremonium, Aspergillus, Beauveria, Botrytis, Chaetomium, Fusarium, Geotrichum, Gliocladium, Nodulisporium, Paecilomyces, Penicillium, Raffaelea, Scytalidium, Trichoderma e da ordem Mucorales. Foi igualmente identificado um actinomiceta do género Streptomyces. Alguns destes géneros são referidos pela primeira vez nesta interacção. No presente trabalho caracterizam-se os fungos isolados e discute-se a sua contribuição para o estabelecimento das populações do insecto e enfraquecimento das árvores.

  14. Identification and Function Reasearch of fabA and fabB of Sinorhizobium meliloti%苜蓿中华根瘤菌fabA和fabB基因功能的鉴定

    Institute of Scientific and Technical Information of China (English)

    胡喆; 马金成; 蒋晶晶; 王海洪


    在大肠杆菌(Escherichia coli)脂肪酸合成酶体系中,fabA基因编码有双功能的3-羟基脂酰ACP脱水异构酶,其异构产物能被fabB基因编码的3-酮基脂酰ACP合成酶Ⅰ延伸,合成不饱和脂肪酸,该FabA-FabB途径被认为是缺氧条件下不饱和脂肪酸合成的经典途径.生物信息学分析发现,苜蓿中华根瘤菌(Sinorhizobium meliloti)的SmFabA与EcFabA相似性达到60.6%,具有相同的保守活性位点和两个保守的α螺旋结构;SmFabB与EcFabB相似性达到61.1%,具有相同的Cys-His-His活性中心.用携带SmfabA和SmfabB的质粒载体遗传互补大肠杆菌湿度敏感突变株CY57和CY242,在添加三氯森(TCL)抑制烯脂酰ACP还原酶活性的条件下,转化子能在42℃恢复生长,且放射性薄层层析能检测到转化子中不饱和脂肪酸棕榈油酸(A9C16:1)和十八碳烯酸(△11C18:1)的合成.体外重建脂肪酸合成反应表明,SmFabA能催化羟脂酰ACP的脱水反应且能够使反-2-癸烯酰ACP异构化,SmFabB能催化不同链长的脂酰ACP和丙二酸单酰ACP的聚合反应.另外,未得到SmFabA和SmFabB的突变株,表明SmFabA和SmFabB可能是苜蓿中华根瘤菌脂肪酸合成酶系中必不可少的关键蛋白.上述结果证实了苜蓿中华根瘤菌fabA和fabB两个基因在不饱和脂肪酸合成中的功能.

  15. Influence of substrate and relative humidity on the efficacy of three entomopathogenic fungi for the hide beetle, Dermestes maculatus (United States)

    Dermestes maculatus is carrion feeder that is also a pest of poultry houses, museums, silkworm culture, and many stored foods. The Hypocreales, Beauveria bassiana, Metarhizium anisopliae, and Isaria fumosorosea, were tested for efficacy against D. maculatus larvae on concrete, plastic, leather, and ...

  16. The cosmology of the Fab-Four

    CERN Document Server

    Copeland, Edmund J; Saffin, Paul M


    We have recently proposed a novel self tuning mechanism to alleviate the famous cosmological constant problem, based on the general scalar tensor theory proposed by Horndeski. The self-tuning model ends up consisting of four geometric terms in the action, with each term containing a free potential function of the scalar field; the four together being labeled as the Fab-Four. In this paper we begin the important task of deriving the cosmology associated with the Fab-Four Lagrangian. Performing a phase plane analysis of the system we are able to obtain a number of fixed points for the system, with some remarkable new solutions emerging from the trade-off between the various potentials. As well as obtaining inflationary solutions we also find conventional radiation/matter-like solutions, but in regimes where the energy density is dominated by a cosmological constant, and where we do not have any explicit forms of radiation or matter. Stability conditions for matter solutions are obtained and we show how it is po...

  17. Bruchid (Coleoptera: Bruchidae) ovicidal phenylbutanoid from Zingiber purpureum. (United States)

    Bandara, K A Nimal P; Kumar, Vijaya; Saxena, Ramesh C; Ramdas, Puthenveetil K


    The larvicidal activity of the dichloromethane extract of Zingiber purpureum Roscoe (Zingiberaceae) rhizome against the second instar of Aedes aegypti (L.) (Diptera: Culicidae) is shown to be due to 4-(3',4'-dimethoxyphenyl)buta-1,3-diene. The diene also showed ovicidal activity against the bruchid Callosobruchus maculatus (F.) (Coleoptera: Bruchidae). Most of the eggs laid by bruchids on treated cowpea seeds were transparent, and very few of them contained developing embryos. The few larvae produced from these embryos were unable to penetrate the seed coat and enter the seed. Similar effects were seen when adults were exposed to the compound and then placed on untreated cowpea seeds, suggesting that a new type of maternally mediated ovicidal effect was involved. Coated and impregnated granular formulations of the extract were evaluated for use in the control of bruchid infestation of stored cowpea seeds. Coated granules showed activity similar to that of the crude extract but were found to lose activity rapidly. Impregnated granules were found to be less active than the crude extract.

  18. Een inventarisatie naar de bekendheid van Functionele Agrobiodiversiteit (FAB) en de mogelijkheden om FAB met andere agroranden te combineren

    NARCIS (Netherlands)

    Geerts, R.H.E.M.; Meerburg, B.G.


    Op dit moment zijn er regio's waar het FAB concept al breed wordt toegepast en goed is ingebed (West-Brabant/Zeeland), waar het in opmars is (Flevoland en Zuid-Holland) en enkele regio's waar het FAB gedachtegoed slechts matig aanwezig is (Groningen / Drenthe).

  19. Why do female Callosobruchus maculatus kick their mates?

    Directory of Open Access Journals (Sweden)

    Emile van Lieshout

    Full Text Available Sexual conflict is now recognised as an important driver of sexual trait evolution. However, due to their variable outcomes and effects on other fitness components, the detection of sexual conflicts on individual traits can be complicated. This difficulty is exemplified in the beetle Callosobruchus maculatus, where longer matings increase the size of nutritious ejaculates but simultaneously reduce female future receptivity. While previous studies show that females gain direct benefits from extended mating duration, females show conspicuous copulatory kicking behaviour, apparently to dislodge mating males prematurely. We explore the potential for sexual conflict by comparing several fitness components and remating propensity in pairs of full sibling females where each female mated with a male from an unrelated pair of full sibling males. For one female, matings were terminated at the onset of kicking, whereas the other's matings remained uninterrupted. While fecundity (number of eggs was similar between treatments, uninterrupted matings enhanced adult offspring numbers and fractionally also longevity. However, females whose matings were interrupted at the onset of kicking exhibited an increased propensity to remate. Since polyandry can benefit female fitness in this species, we argue that kicking, rather than being maladaptive, may indicate that females prefer remating over increased ejaculate size. It may thus be difficult to assess the presence of sexual conflict over contested traits such as mating duration when females face a trade off between direct benefits gained from one mating and indirect benefits from additional matings.

  20. The mimetic repertoire of the spotted bowerbird Ptilonorhynchus maculatus (United States)

    Kelley, Laura A.; Healy, Susan D.


    Although vocal mimicry in songbirds is well documented, little is known about the function of such mimicry. One possibility is that the mimic produces the vocalisations of predatory or aggressive species to deter potential predators or competitors. Alternatively, these sounds may be learned in error as a result of their acoustic properties such as structural simplicity. We determined the mimetic repertoires of a population of male spotted bowerbirds Ptilonorhynchus maculatus, a species that mimics predatory and aggressive species. Although male mimetic repertoires contained an overabundance of vocalisations produced by species that were generally aggressive, there was also a marked prevalence of mimicry of sounds that are associated with alarm such as predator calls, alarm calls and mobbing calls, irrespective of whether the species being mimicked was aggressive or not. We propose that it may be the alarming context in which these sounds are first heard that may lead both to their acquisition and to their later reproduction. We suggest that enhanced learning capability during acute stress may explain vocal mimicry in many species that mimic sounds associated with alarm.

  1. Phylogeographical population structure of tiger quolls Dasyurus maculatus (Dasyuridae: Marsupialia), an endangered carnivorous marsupial. (United States)

    Firestone, K B; Elphinstone, M S; Sherwin, W B; Houlden, B A


    Tiger quolls, Dasyurus maculatus, are the largest carnivorous marsupials still extant on the mainland of Australia, and occupy an important ecological niche as top predators and scavengers. Two allopatric subspecies are recognized, D.m. gracilis in north Queensland, and D.m. maculatus in the southeast of the mainland and Tasmania. D.m. gracilis is considered endangered while D.m. maculatus is listed as vulnerable to extinction; both subspecies are still in decline. Phylogeographical subdivision was examined to determine evolutionarily significant units (ESUs) and management units (MUs) among populations of tiger quolls to assist in the conservation of these taxa. Ninety-three tiger quolls from nine representative populations were sampled from throughout the species range. Six nuclear microsatellite loci and the mitochondrial DNA (mtDNA) control region (471 bp) were used to examine ESUs and MUs in this species. We demonstrated that Tasmanian tiger quolls are reciprocally monophyletic to those from the mainland using mtDNA analysis, but D.m. gracilis was not monophyletic with respect to mainland D.m. maculatus. Analysis of microsatellite loci also revealed significant differences between the Tasmanian and mainland tiger quolls, and between D.m. gracilis and mainland D.m. maculatus. These results indicate that Tasmanian and mainland tiger quolls form two distinct evolutionary units but that D.m. gracilis and mainland D.m. maculatus are different MUs within the same ESU. The two marker types used in this study revealed different male and female dispersal patterns and indicate that the most appropriate units for short-term management are local populations. A revised classification and management plan are needed for tiger quolls, particularly in relation to conservation of the Tasmanian and Queensland populations.

  2. HIV-2 neutralization by intact V3-specific Fab fragments

    Directory of Open Access Journals (Sweden)

    Sourial Samer


    Full Text Available Abstract The V3 region of both HIV-1 gp120 and HIV-2 gp125 surface glycoprotein has been described as a target for neutralizing antibodies. In this study a conformation-sensitive (3C4 and a linear site-specific (7C8 anti-HIV-2 V3 monoclonal antibody (mAb were characterized. The neutralization capacity of the purified mAbs and their respective papain-generated Fab fragments was analyzed. The Fabs were further characterized by sequence analysis. Our results demonstrate that neither purified mAbs were capable of neutralizing HIV-2, while intact Fab fragments from both mAbs blocked in vitro infection of HIV-2 isolates. Moreover, the conformation sensitive 3C4 Fab neutralized both subtype A and B HIV-2 isolates and SIVsm. Sequence analysis of the hypervariable regions of 3C4 Fab and 7C8 Fab revealed that the third CDR of the heavy chain (CDRH3 of the antibodies was not as long as many of the previously characterized neutralizing antibodies. Our findings suggest that whole 7C8 and 3C4 mAbs are sterically hindered from neutralizing HIV-2, whereas the smaller size of Fab fragments enables access to the V3 region on the virion surface.

  3. Adult Diapause in Coleoptera

    Directory of Open Access Journals (Sweden)

    Ivo Hodek


    Full Text Available Recent studies dealing with adult (reproductive diapause in the Coleoptera are reviewed, as a kind of supplement to the classic compendia. In the first two sections, the general characteristics of adult diapause are described and principal terms explained. Original articles dealing with 19 species from nine coleopteran families (Coccinellidae, Chrysomelidae, Bruchidae, Curculionidae, Carabidae, Silphidae, Scolytidae, Scarabaeidae, and Endomychidae are reviewed. Finally attempts are made at generalisations from the papers reviewed, and hypotheses on diapause evolution are inferred. A polyphenic character of diapause is a prominent feature in C. septempunctata and L. decemlineata, but has been found also in other Coleoptera and in insects generally and often generates voltinism heterogeneity within populations.

  4. Semiconductor industry wafer fab exhaust management

    CERN Document Server

    Sherer, Michael J


    Given the myriad exhaust compounds and the corresponding problems that they can pose in an exhaust management system, the proper choice of such systems is a complex task. Presenting the fundamentals, technical details, and general solutions to real-world problems, Semiconductor Industry: Wafer Fab Exhaust Management offers practical guidance on selecting an appropriate system for a given application. Using examples that provide a clear understanding of the concepts discussed, Sherer covers facility layout, support facilities operations, and semiconductor process equipment, followed by exhaust types and challenges. He reviews exhaust point-of-use devices and exhaust line requirements needed between process equipment and the centralized exhaust system. The book includes information on wet scrubbers for a centralized acid exhaust system and a centralized ammonia exhaust system and on centralized equipment to control volatile organic compounds. It concludes with a chapter devoted to emergency releases and a separ...

  5. How Fabulous Is Fab 5 Cosmology?

    CERN Document Server

    Linder, Eric V


    Extended gravity origins for cosmic acceleration can solve some fine tuning issues and have useful characteristics, but generally have little to say regarding the cosmological constant problem. Fab 5 gravity can be ghost free and stable, have attractor solutions in the past and future, and possess self tuning that solves the original cosmological constant problem. Here we show however it does not possess all these qualities at the same time. We also demonstrate that the self tuning is so powerful that it not only cancels the cosmological constant but also all other energy density, and we derive the scalings of its approach to a renormalized de Sitter cosmology. While this strong cancellation is bad for the late universe, it greatly eases early universe inflation.

  6. Involvement of larder beetles (Coleoptera: Dermestidae) on human cadavers: a review of 81 forensic cases. (United States)

    Charabidze, Damien; Colard, Thomas; Vincent, Benoit; Pasquerault, Thierry; Hedouin, Valery


    From 1994 to 2013, French forensic entomology laboratories investigated 1,093 cases. Larder beetles (Coleoptera: Dermestidae) were observed in 81 (7.5%) of these cases. To describe and analyze these 81 cases, eight parameters were used: city, location (indoor or outdoor), decay stage (fresh, decay, or dry), dermestid species and instar (adults and/or larvae), presence of living calliphorid larvae, presence of calliphorid pupae or adults, and presence of other necrophagous species. Eight Dermestidae species were observed: Dermestes frischii (42% of cases), Dermestes undulatus (35.8%), Dermestes peruvianus (12.3%), Dermestes lardarius (9.9%), Dermestes haemorrhoidalis (8.6%), Dermestes maculatus (7.4%), Dermestes bicolor (3.7%), and Dermestes ater (1.2%). Larder beetles primarily developed on human cadavers in outdoor locations in areas with a dry climate and were never reported in oceanic areas (which are characterized by frequent rainfall and high ambient humidity). The number of dermestid species on a single corpse never exceeded three. Typically, one species was found per corpse. Species differed between indoor and outdoor cases, with D. frischii and D. undulatus dominant in outdoor cases, while D. peruvianus dominant in indoor cases. Calliphoridae was found in 88% of the cases, while Hydrotaea and Piophilidae were observed 40% of the time. Regarding Coleoptera, Necrobia spp. (Coleoptera: Cleridae) was observed in 46% of the cases. Lastly, we observed a typical decomposition pattern, with preferential feeding areas on the face, hands, and feet (i.e., the extremities). Pupation chambers on or inside the bones were not observed.

  7. Yeast mating for combinatorial Fab library generation and surface display

    Energy Technology Data Exchange (ETDEWEB)

    Feldhaus, Jane M.; Lou, Jianlong; Coleman, James R.; Siegel, Robert W.; Marks, James D.; Feldhaus, Michael


    Yeast display of antibody fragments has proven to be an efficient and productive means for directed evolution of single chain Fv (scFv) antibodies for increased affinity and thermal stability, and more recently for the display and screening of a non-immune library. In this paper, we describe an elegant and simple method for constructing large combinatorial Fab libraries for display on the surface of Saccharomyces cerevisiae, from modestly sized, and easily constructed, heavy and light chain libraries. To this end, we have constructed a set of yeast strains and a two vector system for heavy chain and light chain surface display of Fab fragments with free native amino termini. Through yeast mating of the haploid libraries, a very large heterodimeric immune Fab library was displayed on the diploids and high affinity antigen specific Fabs were isolated from the library.

  8. The Insecticidal Activity of Tea Tree Oil (Melaleuca Alternifolia Against the Common Pest in Mummies (Dermestes Maculatus

    Directory of Open Access Journals (Sweden)



    Full Text Available Egyptian mummies show different signs of deterioration caused by insects, such as missing parts, gaps and accumulated dust. Dermestes maculatus is one of the serious pests that cause damage to Egyptian mummies. To assess the insecticidal activity of tea tree oil against the larvae of the museum insect pest Dermestes maculatus (isolated from Egyptian mummies we tested it under different concentration and treatment times by the bioassay methods. Our results showed that tea tree oil diluted in ethanol was highly toxic to Dermestes maculatus larvae. Insecticidal activity depended on both concentration and exposure time. By increasing the concentration level and the exposure time we obtained a higher mortality rate.

  9. A Case Study of a High School Fab Lab (United States)

    Lacy, Jennifer E.

    This dissertation examines making and design-based STEM education in a formal makerspace. It focuses on how the design and implementation of a Fab Lab learning environment and curriculum affect how instructors and students see themselves engaging in science, and how the Fab Lab relates to the social sorting practices that already take place at North High School. While there is research examining design-based STEM education in informal and formal learning environments, we know little about how K-12 teachers define STEM in making activities when no university or museum partnership exists. This study sought to help fill this gap in the research literature. This case study of a formal makerspace followed instructors and students in one introductory Fab Lab course for one semester. Additional observations of an introductory woodworking course helped build the case and set it into the school context, and provided supplementary material to better understand the similarities and differences between the Fab Lab course and a more traditional design-based learning course. Using evidence from observational field notes, participant interviews, course materials, and student work, I found that the North Fab Lab relies on artifacts and rhetoric symbolic of science and STEM to set itself apart from other design-based courses at North High School. Secondly, the North Fab Lab instructors and students were unable to explain how what they were doing in the Fab Lab was science, and instead relied on vague and unsupported claims related to interdisciplinary STEM practices and dated descriptions of science. Lastly, the design and implementation of the Fab Lab learning environment and curriculum and its separation from North High School's low tech, design-based courses effectively reinforced social sorting practices and cultural assumptions about student work and intelligence.

  10. Import of exotic and zoonotic trematodes (Heterophyidae: Centrocestus sp.) in Xiphophorus maculatus

    DEFF Research Database (Denmark)

    Mehrdana, Foojan; Jensen, Hannah M.; Kania, Per Walter


    Ornamental fish, Xiphophorus maculatus, were imported from Singapore to Denmark for distribution to local aquarists. Importers observed lethargic and erratic swimming patterns among fish and forwarded a total of 30 fish for pathological examination to a university diagnostic service. All fish were...

  11. Erstnachweise von Paratrachelas maculatus in Österreich und Deutschland (Araneae, Corinnidae

    Directory of Open Access Journals (Sweden)

    Bauer, Tobias


    Full Text Available Three adult females of Paratrachelas maculatus (Thorell, 1875 were found inside a house in the south of Vienna, in a cellar in Cologne and in a house in Rüsselsheim. Additional notes on diet in captivity are presented.

  12. Increased Fab thermoresistance via VH-targeted directed evolution. (United States)

    Entzminger, Kevin C; Johnson, Jennifer L; Hyun, Jeongmin; Lieberman, Raquel L; Maynard, Jennifer A


    Antibody aggregation is frequently mediated by the complementarity determining regions within the variable domains and can significantly decrease purification yields, shorten shelf-life and increase the risk of anti-drug immune responses. Aggregation-resistant antibodies could offset these risks; accordingly, we have developed a directed evolution strategy to improve Fab stability. A Fab-phage display vector was constructed and the VH domain targeted for mutagenesis by error-prone PCR. To enrich for thermoresistant clones, the resulting phage library was transiently heated, followed by selection for binding to an anti-light chain constant domain antibody. Five unique variants were identified, each possessing one to three amino acid substitutions. Each engineered Fab possessed higher, Escherichia coli expression yield, a 2-3°C increase in apparent melting temperature and improved aggregation resistance upon heating at high concentration. Select mutations were combined and shown to confer additive improvements to these biophysical characteristics. Finally, the wild-type and most stable triple variant Fab variant were converted into a human IgG1 and expressed in mammalian cells. Both expression level and aggregation resistance were similarly improved in the engineered IgG1. Analysis of the wild-type Fab crystal structure provided a structural rationale for the selected residues changes. This approach can help guide future Fab stabilization efforts.

  13. Implementation of high-resolution reticle inspection in wafer fabs (United States)

    Dayal, Aditya; Bergmann, Nathan M.; Sanchez, Peter


    Many advanced wafer fabs are currently fabricating devices with 130nm or smaller design rules. To meet the challenges at these sub-wavelength technology nodes, fabs are using a variety of resolution enhancement techniques (RETs) in lithography and exploring new methods of processing, inspecting and requalifying photomasks. The acceleration of the lithography roadmap imposes more stringent requirements on mask qualification and requalification to ensure that device yields are not compromised: mask inspection tools of today need to find smaller defects on reticles against considerably more complicated patterns or tighter critical dimensions (CDs). In this paper we describe the early stages of implementation and proliferation of advanced reticle inspection tools at high volume manufacturing wafer fabs. The fabs run incoming multi-surface contamination inspections on masks sent from the mask shop (Intel Mask Operations, IMO), and follow them up with periodic inspections/review to make sure any new contaminant or damage does not go undetected. When necessary, images of defects are electronically presented to engineers at IMO for review. Reticle requalification with these inspection tools reduces or eliminates the need for print test verification. We describe the tools and procedure used to streamline reticle requalification at the fabs and improve the feedback loop between the fabs and the mask shop.

  14. Site-specific conjugation of bifunctional chelator BAT to mouse IgG1 Fab' fragment

    Institute of Scientific and Technical Information of China (English)

    Jun LI; Xue-hao WANG; Xiao-ming WANG; Zhao-lai CHEN


    Aim: To perform a site-specific conjugation of Fab' fragments of a mouse monoclonal antibody(MoAb) B43(of IgG1 subtype) to a bifunctional chelator 6-[p-(bromoacetamido) benzyl]-l,4,8,11-tetraazacyclotetradecane-N,N',N",N'"-tetraacetic acid (BAT) via the thiol groups in the hinge distal to the antigenbinding site of the Fab'. Methods: B43 was cleaved using a simple 2-step method.First, stable F(ab')2 was produced by pepsin treatment. Fab' with free thiol in the hinge region was then obtained by cysteine reduction of F(ab')2. Second, a sitespecific conjugation of Fab' to thiol-specific BAT was performed in a one-step reaction. Results: The Fab' fragment had approximately 1.8 free thiol groups per molecule after cysteine reduction. The conjugation efficiency and the chemical yield were approximately 1.28 moles chelator/Fab' and 74% of the initial concentration of Fab', respectively. The F(ab')2, Fab' and Fab'-BAT all maintained reasonable antigen-binding properties. 67Cu labeling of the conjugate under standard conditions did not impair the immunoreactivity of Fab'-BAT. Conclusion: This is a simple and efficient method for producing immunoreactive conjugates of Fab'-BAT, which can be used to make radiometal-labeled conjugates for further diagnostic and therapeutic applications.

  15. 抗内毒素Fab'的制备%Production of Fab' of the chicken egg yolk antibody against lipopolysaccharide

    Institute of Scientific and Technical Information of China (English)

    马思远; 张雅萍


    目的 研究抗内毒素卵黄免疫球蛋白(IgY)的活性片断Fab',探讨防治内毒素血症的新途径.方法 用内毒素(LPS)作为抗原免疫25周龄德国罗曼鸡,改良水溶法提取抗内毒素IgY,胃蛋白酶切后提取Fab'片断,光密度法测抗内毒素Fab'的浓度和含量、ELISA检测抗内毒素Fab'效价、SDS-聚丙烯酰胺凝胶电泳检测其分子量及纯度. 结果 抗内毒素Fab'含量为4.2 mg/mL蛋黄液,效价为1∶51 200,纯度为92%,相对分子质量为44 000. 结论 抗内毒素Fab'产量大、效价高、特异性强.

  16. The attraction of virgin female hide beetles (Dermestes maculatus to cadavers by a combination of decomposition odour and male sex pheromones

    Directory of Open Access Journals (Sweden)

    von Hoermann Christian


    Full Text Available Abstract Introduction The hide beetle Dermestes maculatus (Coleoptera: Dermestidae feeds as an adult and larva on decomposing animal remains and can also be found on human corpses. Therefore, forensic entomological questions with regard to when and how the first receptive females appear on carcasses are important, as the developmental stages of their larvae can be used to calculate the post-mortem interval. To date, we know that freshly emerged males respond to the cadaver odour of post-bloated carcasses (approximately 9 days after death at Tmean = 27°C, being attracted by benzyl butyrate. This component occurs at its highest concentration at this stage of decay. The aim of our study was to determine the principle of attraction of virgin females to the feeding and breeding substrate. For this purpose, we tested the response of these females to headspace samples of piglet cadavers and male sex pheromones [(Z9-unsaturated fatty acid isopropyl esters] in a Y-olfactometer. Because we expected that such an odour combination is of importance for virgin female attraction, we tested the following two questions: 1 Are virgin female hide beetles attracted by a combination of cadaver odour and male sex pheromones? 2 During which decomposition stage do the first virgin females respond to cadaver odour when combined with male sex pheromones? Results We found that young virgin females were attracted to the cadaver by a combination of cadaver odour and male sex pheromones. Neither cadaver odour alone nor male sex pheromones alone was significantly more attractive than a solvent control. Our results also gave a weak indication that the first young virgin females respond as early as the post-bloating stage to its associated decomposition odour when combined with male sex pheromones. Conclusions Our results indicate that freshly emerged males possibly respond to cadaver odour and visit carcasses before virgin females. Being attracted to cadavers when male sex

  17. Self-tuning and the derivation of the Fab Four

    CERN Document Server

    Charmousis, Christos; Padilla, Antonio; Saffin, Paul M


    We have recently proposed a special class of scalar tensor theories known as the Fab Four. These arose from attempts to analyse the cosmological constant problem within the context of Horndeski's most general scalar tensor theory. The Fab Four together give rise to a model of self-tuning, with the relevant solutions evading Weinberg's no-go theorem by relaxing the condition of Poincare invariance in the scalar sector. The Fab Four are made up of four geometric terms in the action with each term containing a free potential function of the scalar field. In this paper we rigorously derive this model from the general model of Horndeski, proving that the Fab Four represents the only classical scalar tensor theory of this type that has any hope of tackling the cosmological constant problem. We present the full equations of motion for this theory, and give an heuristic argument to suggest that one might be able to keep radiative corrections under control. We also give the Fab Four in terms of the potentials presente...

  18. Insecticidal and repellent activities of the essential oil of Callistemon citrinus (Myrtaceae) against Callosobruchus maculatus (F.) (Coleoptera: Bruchidae). (United States)

    Zandi-Sohani, N; Hojjati, M; Carbonell-Barrachina, Á A


    The essential oil of Callistemon citrinus (Curtis) leaves was extracted by hydro distillation and tested on female and male adults of Callosobruchus macullatus (F.) for insecticidal and repellent effects. GC-MS analysis was used to identify and quantify the volatile composition of the essential oil. Results showed that 1,8-cineole (34.2%) and α-pinene (29.0%) were the major components of the oil. Callistemon citrinus oil was found to be toxic to adult insects when applied by fumigation. Responses varied according to the gender of the insect and exposure time. LC50 values were 12.88 and 84.4 μL. L(-1) for males and females, respectively. An increase in exposure time from 3 to 24 h caused an increase in mortality from 50% to 100% in males and from 15.5% to 85.2% in females, at the highest concentration (500 μL. L(-1)). The essential oil also had a repellent effect against C. macullatus in a filter paper arena test. After 2 and 4 h, 86% and 94%, respectively, repellent effects were demonstrated at the highest concentration of 0.4 μL .cm(-2). These observations suggest that C. citrinus essential oil may be usefully applied to control storage pests.

  19. Bioactivity of Lantana camara L. essential oil against Callosobruchus maculatus (Fabricius Bioactividad de aceite esencial de Lantana camara L. contra Callosobruchus maculatus (Fabricius

    Directory of Open Access Journals (Sweden)

    Nooshin Zandi-Sohani


    Full Text Available Lantana camara L. is a widespread plant species mostly native to subtropical and tropical regions of the world. In this study, insecticidal and repellent activities of L. camara essential oil were evaluated against Callosobruchus maculatus (Fabricius (Col: Bruchidae. Analysis of chemical composition by gas chromatography/mass spectrometry (GC/MS showed high amounts of sesquiterpenes, mainly α-humelene (23.3% and cis-caryophyllene (16.2%. The results showed that the essential oil of L. camara has strong repellent activity against adults of C. maculatus at all tested concentrations. After 2 and 4 h, 97.4 and 100% repellency was seen at highest concentrations of 0.4 μL cm-2, respectively. Moreover, the oil was found to be toxic to adults when applied by fumigation. Responses varied with the gender of insect and exposure time. The LC50 values were 282.7 and 187.9 μL L-1 for females and males, respectively. An increase in the exposure time from 3 to 24 h caused increasing in mortality from 23.6 to 100% in males and from 14.1 to 97.1% in females, at highest concentration (1160 μL L-1. According to these results, L. camara essential oil may be useful as an alternative for bean protection against C. maculatus.Lantana camara L. es una especie vegetal nativa de las regiones subtropicales y tropicales del mundo. En este estudio se evaluaron las actividades insecticida y repelente del aceite esencial de L. camara contra Callosobruchus maculatus (Fabricius. El análisis de la composición volátil de este aceite esencial mediante cromatografía de gas/ espectrometría de masa (GC/MS demostró la presencia de elevadas cantidades de sesquiterpenos, principalmente α-humuleno (23.3% y ci's-cariofileno (16.2%. Los resultados obtenidos demostraron que el aceite esencial de L. camara tiene una elevada actividad repelente contra los adultos de C. maculatus a todas las concentraciones ensayadas. Después de 2 y 4 h la concentración de 0.4 μL cm-2 causó 97

  20. The diet of spotted cuscus (Spilocuscus maculatus in natural and captivity habitat

    Directory of Open Access Journals (Sweden)



    Full Text Available Saragih EW, Sadsoeitoeboen MJ, Pattiselanno F. 2010. The diet of spotted cuscus (Spilocuscus maculatus in natural and captivity habitat. Nusantara Bioscience 2: 78-83. The ex-situ conservation of cuscus (Spilocuscus maculatus under captivating condition is an alternative solution to protect cuscus from extinction. Diets became the main factor in order to support the domestication process. Particular studies on habitat and diet of cuscus have been carried out however there is still limited information on the nutrition aspects of cuscus food. This study aimed to determine the diet type, palatability and nutrient in both natural habitat and captivating condition. The results indicated that there were 19 and 8 plant species identified as cuscus diets in both natural habitat and captivating condition. Cuscus prefers fruits with astringent and sour taste which is contained high crude fiber and low fat.

  1. Correlation between the length and weight of Arius maculatus off the southwestern coast of Taiwan

    Directory of Open Access Journals (Sweden)

    Wu-Shan Chu


    Full Text Available In this study, Arius maculatus were collected using the samples caught by bottom trawling off the southwestern coast of Taiwan. Length-weight and length-length, fork (FL, standard (SL and total (TL lengths, relationships were determined. The relationships between the lengths were all significantly linear (p<0.01, the b value in the length-weight relationship for this value was significantly lower than 3 in the fall (p<0.01, when the temporal changes were taken into account, indicating that only the sampling time affected the growth pattern of A. maculatus. The growth was isometric in the spring, summer and winter, but it was negative and allometric in the fall.

  2. 21 CFR 866.5520 - Immunoglobulin G (Fab fragment specific) immunological test system. (United States)


    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Immunoglobulin G (Fab fragment specific... Test Systems § 866.5520 Immunoglobulin G (Fab fragment specific) immunological test system. (a) Identification. An immunoglobulin G (Fab fragment specific) immunological test system is a device that...

  3. 20 CFR 30.316 - How does the FAB issue a final decision on a claim? (United States)


    ... 20 Employees' Benefits 1 2010-04-01 2010-04-01 false How does the FAB issue a final decision on a... Adjudicatory Process Hearings and Final Decisions on Claims § 30.316 How does the FAB issue a final decision on... waives any objections to all or part of the recommended decision, the FAB may issue a final...

  4. UVB-induced gene expression in the skin of Xiphophorus maculatus Jp 163 B. (United States)

    Yang, Kuan; Boswell, Mikki; Walter, Dylan J; Downs, Kevin P; Gaston-Pravia, Kimberly; Garcia, Tzintzuni; Shen, Yingjia; Mitchell, David L; Walter, Ronald B


    Xiphophorus fish and interspecies hybrids represent long-standing models to study the genetics underlying spontaneous and induced tumorigenesis. The recent release of the Xiphophorus maculatus genome sequence will allow global genetic regulation studies of genes involved in the inherited susceptibility to UVB-induced melanoma within select backcross hybrids. As a first step toward this goal, we report results of an RNA-Seq approach to identify genes and pathways showing modulated transcription within the skin of X. maculatus Jp 163 B upon UVB exposure. X. maculatus Jp 163 B were exposed to various doses of UVB followed by RNA-Seq analysis at each dose to investigate overall gene expression in each sample. A total of 357 genes with a minimum expression change of 4-fold (p-adjbasal expression level of each transcript for each skin sample, (2) the changes in expression levels for each gene in the transcriptome upon exposure to increasing doses of UVB, and (3) clusters of genes that exhibit similar patterns of change in expression upon UVB exposure. These data provide a foundation for understanding the molecular genetic response of fish skin to UVB exposure.

  5. Pollution prevention opportunity assessment for MicroFab and SiFab facilities at Sandia National Laboratories.

    Energy Technology Data Exchange (ETDEWEB)

    Gerard, Morgan Evan


    This Pollution Prevention Opportunity Assessment (PPOA) was conducted for the MicroFab and SiFab facilities at Sandia National Laboratories/New Mexico in Fiscal Year 2011. The primary purpose of this PPOA is to provide recommendations to assist organizations in reducing the generation of waste and improving the efficiency of their processes and procedures. This report contains a summary of the information collected, the analyses performed, and recommended options for implementation. The Sandia National Laboratories Environmental Management System (EMS) and Pollution Prevention (P2) staff will continue to work with the organizations to implement the recommendations.

  6. Production of anti-horse antibodies induced by IgG, F(ab')2 and Fab applied repeatedly to rabbits. Effect on antivenom pharmacokinetics. (United States)

    Vázquez, Hilda; Olvera, Felipe; Alagón, Alejandro; Sevcik, Carlos


    We separated whole IgG, Fab and F(ab')2 fragments from horse plasma. We previously studied the pharmacokinetics of these immunoglobulins and fragments in rabbits and shown that Fab and F(ab')2 pharmacokinetics were well described by a three-exponential kinetics, while IgG and IgG(T) pharmacokinetics, however, deviated from the three-exponential kinetics 120 h after injecting a bolus of the immunotherapeutics; this departure was shown to be due to a surge of anti-horse antibodies occurring after 120 h, peaking at ≈260 h and decaying slowly afterward (Vázquez et al., 2010). We now describe antivenom pharmacokinetics and anti-horse IgG production in rabbits receiving three boluses (300 μg/kg, I.V.) of Fab, F(ab')2 or IgG separated by 21 days.

  7. Suppression of fabB Mutation by fabF1 Is Mediated by Transcription Read-through in Shewanella oneidensis. (United States)

    Li, Meng; Meng, Qiu; Fu, Huihui; Luo, Qixia; Gao, Haichun


    As type II fatty acid synthesis is essential for the growth of Escherichia coli, its many components are regarded as potential targets for novel antibacterial drugs. Among them, β-ketoacyl-acyl carrier protein (ACP) synthase (KAS) FabB is the exclusive factor for elongation of the cis-3-decenoyl-ACP (cis-3-C10-ACP). In our previous study, we presented evidence to suggest that this may not be the case in Shewanella oneidensis, an emerging model gammaproteobacterium renowned for its respiratory versatility. Here, we identified FabF1, another KAS, as a functional replacement for FabB in S. oneidensis In fabB(+) or desA(+) (encoding a desaturase) cells, which are capable of making unsaturated fatty acids (UFA), FabF1 is barely produced. However, UFA auxotroph mutants devoid of both fabB and desA genes can be spontaneously converted to suppressor strains, which no longer require exogenous UFAs for growth. Suppression is caused by a TGTTTT deletion in the region upstream of the fabF1 gene, resulting in enhanced FabF1 production. We further demonstrated that the deletion leads to transcription read-through of the terminator for acpP, an acyl carrier protein gene immediately upstream of fabF1 There are multiple tandem repeats in the region covering the terminator, and the TGTTTT deletion, as well as others, compromises the terminator efficacy. In addition, FabF2 also shows an ability to complement the FabB loss, albeit substantially less effectively than FabF1.

  8. GTP-specific fab fragment-based GTPase activity assay. (United States)

    Kopra, Kari; Rozwandowicz-Jansen, Anita; Syrjänpää, Markku; Blaževitš, Olga; Ligabue, Alessio; Veltel, Stefan; Lamminmäki, Urpo; Abankwa, Daniel; Härmä, Harri


    GTPases are central cellular signaling proteins, which cycle between a GDP-bound inactive and a GTP-bound active conformation in a controlled manner. Ras GTPases are frequently mutated in cancer and so far only few experimental inhibitors exist. The most common methods for monitoring GTP hydrolysis rely on luminescent GDP- or GTP-analogs. In this study, the first GTP-specific Fab fragment and its application are described. We selected Fab fragments using the phage display technology. Six Fab fragments were found against 2'/3'-GTP-biotin and 8-GTP-biotin. Selected antibody fragments allowed specific detection of endogenous, free GTP. The most potent Fab fragment (2A4(GTP)) showed over 100-fold GTP-specificity over GDP, ATP, or CTP and was used to develop a heterogeneous time-resolved luminescence based assay for the monitoring of GTP concentration. The method allows studying the GEF dependent H-Ras activation (GTP binding) and GAP-catalyzed H-Ras deactivation (GTP hydrolysis) at nanomolar protein concentrations.

  9. 20 CFR 30.319 - May a claimant request reconsideration of a final decision of the FAB? (United States)


    ... final decision of the FAB? 30.319 Section 30.319 Employees' Benefits OFFICE OF WORKERS' COMPENSATION... reconsideration of a final decision of the FAB? (a) A claimant may request reconsideration of a final decision of the FAB by filing a written request with the FAB within 30 days from the date of issuance of...

  10. Improved Fab presentation on phage surface with the use of molecular chaperone coplasmid system. (United States)

    Loh, Qiuting; Leong, Siew Wen; Tye, Gee Jun; Choong, Yee Siew; Lim, Theam Soon


    The low presentation efficiency of Fab (fragment antigen binding) fragments during phage display is largely due to the complexity of disulphide bond formation. This can result in the presentation of Fab fragments devoid of a light chain during phage display. Here we propose the use of a coplasmid system encoding several molecular chaperones (DsbA, DsbC, FkpA, and SurA) to improve Fab packaging. A comparison was done using the Fab fragment from IgG and IgD. We found that the use of the coplasmid during phage packaging was able to improve the presentation efficiency of the Fab fragment on phage surfaces. A modified version of panning using the coplasmid system was evaluated and was successful at enriching Fab binders. Therefore, the coplasmid system would be an attractive alternative for improved Fab presentation for phage display.

  11. Protein crystallization with microseed matrix screening: application to human germline antibody Fabs

    Energy Technology Data Exchange (ETDEWEB)

    Obmolova, Galina, E-mail:; Malia, Thomas J.; Teplyakov, Alexey; Sweet, Raymond W.; Gilliland, Gary L., E-mail: [Janssen Research and Development LLC, 1400 McKean Road, Spring House, PA 19477 (United States)


    The power of microseed matrix screening is demonstrated in the crystallization of a panel of antibody Fab fragments. The crystallization of 16 human antibody Fab fragments constructed from all pairs of four different heavy chains and four different light chains was enabled by employing microseed matrix screening (MMS). In initial screening, diffraction-quality crystals were obtained for only three Fabs, while many Fabs produced hits that required optimization. Application of MMS, using the initial screens and/or refinement screens, resulted in diffraction-quality crystals of these Fabs. Five Fabs that failed to give hits in the initial screen were crystallized by cross-seeding MMS followed by MMS optimization. The crystallization protocols and strategies that resulted in structure determination of all 16 Fabs are presented. These results illustrate the power of MMS and provide a basis for developing future strategies for macromolecular crystallization.

  12. Notes on Dutch Cryptophagidae (Coleoptera).

    NARCIS (Netherlands)

    Vorst, O.; Johnson, C.


    Aantekeningen over Nederlandse Cryptophagidae (Coleoptera) Zeven soorten cryptophagiden worden hier voor het eerst gemeld voor de Nederlandse fauna. Van twee soorten is duidelijk dat ze hier al geruime tijd voorkomen. Zo werd het enige Nederlandse exemplaar van Atomaria atra al in 1949 verzameld,

  13. Beschermde kevers in Nederland (Coleoptera)

    NARCIS (Netherlands)

    Huijbregts, J.


    Protected beetles in the Netherlands (Coleoptera) The status of all by law protected beetles of the Netherlands is discussed. Several specimens of Cerambyx cerdo are known from the Netherlands, but all of these are considered to have been imported with oak-wood. Taking the European distribution into

  14. Description of Acleotrema maculatus sp. nov. (Monogenea: Diplectanidae infecting the spotted coral grouper Plectropomus maculatus (F:Serranidea from the Red Sea and its histopathological impact

    Directory of Open Access Journals (Sweden)

    Kareem Morsy


    Full Text Available Acleotrema maculatus sp. nov. (Monogenea: Diplectanidae was described from the gills of The spotted coral grouper Plectropomus maculatus (F:Serranidea, Forsskal, 1775. Fish were collected from boot landing sites and fishermen at different water locations along the Red Sea at Hurghada City, Egypt. The morphology and morphometric characterization of the recovered worms were described by means of light microscopy. Eight (53.3% out 15 specimens of P. maculatus were infected. Most of the infected fish had very pale gills. Morphologically, the adult worm of A. maculatus sp. nov., possessed a body which was elongated, fusiform with a total length 0.86–0.90 (0.88 ± 0.02 mm, and a maximum width 0.09–0.13 (0.11 ± 0.02 mm at the level of ovary. Haptor, broad, differentiated from the rest of the body, measured 0.04–0.08 (0.06 ± 0.02 mm and provided with continuous rows of squamodiscs. Two pairs of lateral hamuli, three bars and 14 marginal hooklets were also observed. Lateral (dorsal bars two, stout, dumbbell-shaped, measured 0.052–0.056 (0.054 ± 0.002 mm in length. Ventral bar slender, with transverse groove, measured 0.09–0.11 (0.10 ± 0.01 mm in length. Ventral hamuli measured 0.02–0.06 (0.04 ± 0.02 mm in total length with an outer root that was very long measured 0.016–0.02 (0.018 ± 0.002, stout, slightly notched at broad proximal end; inner root was conical and measured 0.013–0.017 (0.015 ± 0.002 mm, with shaft measured 0.025–0.029 (0.027 ± 0.002 mm and point length 0.007–0.009 (0.008 ± 0.002 mm. Dorsal hamuli measured 0.034–0.038 (0.036 ± 0.002 in total length; base large, stout, with only lateral rudiment of roots; blade and point were long and curved with shaft length measured 0.024–0.028 (0.026 ± 0.002 mm and 0.02–0.06 (0.04 ± 0.02 mm point length. The worm is hermaphrodite, male copulatory organ measured 0.058–0.062 (0.060 ± 0.002 mm in length with a sclerotized part composed of

  15. Characterization of molecular mechanisms controlling fabAB transcription in Pseudomonas aeruginosa.

    Directory of Open Access Journals (Sweden)

    Herbert P Schweizer

    Full Text Available BACKGROUND: The FabAB pathway is one of the unsaturated fatty acid (UFA synthesis pathways for Pseudomonas aeruginosa. It was previously noted that this operon was upregulated in biofilms and repressed by exogenous UFAs. Deletion of a 30 nt fabA upstream sequence, which is conserved in P. aeruginosa, P. putida, and P. syringae, led to a significant decrease in fabA transcription, suggesting positive regulation by an unknown positive regulatory mechanism. METHODS/PRINCIPAL FINDINGS: Here, genetic and biochemical approaches were employed to identify a potential fabAB activator. Deletion of candidate genes such as PA1611 or PA1627 was performed to determine if any of these gene products act as a fabAB activator. However, none of these genes were involved in the regulation of fabAB transcription. Use of mariner-based random mutagenesis to screen for fabA activator(s showed that several genes encoding unknown functions, rpoN and DesA may be involved in fabA regulation, but probably via indirect mechanisms. Biochemical attempts performed did fail to isolate an activator of fabAB operon. CONCLUSION/SIGNIFICANCE: The data suggest that fabA expression might not be regulated by protein-binding, but by a distinct mechanism such as a regulatory RNA-based mechanism.

  16. Insecticidal activity and chemical composition of the Morinda lucida essential oil against pulse beetle Callosobruchus maculatus. (United States)

    Owolabi, Moses S; Padilla-Camberos, Eduardo; Ogundajo, Akintayo L; Ogunwande, Isiaka A; Flamini, Guido; Yusuff, Olaniyi K; Allen, Kirk; Flores-Fernandez, Karen Isabel; Flores-Fernandez, Jose Miguel


    Insecticidal activity of essential oil extracted from Morinda lucida was tested on pulse beetle Callosobruchus maculatus, which is a pest that causes serious damage to several pulses. The insecticidal activity was compared with two pesticides, Phostoxin and Primo-ban-20. 120 mixed sex adult C. maculatus were introduced, along with 30 g of cowpeas. Four concentrations (0.40, 0.20, 0.10, and 0.05 μg/mL) of the M. lucida essential oil, Phostoxin, and Primo-ban-20 were tested. Essential oil chemical composition was analyzed by GC-MS. M. lucida essential oil showed a high toxicological effect, producing 100% mortality after 72 hours at a dose of 0.20 μg/mL. M. lucida essential oil had a potent insecticidal activity (LC90 = 0.629 μg/mL) compared to both pesticides, Phostoxin (LC90 = 0.652 μg/mL) and Primo-ban-20 (LC90 = 0.726 μg/mL), at 24 h. The main compounds of the essential oil were the oxygenated monoterpenoids, 1,8-cineole (43.4%), and α-terpinyl acetate (14.5%), and the monoterpene hydrocarbons, mostly sabinene (8.2%) and β-pinene (4.0%). Results clearly indicate that M. lucida essential oil can be used as an effective alternative for pulse beetle C. maculatus control, and it could be tested against other pulse beetles affecting Asia and Africa and throughout the world, thereby reducing use of synthetic pesticides.


    Directory of Open Access Journals (Sweden)

    Widiarti Widiarti


    Full Text Available The effectiveness of vector control depends on the susceptibility of the malaria vectors to the insecticide used. Synthetic pyrethroid insecticide are recommended and used for vector control in Java-Bali by CDC. However baseline information on the susceptibility of Anopheles aconitus and Anopheles maculatus to that insecticide were not reported yet. This research therefore was conducted to determine the susceptibility status of malaria vector to the synthetic pyrethroid insecticide. Adults bioassays were carried out using standard WHO test methods. Field blood fed female malaria vector were exposed for one hour to 0.75% permethrin and 0.05% deltamethrin insecticide papers. Susceptibility test of An. aconitus from Wonosobo, Banjarnegara, Purworejo, Pekalongan, Jepara and Kulonprogo regency to 0.75% permethrin insecticide resulted in mortality of 86.0%, 94.0%, 92.5%, 96.0%, 96.0% and 96.25% respectively. Twenty four hours of exposure to 0.05% deltamethrin is required to kill 100%, 97.0%, 97.5%, 94.66%, 66.0% and 100% respectively. The susceptibility test of An. maculatus from, Purworejo (2 locations and Kulonprogo regency to 0. 75% permethrin insecticide resulted in mortality of 85.0% & 100% and 85.0% respectively. Twenty four hours exposure to 0.05% deltamethrin is required to kill 97.5 & 100% and 97.5% respectively. It can be concluded that An. aconitus and An. maculatus mosquito collected from the field in Central Java and Yogyakarta Province shows a reduced susceptibility to both synthetic pyrethroids insecticides. Keywords: Malaria Susceptibility Test, Permethrin dan Deltamethrin

  18. Insecticidal Activity and Chemical Composition of the Morinda lucida Essential Oil against Pulse Beetle Callosobruchus maculatus

    Directory of Open Access Journals (Sweden)

    Moses S. Owolabi


    Full Text Available Insecticidal activity of essential oil extracted from Morinda lucida was tested on pulse beetle Callosobruchus maculatus, which is a pest that causes serious damage to several pulses. The insecticidal activity was compared with two pesticides, Phostoxin and Primo-ban-20. 120 mixed sex adult C. maculatus were introduced, along with 30 g of cowpeas. Four concentrations (0.40, 0.20, 0.10, and 0.05 μg/mL of the M. lucida essential oil, Phostoxin, and Primo-ban-20 were tested. Essential oil chemical composition was analyzed by GC-MS. M. lucida essential oil showed a high toxicological effect, producing 100% mortality after 72 hours at a dose of 0.20 μg/mL. M. lucida essential oil had a potent insecticidal activity (LC90 = 0.629 μg/mL compared to both pesticides, Phostoxin (LC90 = 0.652 μg/mL and Primo-ban-20 (LC90 = 0.726 μg/mL, at 24 h. The main compounds of the essential oil were the oxygenated monoterpenoids, 1,8-cineole (43.4%, and α-terpinyl acetate (14.5%, and the monoterpene hydrocarbons, mostly sabinene (8.2% and β-pinene (4.0%. Results clearly indicate that M. lucida essential oil can be used as an effective alternative for pulse beetle C. maculatus control, and it could be tested against other pulse beetles affecting Asia and Africa and throughout the world, thereby reducing use of synthetic pesticides.

  19. Surviving historical Patagonian landscapes and climate: molecular insights from Galaxias maculatus

    Directory of Open Access Journals (Sweden)

    Carrea Cecilia


    Full Text Available Abstract Background The dynamic geological and climatic histories of temperate South America have played important roles in shaping the contemporary distributions and genetic diversity of endemic freshwater species. We use mitochondria and nuclear sequence variation to investigate the consequences of mountain barriers and Quaternary glacial cycles for patterns of genetic diversity in the diadromous fish Galaxias maculatus in Patagonia (~300 individuals from 36 locations. Results Contemporary populations of G. maculatus, east and west of the Andes in Patagonia, represent a single monophyletic lineage comprising several well supported groups. Mantel tests using control region data revealed a strong positive relationship when geographic distance was modeled according to a scenario of marine dispersal. (r = 0.69, P = 0.055. By contrast, direct distance between regions was poorly correlated with genetic distance (r = -0.05, P = 0.463. Hierarchical AMOVAs using mtDNA revealed that pooling samples according to historical (pre-LGM oceanic drainage (Pacific vs. Atlantic explained approximately four times more variance than pooling them into present-day drainage (15.6% vs. 3.7%. Further post-hoc AMOVA tests revealed additional genetic structure between populations east and west of the Chilean Coastal Cordillera (coastal vs. interior. Overall female effective population size appears to have remained relatively constant until roughly 0.5 Ma when population size rapidly increased several orders of magnitude [100× (60×-190×] to reach contemporary levels. Maximum likelihood analysis of nuclear alleles revealed a poorly supported gene tree which was paraphyletic with respect to mitochondrial-defined haplogroups. Conclusions First diversifying in the central/north-west region of Patagonia, G. maculatus extended its range into Argentina via the southern coastal regions that join the Pacific and Atlantic oceans. More recent gene flow between northern

  20. Spherical Cows in the Sky with Fab Four

    CERN Document Server

    Kaloper, Nemanja


    We explore spherically symmetric static solutions in a subclass of unitary scalar-tensor theories of gravity, called the `Fab Four' models. The weak field large distance solutions may be phenomenologically viable, but only if the Gauss-Bonnet term is negligible. Only in this limit will the Vainshtein mechanism work consistently. Further, classical constraints and unitarity bounds constrain the models quite tightly. Nevertheless, in the limits where the range of individual terms at large scales is respectively Kinetic Braiding, Horndeski, and Gauss-Bonnet, the horizon scale effects may occur while the theory satisfies Solar system constraints and, marginally, unitarity bounds. On the other hand, to bring the cutoff down to below a millimeter constrains all the couplings scales such that `Fab Fours' can't be heard outside of the Solar system.

  1. Evaluation of strategies to control Fab light chain dimer during mammalian expression and purification: A universal one-step process for purification of correctly assembled Fab. (United States)

    Spooner, Jennifer; Keen, Jenny; Nayyar, Kalpana; Birkett, Neil; Bond, Nicholas; Bannister, David; Tigue, Natalie; Higazi, Daniel; Kemp, Benjamin; Vaughan, Tristan; Kippen, Alistair; Buchanan, Andrew


    Fabs are an important class of antibody fragment as both research reagents and therapeutic agents. There are a plethora of methods described for their recombinant expression and purification. However, these do not address the issue of excessive light chain production that forms light chain dimers nor do they describe a universal purification strategy. Light chain dimer impurities and the absence of a universal Fab purification strategy present persistent challenges for biotechnology applications using Fabs, particularly around the need for bespoke purification strategies. This study describes methods to address light chain dimer formation during Fab expression and identifies a novel CH 1 affinity resin as a simple and efficient one-step purification for correctly assembled Fab.

  2. A single-domain antibody-linked Fab bispecific antibody Her2-S-Fab has potent cytotoxicity against Her2-expressing tumor cells. (United States)

    Li, Aifen; Xing, Jieyu; Li, Li; Zhou, Changhua; Dong, Bin; He, Ping; Li, Qing; Wang, Zhong


    Her2, which is frequently overexpressed in breast cancer, is one of the most studied tumor-associated antigens for cancer therapy. Anti-HER2 monoclonal antibody, trastuzumab, has achieved significant clinical benefits in metastatic breast cancer. In this study, we describe a novel bispecific antibody Her2-S-Fab targeting Her2 by linking a single domain anti-CD16 VHH to the trastuzumab Fab. The Her2-S-Fab antibody can be efficiently expressed and purified from Escherichia coli, and drive potent cancer cell killing in HER2-overexpressing cancer cells. In xenograft model, the Her2-S-Fab suppresses tumor growth in the presence of human immune cells. Our results suggest that the bispecific Her2-S-Fab may provide a valid alternative to Her2 positive cancer therapy.

  3. Detection of experimental myocarditis by monoclonal antimyosin antibody, Fab fragment

    Energy Technology Data Exchange (ETDEWEB)

    Rezkalla, S.; Kloner, R.A.; Khaw, B.A.; Haber, E.; Fallon, J.T.; Smith, F.E.; Khatib, R.


    The purpose of this study was to determine whether monoclonal antimyosin Fab (antigen binding fragment) was capable of labeling hearts with experimental coxsackievirus myocarditis, and to determine whether Fab could be used for detecting myocardial damage in either early or chronic phases of the disease. Sixty-five, 3-week-old cesarean-derived 1 (CD 1) mice were divided into two groups: group I (noninfected animals) and group II (infected with coxsackievirus B3). Mice from each group were killed on days 7, 17, 30, or 90 of infection. Forty-eight hours before killing, mice were injected with monoclonal I-125 antimyosin, Fab (25 microCi/injection) and radioactivity was counted in the heart. Selected heart sections were also examined by autoradiography. Heart radioactivity, count/m/mg (m +/- SEM) on days 7, 17, 30, and 90 of infection was 10.8 +/- 1.7, 21.3 +/- 1.1, 11.2 +/- 3.4, and 12.4 +/- 1.5 for group I, versus 36.7 +/- 8.0 (p less than 0.01), 50.0 +/- 4.5 (p less than 0.001), 33.4 +/- 16.1 (p = NS), and 40.6 +/- 8.5 (p less than 0.01) for group II, respectively. Autoradiography revealed focal uptake within areas of necrotic myocardium. We conclude that I125 Fab may be useful in detecting myocardial damage in the experimental model of murine myocarditis up to day 90 of infection.

  4. Preparation of F(ab')2 fragments of immunoglobulin G. (United States)

    Killion, J J; Holtgrewe, E M


    We describe a simple protocol for the preparation of F(ab')2 fragments of immunoglobulin G, based upon the known Fc- binding properties of protein A-Sepharose. The fragment preparations of xenogeneic and allogeneic anti-IgG were noncytotoxic to intact target cells, and were able to block the cytotoxicity of intact antibody. This method should therefore be useful for functional studies not requiring biochemical homogeneity.

  5. Immobilization and functional reconstitution of antibody Fab fragment by solid-phase refolding. (United States)

    Kumada, Yoichi; Hamasaki, Kyoto; Nakagawa, Aya; Sasaki, Eiju; Shirai, Tatsunori; Okumura, Masahiro; Inoue, Manami; Kishimoto, Michimasa


    In this study, we demonstrated the successful preparation of a Fab antibody-immobilized hydrophilic polystyrene (phi-PS) plate via one- and two-step solid-phase refolding methods. Both polystyrene-binding peptide (PS-tag)-fused Fd fragment of heavy chain (Fab H-PS) and full-length of light-chain (Fab L-PS) were individually produced in insoluble fractions of Escherichia coli cells, and they were highly purified in the presence of 8M of urea. Antigen-binding activities of Fab antibody immobilized were correctly recovered by the one-step solid-phase refolding method that a mixture of Fab H-PS and Fab L-PS was immobilized in the presence of 0.5-2M urea, followed by surface washing of the phi-PS plate with PBST. These results indicate that by genetic fusion of a PS-tag, a complex between Fab H and Fab L was efficiently immobilized on the surface of a phi-PS plate even in the presence of a low concentration of urea, and was then correctly refolded to retain its high antigen-binding activity via removal of the urea. A two-step solid-phase refolding method whereby Fab H-PS and Fab L-PS were successively refolded on the surface of a phi-PS plate also resulted in Fab antibody formation on the plate. Furthermore, both the binding affinity and the specificity of the Fab antibody produced by the two-step method were highly maintained, according to the results of sandwich ELISA and competitive ELISA using Fab antibody-immobilized plate via two-step solid-phase refolding. Thus, the solid-phase refolding method demonstrated in this study should be quite useful for the preparation of a Fab antibody-immobilized PS surface with high efficiency from individually produced Fab H-PS and Fab L-PS. This method will be applicable to the preparation of a large Fab antibody library on the surface of a PS plate for use in antibody screening.

  6. Strategy optimization for mask rule check in wafer fab (United States)

    Yang, Chuen Huei; Lin, Shaina; Lin, Roger; Wang, Alice; Lee, Rachel; Deng, Erwin


    Photolithography process is getting more and more sophisticated for wafer production following Moore's law. Therefore, for wafer fab, consolidated and close cooperation with mask house is a key to achieve silicon wafer success. However, generally speaking, it is not easy to preserve such partnership because many engineering efforts and frequent communication are indispensable. The inattentive connection is obvious in mask rule check (MRC). Mask houses will do their own MRC at job deck stage, but the checking is only for identification of mask process limitation including writing, etching, inspection, metrology, etc. No further checking in terms of wafer process concerned mask data errors will be implemented after data files of whole mask are composed in mask house. There are still many potential data errors even post-OPC verification has been done for main circuits. What mentioned here are the kinds of errors which will only occur as main circuits combined with frame and dummy patterns to form whole reticle. Therefore, strategy optimization is on-going in UMC to evaluate MRC especially for wafer fab concerned errors. The prerequisite is that no impact on mask delivery cycle time even adding this extra checking. A full-mask checking based on job deck in gds or oasis format is necessary in order to secure acceptable run time. Form of the summarized error report generated by this checking is also crucial because user friendly interface will shorten engineers' judgment time to release mask for writing. This paper will survey the key factors of MRC in wafer fab.

  7. Disulfide cross-linked Fab-aggregates: preparation and biodistribution. (United States)

    Dalkara, S; Petrov, A; Trubetskoy, V S; Khaw, B A; Torchilin, V P


    The high-molecular-weight soluble aggregates of Fab fragments of murine antibodies against cardiac myosin were prepared as a potential long-circulating and low immunogenic pharmaceutical carriers by conjugation of thiolated Fab and Fab modified with succinimidyl 3-(2-pyridyldithio)propionate. The clearance time and biodistribution of 111In-radiolabeled aggregates were studied in normal and nude-mice bearing human breast tumor implant and in rabbits with experimental myocardial infarction. The aggregates had a prolonged circulation time (half clearance time ca. 3-5 h) and ability to concentrate in the tumor and in the necrotic area of infarcted myocardium. Similar tumor-to-normal and infarct-to-normal accumulation ratios (ca. 3 h in both cases) suggest that combination of long circulation with impaired filtration in necrotic tissues is responsible for this accumulation rather than a specific interaction. The aggregates prepared may serve as long-circulating drug carriers able to deliver pharmaceuticals into areas with affected and leaky vasculature.

  8. A novel bispecific antibody, S-Fab, induces potent cancer cell killing. (United States)

    Li, Li; He, Ping; Zhou, Changhua; Jing, Li; Dong, Bin; Chen, Siqi; Zhang, Ning; Liu, Yawei; Miao, Ji; Wang, Zhong; Li, Qing


    Bispecific antibodies that engage immune cells to kill cancer cells have been actively studied in cancer immunotherapy. In this study, we present a novel bispecific format, S-Fab, fabricated by linking a single-domain anti-carcinoembryonic antigen VHH to a conventional anti-CD3 Fab. In contrast to most bispecific antibodies, the S-Fab bispecific antibody can be efficiently expressed and purified from bacteria. The purified S-Fab is stable in serum and is able to recruit T cells to drive potent cancer cell killing. In xenograft models, the S-Fab antibody suppresses tumor growth in the presence of human immune cells. Our study suggested that the bispecific S-Fab format can be applied to a wide range of immunotherapies.

  9. Haematological changes in Bufo maculatus treated with sublethal concentrations of Cadmium

    Directory of Open Access Journals (Sweden)

    Lawrence Ikechukwu Ezemonye


    Full Text Available Adult Bufo maculatus was exposed to sublethal cadmium concentrations of 0.25, 0.50, 1.00 and 2.00 mg/L. The toxicant from which the cadmium concentrations were prepared was cadmium chloride (CdCl2.H2O. There were three replicate tanks per treatment and three individuals per tank including control groups. The hematologic alterations based on the examination of blood indices during the 28 days of exposure showed that total erythrocyte count (TEC, hematocrit (Hct and hemoglobin (Hb concentration decreased (P<0.05 relative to controls. The decline was concentration- dependent as concentration of cadmium increased. The decline in hemoglobin and hematocrit in the experimental organism could be due to a decrease in the synthesis or release of erythrocytes into the circulation or an increase in the rate of erythrocyte destruction inflicted by cadmium toxicity. There was significant (P<0.05 elevation in total leuko- leukocyte count (TLC with increase in the concen- cyte concentration of cadmium. The increase in total leukocyte count observed in this study could be attributed to a stimulation of the immune system in response to tissue damage caused by cadmium toxicity. The study has shown that the exposure of the Bufo maculatus toad to cadmium can inflict alterations in the hematologic indices, which could induce unfavorable physiological changes in the amphibian, which may lead to death. There is, therefore, the need to protect amphibians in order to sustain the biodiversity in the Nigerian Niger Delta ecological zone.

  10. Acanthostomoides apophalliformis (Trematoda: Cryptogonimidae does not cause detectable mortality in Galaxias maculatus (Teleostomi: Galaxiidae

    Directory of Open Access Journals (Sweden)

    Jorge E. Revenga


    Full Text Available The present study was conducted to determine if parasite induced mortality existed in a "puyen" population in Moreno Lake, southern Argentina. Data on both parasite abundance and degree of aggregation of parasite frequency distribution showed the absence of fish mortality induced by A. apophalliformis.Acanthostomoides apophalliformis (Trematoda: Cryptogonimidae não causa mortalidade detectável no peixe Galaxias maculatus (Teleostomi: Galaxiidae de Argentina do Sul. O "puyen", Galaxias maculatus, é um peixe nativo pequeno que habita a América do Sul e a Oceania. Acanthostomoides apophalliformis é um trematódeo digenético que parasita, freqüentemente, os "puyenes". O estudo atual foi conduzido para determinar se existe mortalidade induzida por parasito no população do "puyen" no lago Moreno, no sul de Argentina. Os dados de abundância do parasito e no grau de aggregação da distribuição de freqüência do parasito, sugerem a ausência de mortalidade induzida por A. apophalliformis em "puyenes".

  11. Effect of body size and temperature on respiration of Galaxias maculatus (Pisces: Galaxiidae) (United States)

    Milano, D.; Vigliano, P.H.; Beauchamp, David A.


    Body mass and temperature are primary determinants of metabolic rate in ectothermic animals. Oxygen consumption of post-larval Galaxias maculatus was measured in respirometry trials under different temperatures (5–21°C) and varying body masses (0.1–>1.5 g) spanning a relevant range of thermal conditions and sizes. Specific respiration rates (R in g O2 g−1 d−1) declined as a power function of body mass and increased exponentially with temperature and was expressed as: R = 0.0007 * W −0.31 * e 0.13 * T. The ability of this model to predict specific respiration rate was evaluated by comparing observed values with those predicted by the model. Our findings suggest that the respiration rate of G. maculatus is the result of multiple interactive processes (intrinsic and extrinsic factors) that modulate each other in ‘meta-mechanistic’ ways; this would help to explain the species’ ability to undergo the complex ontogenetic habitat shifts observed in the lakes of the Andean Patagonic range.

  12. Novel Infection Site and Ecology of Cryptic Didymocystis sp. (Trematoda) in the Fish Scomberomorus maculatus. (United States)

    Schrandt, Meagan N; Andres, Michael J; Powers, Sean P; Overstreet, Robin M


    An undescribed, cryptic species of Didymocystis, as determined from sequences of 2 ribosomal genes and superficially similar to Didymocystis scomberomori ( MacCallum and MacCallum, 1916 ), infected the skin of the Spanish mackerel, Scomberomorus maculatus , in the north-central Gulf of Mexico (GOM). An analysis of 558 fish from 2011 to 2013 from Louisiana, Mississippi, Alabama, and the Florida panhandle showed the prevalence of the trematode varied both spatially and temporally but not with sex of the fish host. Month, year, and geographic location were identified by a negative binomial generalized linear model as indicators of the abundance and intensity of infection. Prevalence, abundance, and intensity of infection were greatest in spring and fall months off the Florida panhandle. Furthermore, the abundance and intensity of infection correlated negatively with fork length, weight, and gonad weight of mature fish but positively with longitude. Therefore, smaller adult fish tended to be more infected than larger adults, and prevalence and intensity increased from west to east (Louisiana to Florida). Spatial and temporal trends seemed to result from physical factors (e.g., water temperature, salinity, bottom type), but they also coincided with the annual migration of S. maculatus as fish moved northward along the GOM coastline from the southern tip of Florida in the spring months and returned in the fall, being present in the north-central GOM from late spring through fall. This pattern suggests the possibility that acquisition of infections occurred from a molluscan host in waters off the Florida panhandle.

  13. New morphological data on Cucullanus pinnai pinnai (Nematoda parasitizing Pimelodus maculatus (Pimelodidae in southeastern Brazil

    Directory of Open Access Journals (Sweden)

    Vivian Suane de Freitas Vieira

    Full Text Available This paper describes the morphology of Cucullanus pinnai pinnai parasitizing Pimelodus maculatus in the Guandu River, Brazil, based on differential interference contrast (DIC microscopy and scanning electron microscopy (SEM, providing new morphological data about this species of parasite. Nematodes were collected between May and October 2012 from specimens of Pimelodus maculatus in the Guandu River (22°48’2”S, 43°37’35”W, in the state of Rio de Janeiro, Brazil. Some characteristics of specimens of Cucullanus in this study fall within the range of morphological variations of previously studied C. pinnai pinnai. Most of the specimens studied here had excretory pore and deirids located at the posterior end of the oesophagus, a feature not recorded in previous studies of this species. In addition, the size of the gubernaculum was larger than the other specimens previously studied. The SEM and DIC analyses of C. pinnai revealed several morphological details of the cephalic region and the tail papillae. With regard to the polymorphism of C. pinnai, morphological and genetic studies of this cucullanid nematode are needed, involving large numbers of host species and a wide geographical distribution.

  14. Biocontrol in store: spatial and behavioural aspects of foraging by Uscana lariophaga, egg parasitoid of Callosobruchus maculatus, in stored cowpea

    NARCIS (Netherlands)

    Stolk, C.


    Cowpea ( Vigna unguiculata Walpers), an important crop for West African subsistence farmers, is often infested in storage by the bruchid beetle Callosobruchus maculatus Fabricius. The indigenous egg parasitoid Uscana

  15. Toxicity and repellence of African plants traditionally used for the protection of stored cowpea against Callosobruchus maculatus

    NARCIS (Netherlands)

    Boeke, S.J.; Baumgart, I.R.; Loon, van J.J.A.; Huis, van A.; Dicke, M.; Kossou, D.K.


    In a search for botanical products to control the main insect pest of stored cowpea, Callosobruchus maculatus, 33 traditionally used African plants were tested in the laboratory for their toxic and repellent effects against this beetle. Toxicity was evaluated measuring life history parameters in a n

  16. Toxicity and repellence of African plants traditionally used for the protection of stored cowpea against Callosobruchus maculatus

    NARCIS (Netherlands)

    Boeke, S.J.; Baumgart, I.R.; Loon, van J.J.A.; Huis, van A.; Dicke, M.; Kossou, D.K.


    In a search for botanical products to control the main insect pest of stored cowpea, Callosobruchus maculatus, 33 traditionally used African plants were tested in the laboratory for their toxic and repellent effects against this beetle. Toxicity was evaluated measuring life history parameters in a

  17. Assessment of Digoxin-Specific Fab Fragment Dosages in Digoxin Poisoning. (United States)

    Nordt, Sean Patrick; Clark, Richard F; Machado, Carol; Cantrell, F Lee


    Digoxin poisoning still remains a common cause of morbidity and mortality. Fortunately, digoxin-specific Fab fragments are commercially available as an antidote. However, these Fab fragments are several thousand dollars per vial. There is a standardized formula to calculate appropriate Fab fragment dosage based on the serum digoxin concentration. This can greatly reduce the amount of Fab fragment administered. There is also an empiric dosing guideline recommending 6-10 vials be given; however, this may result in higher amounts of Fab fragments being administered than required. We performed this study to assess the amounts of digoxin-specific Fab fragments administered in the treatment of digoxin poisonings recorded in a poison control system database from January 1, 2000, to December 31, 2009, in which digoxin serum concentrations were available. This was a retrospective study of 278 patients, 107 with acute poisonings (group A) and 171 following chronic poisoning (group B). In group A, the calculated Fab dose was higher than the calculated dose based on available concentrations in 39 (36%) of group A and 15 (9%) of group B patients. The average wholesale price cost of the excessive dosages ranged from $4818 to as high as $50,589 per patient. Our data suggests that clinician education on digoxin poisoning and the use of the standardized formula to calculate the Fab dose may decrease over utilization and decrease costs associated with the administration of digoxin-specific Fab fragments in the treatment of digoxin poisonings.

  18. Complex Binding of the FabR Repressor of Bacterial Unsaturated Fatty Acid Biosynthesis to its Cognate Promoters


    Feng, Youjun; Cronan, John E.


    Two transcriptional regulators, the FadR activator and the FabR repressor control biosynthesis of unsaturated fatty acids in Escherichia coli. FabR represses expression of the two genes, fabA and fabB, required for unsaturated fatty acid synthesis and has been reported to require the presence of an unsaturated thioester (of either acyl carrier protein or CoA) in order to bind the fabA and fabB promoters in vitro. We report in vivo experiments in which unsaturated fatty acid synthesis was bloc...

  19. Apoptosis function on tumor cell by SEA-Fab' coupled protein%SWA-Fab'对肿瘤细胞的凋亡作用

    Institute of Scientific and Technical Information of China (English)

    舒晓刚; 王国斌


    目的研究SEA-Fab'对肿瘤细胞的凋亡作用及单克隆抗体于肿瘤定向治疗作用.方法实验分为三组:SEA-Fab'、SEA及对照组,三组肿瘤细胞分别用SEA-Fab'、SEA及PBMC+Walker-256细胞处理,在24~72 h内观察肿瘤细胞的凋亡情况.结果SEA-Fab'组、SEA组的肿瘤细胞指数分别为(34.6%~68.9%)和(15.5%~31.9%)高于对照组(5.5%~12.5%),差异存在显著性,SEA-Fab'组高于SEA组差异显著.结论SEA-Fab'在激活T细胞和杀死肿瘤细胞效果好于SEA,将来有可能利用单克隆抗体来作定向靶位治疗.%[Objective] To Study the anti-tumor effect of the SEA-Fab' coupled protein and the possibility of themonoclonal antibody Fab's targeted in immunotherapy of human tumor. [Methods] The experiment group was treat-ed by SEA-Fab' and SEA, the contrast group was treated by PBMC+Walker-256 cell. The apoptotic index was ob-served in 24~72 h. [Results] The poptotic index was significantly higher in the SEA-Fab's (34.6~68.9%) and SEAgroups (15.5~31.9%) compared with the PBMC+Wallker-256 cell group (5.5~12.8%). There was significant differ-ence between SEA-Fab group and SEA group (P<0.01). [Conclusion] SEA-Fab' is more effective to activate Tcells and kill tumor cell than SEA, there is ossibility of The monoclonal antibody targeted in immunotherapy of hu-man tumor.

  20. Structural Characterisation of the Beta-Ketoacyl-Acyl Carrier Protein Synthases, FabF and FabH, of Yersinia pestis


    Jeffrey D. Nanson; Himiari, Zainab; Swarbrick, Crystall M. D.; Forwood, Jade K.


    Yersinia pestis, the causative agent of bubonic, pneumonic, and septicaemic plague, remains a major public health threat, with outbreaks of disease occurring in China, Madagascar, and Peru in the last five years. The existence of multidrug resistant Y. pestis and the potential of this bacterium as a bioterrorism agent illustrates the need for new antimicrobials. The β-ketoacyl-acyl carrier protein synthases, FabB, FabF, and FabH, catalyse the elongation of fatty acids as part of the type II f...

  1. Neutron Reflection Study of Surface Adsorption of Fc, Fab, and the Whole mAb. (United States)

    Li, Zongyi; Li, Ruiheng; Smith, Charles; Pan, Fang; Campana, Mario; Webster, John R P; van der Walle, Christopher F; Uddin, Shahid; Bishop, Steve M; Narwal, Rojaramani; Warwicker, Jim; Lu, Jian Ren


    Characterizing the influence of fragment crystallization (Fc) and antigen-binding fragment (Fab) on monoclonal antibody (mAb) adsorption at the air/water interface is an important step to understanding liquid mAb drug product stability during manufacture, shipping, and storage. Here, neutron reflection is used to study the air/water adsorption of a mAb and its Fc and Fab fragments. By varying the isotopic contrast, the adsorbed amount, thickness, orientation, and immersion of the adsorbed layers could be determined unambiguously. While Fc adsorption reached saturation within the hour, its surface adsorbed amount showed little variation with bulk concentration. In contrast, Fab adsorption was slower and the adsorbed amount was concentration dependent. The much higher Fc adsorption, as compared to Fab, was linked to its lower surface charge. Time and concentration dependence of mAb adsorption was dominated by Fab behavior, although both Fab and Fc behaviors contributed to the amount of mAb adsorbed. Changing the pH from 5.5 to 8.8 did not much perturb the adsorbed amount of Fc, Fab, or mAb. However, a small decrease in adsorption was observed for the Fc over pH 8-8.8 and vice versa for the Fab and mAb, consistent with a dominant Fab behavior. As bulk concentration increased from 5 to 50 ppm, the thicknesses of the Fc layers were almost constant at 40 Å, while Fab and mAb layers increased from 45 to 50 Å. These results imply that the adsorbed mAb, Fc, and Fab all retained their globular structures and were oriented with their short axial lengths perpendicular to the interface.

  2. An improved single-chain Fab platform for efficient display and recombinant expression. (United States)

    Koerber, James T; Hornsby, Michael J; Wells, James A


    Antibody phage display libraries combined with high-throughput selections have recently demonstrated tremendous promise to create the next generation of renewable, recombinant antibodies to study proteins and their many post-translational modification states; however, many challenges still remain, such as optimized antibody scaffolds. Recently, a single-chain fragment antigen binding (Fab) (scFab) format, in which the carboxy-terminus of the light chain is linked to the amino-terminus of the heavy chain, was described to potentially combine the high display levels of a single-chain fragment variable with the high stability of purified Fabs. However, this format required removal of the interchain disulfide bond to achieve modest display levels and subsequent bacterial expression resulted in high levels of aggregated scFab, hindering further use of scFabs. Here, we developed an improved scFab format that retains the interchain disulfide bond by increasing the linker length between the light and heavy chains to improve display and bacterial expression levels to 1-3 mg/L. Furthermore, rerouting of the scFab to the co-translational signal recognition particle pathway combined with reengineering of the signal peptide sequence results in display levels 24-fold above the original scFab format and 3-fold above parent Fab levels. This optimized scFab scaffold can be easily reformatted in a single step for expression in a bacterial or mammalian host to produce stable (Tm of 81 °C), predominantly monomeric (>90%) antibodies at a high yield. Ultimately, this new scFab format will advance high-throughput antibody generation platforms to discover the next generation of research and therapeutic antibodies.

  3. Crystal structure determination of anti-DNA Fab A52. (United States)

    Stanfield, Robyn L; Eilat, Dan


    A52 is a murine monoclonal antibody isolated from autoimmune New Zealand Black/New Zealand White F1 mice that recognizes single and double stranded DNA. This mouse strain spontaneously develops systemic lupus erythematosus-like symptoms and has served as a model for that disease for many years. The 1.62 Å crystal structure of the A52 Fab fragment reveals an H3 complementarity determining region with four closely spaced arginine residues, creating a positively charged surface to accommodate bound DNA.

  4. Generation and characterization of the human neutralizing antibody fragment Fab091 against rabies virus

    Institute of Scientific and Technical Information of China (English)

    Chen LI; Feng ZHANG; Hong LIN; Zhong-can WANG; Xin-jian LIU; Zhen-qing FENG; Jin ZHU; Xiao-hong GUAN


    Aim: To transform the human anti-rabies virus glycoprotein (anti-RABVG) single-chain variable fragment (scFv) into a Fab fragment and to analyze its immunological activity.Methods: The Fab gene was amplified using overlap PCR and inserted into the vector pComb3XSS. The recombinant vector was then transformed into E coli Top10F' for expression and purification. The purified Fab was characterized using SDS-PAGE, Western blotting,indirect ELISA, competitive ELISA, and the fluorescent antibody virus neutralization test (FAVN), respectively, and examined in a Kunming mouse challenge model in vivo.Results: A recombinant vector was constructed. The Fab was expressed in soluble form In E coll Top10F'. Specific binding of the Fab to rabies virus was confirmed by indirect ELISA and immunoprecipitation (IP). The neutralizing antibody titer of Fab was 10.26 IU/mL.The mouse group treated with both vaccine and human rabies immunoglobulin (HRIG)/Fab091 (32 IU/kg) showed protection against rabies, compared with the control group (P<0.05, Logrank test).Conclusion: The antibody fragment Fab was shown to be a neutralizing antibody against RABVG. It can be used together with other monoclonal antibodies for post-exposure prophylaxis of rabies virus in future studies.

  5. Functionally Approached Body (FAB) Strategies for Young Children Who Have Behavioral and Sensory Processing Challenges (United States)

    Pagano, John


    Functionally Approached Body (FAB) Strategies offer a clinical approach to help parents of young children with behavioral and sensory processing strategies. This article introduces the FAB Strategies, clinical strategies developed by the author for understanding and addressing young children's behavioral and sensory processing challenges. The FAB…

  6. Phage-display libraries of murine and human antibody Fab fragments

    DEFF Research Database (Denmark)

    Engberg, J; Andersen, P S; Nielsen, L K


    We provide efficient and detailed procedures for construction, expression, and screening of comprehensive libraries of murine or human antibody Fab fragments displayed on the surface of filamentous phage. In addition, protocols for producing and using ultra-electrocompetent cells, for producing Fab...

  7. Isolation of Osteosarcoma-Associated Human Antibodies from a Combinatorial Fab Phage Display Library

    Directory of Open Access Journals (Sweden)

    Carmela Dantas-Barbosa


    Full Text Available Osteosarcoma, a highly malignant disease, is the most common primary bone tumor and is frequently found in children and adolescents. In order to isolate antibodies against osteosarcoma antigens, a combinatorial osteosarcoma Fab library displayed on the surface of phages was used. After three rounds of selection on the surface of tumor cells, several osteosarcoma-reactive Fabs were detected. From these Fabs, five were better characterized, and despite having differences in their VH (heavy chain variable domain and Vκ (kappa chain variable domain regions, they all bound to a protein with the same molecular mass. Further analysis by cell ELISA and immunocytochemistry suggested that the Fabs recognize a membrane-associated tumor antigen expressed in higher amounts in neoplasic cells than in normal tissue. These results suggest that the human Fabs selected in this work are a valuable tool for the study of this neoplasia.

  8. The Emerging Importance of IgG Fab Glycosylation in Immunity. (United States)

    van de Bovenkamp, Fleur S; Hafkenscheid, Lise; Rispens, Theo; Rombouts, Yoann


    Human IgG is the most abundant glycoprotein in serum and is crucial for protective immunity. In addition to conserved IgG Fc glycans, ∼15-25% of serum IgG contains glycans within the variable domains. These so-called "Fab glycans" are primarily highly processed complex-type biantennary N-glycans linked to N-glycosylation sites that emerge during somatic hypermutation. Specific patterns of Fab glycosylation are concurrent with physiological and pathological conditions, such as pregnancy and rheumatoid arthritis. With respect to function, Fab glycosylation can significantly affect stability, half-life, and binding characteristics of Abs and BCRs. Moreover, Fab glycans are associated with the anti-inflammatory activity of IVIgs. Consequently, IgG Fab glycosylation appears to be an important, yet poorly understood, process that modulates immunity.

  9. The genome of the platyfish, Xiphophorus maculatus, provides insights into evolutionary adaptation and several complex traits. (United States)

    Schartl, Manfred; Walter, Ronald B; Shen, Yingjia; Garcia, Tzintzuni; Catchen, Julian; Amores, Angel; Braasch, Ingo; Chalopin, Domitille; Volff, Jean-Nicolas; Lesch, Klaus-Peter; Bisazza, Angelo; Minx, Pat; Hillier, LaDeana; Wilson, Richard K; Fuerstenberg, Susan; Boore, Jeffrey; Searle, Steve; Postlethwait, John H; Warren, Wesley C


    Several attributes intuitively considered to be typical mammalian features, such as complex behavior, live birth and malignant disease such as cancer, also appeared several times independently in lower vertebrates. The genetic mechanisms underlying the evolution of these elaborate traits are poorly understood. The platyfish, X. maculatus, offers a unique model to better understand the molecular biology of such traits. We report here the sequencing of the platyfish genome. Integrating genome assembly with extensive genetic maps identified an unexpected evolutionary stability of chromosomes in fish, in contrast to in mammals. Genes associated with viviparity show signatures of positive selection, identifying new putative functional domains and rare cases of parallel evolution. We also find that genes implicated in cognition show an unexpectedly high rate of duplicate gene retention after the teleost genome duplication event, suggesting a hypothesis for the evolution of the behavioral complexity in fish, which exceeds that found in amphibians and reptiles.

  10. Food Characterization of the Spotted Pimelodid Fish Pimelodus maculatus from a Polluted Urban River in Argentina

    Directory of Open Access Journals (Sweden)

    David Kuczynski


    Full Text Available On the present study, the nourishment and the digestive system of a population of the spotted pimelodid Pimelodus maculatus Lacepède 1803 from a polluted urban river in Argentina were analyzed (Reconquista River, Buenos Aires province. The specimens are characterized by having benthic habits and an omnivore diet. In their intestinal content, a diversity of components was identified. Such components were grouped into ten alimentary items, with dietary variations according to body size while the longitude of the intestine remained proportional to standard length. Microbiological analysis in the gills, stomach and intestine showed the occurrence of Staphylococcus aureus, Escherichia coli and Salmonella sp. Fungi and yeast were also detected. Bacterial accounts were very high both in the analyzed organs as well in water samples. The ecological and sanitary significance of the identified bacteria is discussed.

  11. Genetic characterization of the Neotropical catfish Pimelodus maculatus (Pimelodidae, Siluriformes) in the Upper Uruguay River (United States)

    Ribolli, Josiane; de Melo, Cláudio Manoel Rodrigues; Zaniboni-Filho, Evoy


    Freshwater fish present unique challenges when one attempts to understand the factors that determine the structure of their populations. Habitat fragmentation is a leading cause of population decline that threatens ecosystems worldwide. In this study, we investigated the conservation status of genetic variability in the Neotropical catfish (Pimelodus maculatus). Specifically, we examined the structure and genetic diversity of this species in a region of the Upper Uruguay River fragmented by natural barriers and dams. There was no genetic structure among the four sites analyzed, indicating the existence of only one population group. A combination of environmental management and genetic monitoring should be used to minimize the impact of impoundment on panmitic populations of migratory fish species. PMID:23271936

  12. 75 FR 21353 - Intel Corporation, Fab 20 Division, Including On-Site Leased Workers From Volt Technical... (United States)


    ... Employment and Training Administration Intel Corporation, Fab 20 Division, Including On-Site Leased Workers... for Worker Adjustment Assistance on March 10, 2010, applicable to workers of Intel Corporation, Fab 20... the Hillsboro, Oregon location of Intel Corporation, Fab 20 Division. The Department has...

  13. Characterization of deamidation at Asn138 in L-chain of recombinant humanized Fab expressed from Pichia pastoris. (United States)

    Ohkuri, Takatoshi; Murase, Eri; Sun, Shu-Lan; Sugitani, Jun; Ueda, Tadashi


    A method was previously established for evaluating Asn deamidation by matrix-assisted laser desorption/ionization time of flight-mass spectrometry using endoproteinase Asp-N. In this study, we demonstrated that this method could be applied to the identification of the deamidation site of the humanized fragment antigen-binding (Fab). First, a system for expressing humanized Fab from methylotrophic yeast Pichia pastoris was constructed, resulting in the preparation of ∼30 mg of the purified humanized Fab from 1 l culture. Analysis of the L-chain derived from recombinant humanized Fab that was heated at pH 7 and 100°C for 1 h showed the deamidation at Asn138 in the constant region. Then, we prepared L-N138D Fab and L-N138A Fab and examined their properties. The circular dichroism (CD) spectrum of the L-N138D Fab was partially different from that of the wild-type Fab. The measurement of the thermostability showed that L-N138D caused a significant decrease in the thermostability of Fab. On the other hand, the CD spectrum and thermostability of L-N138A Fab showed the same behaviour as the wild-type Fab. Thus, it was suggested that the introduction of a negative charge at position 138 in the L-chain by the deamidation significantly affected the stability of humanized Fab.

  14. 20 CFR 30.320 - Can a claim be reopened after the FAB has issued a final decision? (United States)


    ... 20 Employees' Benefits 1 2010-04-01 2010-04-01 false Can a claim be reopened after the FAB has... AMENDED Adjudicatory Process Reopening Claims § 30.320 Can a claim be reopened after the FAB has issued a final decision? (a) At any time after the FAB has issued a final decision pursuant to § 30.316,...

  15. Tibet Orbivirus, a novel Orbivirus species isolated from Anopheles maculatus mosquitoes in Tibet, China.

    Directory of Open Access Journals (Sweden)

    Minghua Li

    Full Text Available BACKGROUND: The genus Orbivirus includes a number of important pathogenic viruses, including Bluetongue virus (BTV, African horse sickness virus (AHSV, and Epizootic hemorrhagic disease virus (EHDV. In this study we describe the isolation and characterization of an Orbivirus strain isolated from Anopheles maculatus mosquitoes collected in Tibet, China. METHODS AND RESULTS: Initial viral screening identified a viral strain (XZ0906 that caused significant cytopathic effect (CPE in BHK-21 cells, including rounding, cell rupture, and floating. Although CPE was not observed in insect cells (C6/36, these cells supported viral replication. Polyacrylamide gel analysis revealed a genome consisting of 10 segments of double-stranded RNA (dsRNA, with a distribution pattern of 3-3-3-1. 454 high throughput sequencing of culture supernatant was used for viral identification. Complete genome sequencing was performed by Sanger sequencing in combination with 5'-RACE and 3'-RACE. Sequence analysis demonstrated that all 5'- and 3'- untranslated regions (UTRs for each of the 10 genome segments contained a series of six highly conserved nucleotides. In addition, homology analysis and phylogenetic analysis based on amino acid sequence was completed, and all results show that virus XZ0906 was not a member of any known species or serotype of Orbivirus, indicating it to be a new species within the genus Orbivirus. CONCLUSIONS: The isolated Orbivirus strain was designated Tibet Orbivirus, TIBOV to denote the location from which it was isolated. TIBOV is a novel orbivirus species which is isolated from Anopheles maculatus mosquitoes collected in Tibet, China.

  16. Isolation Driven Divergence in Osmoregulation in Galaxias maculatus (Jenyns, 1848 (Actinopterygii: Osmeriformes.

    Directory of Open Access Journals (Sweden)

    Ignacio Ruiz-Jarabo

    Full Text Available Marine species have colonized extreme environments during evolution such as freshwater habitats. The amphidromous teleost fish, Galaxias maculatus is found mainly migrating between estuaries and rivers, but some landlocked populations have been described in lakes formed during the last deglaciation process in the Andes. In the present study we use mtDNA sequences to reconstruct the historical scenario of colonization of such a lake and evaluated the osmoregulatory shift associated to changes in habitat and life cycle between amphidromous and landlocked populations.Standard diversity indices including the average number of nucleotide differences (Π and the haplotype diversity index (H indicated that both populations were, as expected, genetically distinctive, being the landlocked population less diverse than the diadromous one. Similarly, pairwise GST and NST comparison detected statistically significant differences between both populations, while genealogy of haplotypes evidenced a recent founder effect from the diadromous stock, followed by an expansion process in the lake. To test for physiological differences, individuals of both populations were challenged with a range of salinities from 0 to 30 ppt for 8 days following a period of progressive acclimation. The results showed that the landlocked population had a surprisingly wider tolerance to salinity, as landlocked fish survival was 100% from 0 to 20 ppt, whereas diadromous fish survival was 100% only from 10 to 15 ppt. The activity of ATPase enzymes, including Na+/K+-ATPase (NKA, and H+-ATPase (HA was measured in gills and intestine. Activity differences were detected between the populations at the lowest salinities, including differences in ATPases other than NKA and HA. Population differences in mortality are not reflected in enzyme activity differences, suggesting divergence in other processes.These results clearly demonstrate the striking adaptive changes of G. maculatus

  17. Fab-based bispecific antibody formats with robust biophysical properties and biological activity. (United States)

    Wu, Xiufeng; Sereno, Arlene J; Huang, Flora; Lewis, Steven M; Lieu, Ricky L; Weldon, Caroline; Torres, Carina; Fine, Cody; Batt, Micheal A; Fitchett, Jonathan R; Glasebrook, Andrew L; Kuhlman, Brian; Demarest, Stephen J


    A myriad of innovative bispecific antibody (BsAb) platforms have been reported. Most require significant protein engineering to be viable from a development and manufacturing perspective. Single-chain variable fragments (scFvs) and diabodies that consist only of antibody variable domains have been used as building blocks for making BsAbs for decades. The drawback with Fv-only moieties is that they lack the native-like interactions with CH1/CL domains that make antibody Fab regions stable and soluble. Here, we utilize a redesigned Fab interface to explore 2 novel Fab-based BsAbs platforms. The redesigned Fab interface designs limit heavy and light chain mixing when 2 Fabs are co-expressed simultaneously, thus allowing the use of 2 different Fabs within a BsAb construct without the requirement of one or more scFvs. We describe the stability and activity of a HER2×HER2 IgG-Fab BsAb, and compare its biophysical and activity properties with those of an IgG-scFv that utilizes the variable domains of the same parental antibodies. We also generated an EGFR × CD3 tandem Fab protein with a similar format to a tandem scFv (otherwise known as a bispecific T cell engager or BiTE). We show that the Fab-based BsAbs have superior biophysical properties compared to the scFv-based BsAbs. Additionally, the Fab-based BsAbs do not simply recapitulate the activity of their scFv counterparts, but are shown to possess unique biological activity.

  18. Comparing domain interactions within antibody Fabs with kappa and lambda light chains. (United States)

    Toughiri, Raheleh; Wu, Xiufeng; Ruiz, Diana; Huang, Flora; Crissman, John W; Dickey, Mark; Froning, Karen; Conner, Elaine M; Cujec, Thomas P; Demarest, Stephen J


    IgG antibodies are multi-domain proteins with complex inter-domain interactions. Human IgG heavy chains (HCs) associate with light chains (LCs) of the κ or λ isotype to form mature antibodies capable of binding antigen. The HC/LC interaction involves 4 domains: VH and CH1 from the HC and VL and CL from the LC. Human Fabs with κ LCs have been well characterized for their unfolding behaviors and demonstrate a significant level of cooperativity and stabilization when all 4 domains are intact. Very little is known regarding the thermodynamic properties of human Fabs with λ LCs. Here, we dissect the domain contributions to Fab stability for both κ and λ LC-containing Fabs. We find the cooperativity of unfolding between the constant domains, CH1/Cλ, and variable domains, VH/Vλ, within λ LC-containing Fabs is significantly weaker than that of κ LC-containing Fabs. The data suggests there may not be an evolutionary necessity for strong variable/constant domain cooperativity within λ LC-containing Fabs. After investigating the biophysical properties of Fabs with mismatched variable and constant domain subunits (e.g., VH/Vκ paired with CH1/Cλ or T cell receptor Cα/Cβ), the major role of the constant domains for both κ- and λ-containing Fabs may be to reduce the hydrophobic exposure at the VH/VL interface. Even though Fabs with these non-native pairings were thermodynamically less stable, they secreted well from mammalian cells as well behaved monodisperse proteins, which was in contrast to what was observed with the VH/Vκ and VH/Vλ scFvs that secreted as a mixture of monomer and aggregates.

  19. Rocinela signata (Isopoda: Aegidae parasitizing the gills of the spotted goatfish Pseudupeneus maculatus (Actinopterygii: Mullidae in Northeastern Brazil

    Directory of Open Access Journals (Sweden)


    Full Text Available ABSTRACT Parasitic isopods of fishes usually show wide distribution and low host specificity. This study investigated the occurrence of gill parasites in 120 specimens of spotted goatfish Pseudupeneus maculatus, marine fish of great economic importance for fishery community. The fish were captured monthly in the Coast of Pernambuco State, Northeast Brazil, between October 2012 and September 2013. Eleven isopods were found and identified as Rocinela signata. We observed lower infections in the gills of P. maculatus (prevalence 8.3%, mean intensity 1.6±0.3 and mean abundance 0.1±0.3 when compared to other studies of different host fishes from geographically close locations. This parasite has been reported from a wide variety of fish species, not only in Brazil, but also in the eastern Atlantic and the Pacific oceans. This is the first report of R. signata in spotted goatfish.

  20. Enzymatic Activity in the Gastrointestinal Tract of Pimelodus maculatus (Teleostei, Siluriformes) in Two Neotropical Reservoirs with Different Trophic Conditions


    Silvana Duarte; Marcelo Bemquerer; Francisco Gerson de Araújo


    Enzymatic activities for digestion of proteins and carbohydrates were compared among three organs of the digestive system of Pimelodus maculatus in two reservoirs with different trophic conditions during the winter of 2006. The aim was to test the hypothesis that enzymatic activity for the digestion of proteins and carbohydrates differed among organs and that such activities differ between the trophic state of the environment. Enzymatic activities were determined through the assays of specifi...

  1. sup 111 In-antimyosin Fab scintigraphy in cardiovascular diseases; Multicenter clinical trial

    Energy Technology Data Exchange (ETDEWEB)

    Kawai, Chuichi; Matsumori, Akira; Endo, Keigo (Kyoto Univ. (Japan). Faculty of Medicine); Nishimura, Tsunehiko


    In a multicenter study, a total of 380 patients with myocardial infarction, myocarditis and cardiomyopathy underwent {sup 111}In-antimyosin Fab myocardial imaging. {sup 111}In-antimyosin Fab was administered intravenously and myocardial images were obtained 48 hours later. Only 3 patients developed mild adverse effects. Human antimouse antibodies were detected in 7 patients. Positive scans in patients with myocardial infarction were seen in 92/119 (77%) within 2 weeks after the onset of myocardial infarction, in 58/71 (82%) at 3-4 weeks, in 20/22 (91%) at 4-8 weeks and 17/31 (55%) thereafter. The location of myocardial damage delineated by {sup 111}In-antimyosin Fab imaging was concordant with the infarct location by ECG and coronary angiography. In patients with myocarditis, {sup 111}In-antimyosin Fab uptake was positive in 7/12 (58%) within 8 weeks and 6/17 (35%) thereafter. Positive {sup 111}In-antimyosin Fab scans were seen in 12/36 (33%) in dilated cardiomyopathy and in 17/19 (89%) in hypertrophic cardiomyopathy. Although the mechanism of persistently positive {sup 111}In-antimyosin Fab images in the subacute to chronic stage of myocardial infarction and myocarditis remains to be clarified, {sup 111}In-antimyosin Fab may be useful for the detection of the disease and in evaluating the prognosis of patients with cardiomyopathy. (author).

  2. Recombinant human Fab fragments neutralize human type 1 immunodeficiency virus in vitro. (United States)

    Barbas, C F; Björling, E; Chiodi, F; Dunlop, N; Cababa, D; Jones, T M; Zebedee, S L; Persson, M A; Nara, P L; Norrby, E


    A panel of 20 recombinant Fab fragments reactive with the surface glycoprotein gp120 of human type 1 immunodeficiency virus (HIV-1) were examined for their ability to neutralize MN and IIIB strains of the virus. Neutralization was determined as the ability of the Fab fragments to inhibit infection as measured in both a p24 ELISA and a syncytium-formation assay. One group of closely sequence-related Fab fragments was found to neutralize virus in both assays with a 50% neutralization titer at approximately 1 micrograms/ml. Another Fab neutralized in the p24 ELISA but not in the syncytium assay. The other Fab fragments showed weak or no neutralizing ability. The results imply that virion aggregation or crosslinking of gp120 molecules on the virion surface is not an absolute requirement for HIV-1 neutralization. Further, all of the Fab fragments were shown to be competitive with soluble CD4 for binding to gp120 and yet few neutralized the virus effectively, implying that the mechanism of neutralization in this case may not involve receptor blocking. The observation of a preponderance of high-affinity Fab fragments with poor or no neutralizing ability could have implications for vaccine strategies. PMID:1384050

  3. Modification of fibrin network ultrastructure by Fab fragments specific for different domain of fibrinogen. (United States)

    Cierniewski, C S; Janiak, A; Wyroba, E


    Kinetics of inhibition of fibrin monomer polymerization produced by Fab fragments prepared from immunochemically purified monospecific antibodies to the surface epitopes of different domains of fibrinogen molecule has been correlated with electron microscopic observations of resulting specimens. Fab fragments prepared from anti FgD antisera were the most efficient inhibitors of thrombin-catalysed conversion of fibrinogen to fibrin; polymerization of fibrin monomers as detected spectrophotometrically was abolished at 2:1 molar ratio of anti FgD Fab fragments to fibra monomer. These Fab fragments acting as a steric hindrance of polymerization sites inhibited the first stage of fibrin monomer aggregation. Interaction of Fab fragments derived from antibodies specific for alpha 239-476 with corresponding segment of fibrinogen molecule resulted in a weak inhibition of fibrin monomer polymerization. However, fibrin obtained in the presence of these Fab fragments was significantly modified and showed no periodicity. This observation may suggest that anti alpha 239-476 Fab impaired the course of the second stage of fibrin monomer polymerization, i.e. lateral association of fibrin fibrils.

  4. Isolation of human anti-serum albumin Fab antibodies with an extended serum-half life. (United States)

    Kang, Hyeon-Ju; Kim, Hye-Jin; Cha, Sang-Hoon


    The serum albumin (SA) has been exploited to generate long-acting biotherapeutics by taking advantage of the FcRn-mediated recycling mechanism in a direct or an indirect way. Since Fab fragments have been proven to be clinically safe for human usage, we assumed that human anti-SA Fab antibodies could have a great potential as a carrier molecule to extend the serum half-life of therapeutic proteins. We, herein, had attempted to isolate anti-SA Fab antibodies from HuDVFab-8L antibody library via a phage display technology, and identified eight discrete human Fab antibodies. One of the Fab antibodies, SL335, showed the strongest binding reactivity to human SA with nM range of affinity at both pH 6 and pH 7.4, and cross-reacted to SAs from various species including rat, mouse, canine and monkey. The in vivo pharmacokinetic assay using a rat model indicated that SL335 has approximately 10 fold longer serum half-life and 26 to 44-fold increase in AUC0 → ∞ compared to the negative control Fab molecule in both intravenous and subcutaneous administrations. Knowing that Fabs have proven to be safe in clinics for a long time, SL335 seems to have a great potential in generating long-acting protein drugs by tagging effector molecules with either chemical conjugation or genetic fusion.

  5. 'Zipbody' leucine zipper-fused Fab in E. coli in vitro and in vivo expression systems. (United States)

    Ojima-Kato, Teruyo; Fukui, Kansuke; Yamamoto, Hiroaki; Hashimura, Dai; Miyake, Shiro; Hirakawa, Yuki; Yamasaki, Tomomi; Kojima, Takaaki; Nakano, Hideo


    A small antibody fragment, fragment of antigen binding (Fab), is favorable for various immunological assays. However, production efficiency of active Fab in microorganisms depends considerably on the clones. In this study, leucine zipper-peptide pairs that dimerize in parallel (ACID-p1 (LZA)/BASE-p1 (LZB) or c-Jun/c-Fos) were fused to the C-terminus of heavy chain (Hc, VH-CH1) and light chain (Lc, VL-CL), respectively, to accelerate the association of Hc and Lc to form Fab in Escherichia coli in vivo and in vitro expression systems. The leucine zipper-fused Fab named 'Zipbody' was constructed using anti-E. coli O157 monoclonal antibody obtained from mouse hybridoma and produced in both in vitro and in vivo expression systems in an active form, whereas Fab without the leucine zipper fusion was not. Similarly, Zipbody of rabbit monoclonal antibody produced in in vitro expression showed significant activity. The purified, mouse Zipbody produced in the E. coli strain Shuffle T7 Express had specificity toward the antigen; in bio-layer interferometry analysis, the KD value was measured to be 1.5-2.0 × 10(-8) M. These results indicate that leucine zipper fusion to Fab C-termini markedly enhances active Fab formation in E. coli.

  6. Immobilization of Fab' fragments onto substrate surfaces: A survey of methods and applications. (United States)

    Crivianu-Gaita, Victor; Thompson, Michael


    Antibody immobilization onto surfaces has widespread applications in many different fields. It is desirable to bind antibodies such that their fragment-antigen-binding (Fab) units are oriented away from the surface in order to maximize analyte binding. The immobilization of only Fab' fragments yields benefits over the more traditional whole antibody immobilization technique. Bound Fab' fragments display higher surface densities, yielding a higher binding capacity for the analyte. The nucleophilic sulfide of the Fab' fragments allows for specific orientations to be achieved. For biosensors, this indicates a higher sensitivity and lower detection limit for a target analyte. The last thirty years have shown tremendous progress in the immobilization of Fab' fragments onto gold, Si-based, polysaccharide-based, plastic-based, magnetic, and inorganic surfaces. This review will show the current scope of Fab' immobilization techniques available and illustrate methods employed to minimize non-specific adsorption of undesirables. Furthermore, a variety of examples will be given to show the versatility of immobilized Fab' fragments in different applications and future directions of the field will be addressed, especially regarding biosensors.

  7. Structural features of Fab fragments of rheumatoid factor IgM-RF in solution

    Energy Technology Data Exchange (ETDEWEB)

    Volkov, V. V., E-mail: [Russian Academy of Sciences, Shubnikov Institute of Crystallography (Russian Federation); Lapuk, V. A. [Russian Academy of Sciences, Zelinskii Institute of Organic Chemistry (Russian Federation); Shtykova, E. V.; Stepina, N. D.; Dembo, K. A.; Sokolova, A. V.; Amarantov, S. V. [Russian Academy of Sciences, Shubnikov Institute of Crystallography (Russian Federation); Timofeev, V. P. [Russian Academy of Sciences, Engelhardt Institute of Molecular Biology (Russian Federation); Ziganshin, R. Kh. [Russian Academy of Sciences, Shemyakin Ovchinnikov Institute of Bioorganic Chemistry (Russian Federation); Varlamova, E. Yu. [Russian Academy of Medical Sciences, Hematology Research Center (Russian Federation)


    The structural features of the Fab fragments of monoclonal (Waldenstroem's disease) immunoglobulin M (IgM) and rheumatoid immunoglobulin M (IgM-RF) were studied by a complex of methods, including small-angle X-ray scattering (SAXS), electron spin resonance (ESR), and mass spectrometry (MS). The Fab-RF fragment was demonstrated to be much more flexible in the region of interdomain contacts, the molecular weights and the shapes of the Fab and Fab-RF macromolecules in solution being only slightly different. According to the ESR data, the rotational correlation time for a spin label introduced into the peptide sequence for Fab is twice as large as that for Fab-RF (21{+-}2 and 11{+-}1 ns, respectively), whereas the molecular weights of these fragments differ by only 0.5% (mass-spectrometric data), which correlates with the results of molecular-shape modeling by small-angle X-ray scattering. The conclusion about the higher flexibility of the Fab-RF fragment contributes to an understanding of the specificity of interactions between the rheumatoid factor and the antigens of the own organism.

  8. The community FabLab platform: applications and implications in biomedical engineering. (United States)

    Stephenson, Makeda K; Dow, Douglas E


    Skill development in science, technology, engineering and math (STEM) education present one of the most formidable challenges of modern society. The Community FabLab platform presents a viable solution. Each FabLab contains a suite of modern computer numerical control (CNC) equipment, electronics and computing hardware and design, programming, computer aided design (CAD) and computer aided machining (CAM) software. FabLabs are community and educational resources and open to the public. Development of STEM based workforce skills such as digital fabrication and advanced manufacturing can be enhanced using this platform. Particularly notable is the potential of the FabLab platform in STEM education. The active learning environment engages and supports a diversity of learners, while the iterative learning that is supported by the FabLab rapid prototyping platform facilitates depth of understanding, creativity, innovation and mastery. The product and project based learning that occurs in FabLabs develops in the student a personal sense of accomplishment, self-awareness, command of the material and technology. This helps build the interest and confidence necessary to excel in STEM and throughout life. Finally the introduction and use of relevant technologies at every stage of the education process ensures technical familiarity and a broad knowledge base needed for work in STEM based fields. Biomedical engineering education strives to cultivate broad technical adeptness, creativity, interdisciplinary thought, and an ability to form deep conceptual understanding of complex systems. The FabLab platform is well designed to enhance biomedical engineering education.

  9. Hepatic targeting and hypocholesterolemic effect of lactosaminated Fab against low density lipoproteins

    Energy Technology Data Exchange (ETDEWEB)

    Bernini, F.; Bocan, T.M.A.; Via, D.P.; Gotto, A.M. Jr.; Smith, L.C.


    Lactosaminated Fab (lac-Fab) specific for human LDL induces plasma clearance and uptake of circulating (/sup 125/-I)-iodo-LDL in rat, a process mediated by galactose receptors of the liver. This study demonstrates that lac-Fab is a specific carrier of LDL to the liver parenchymal cells and exhibits hypocholesterolemic activity in vivo. Rats were injected with fluorescent 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine-LDL (diI-LDL) or 6 mg of LDL plus tracer amounts of (/sup 125/I)-iodo-LDL. After 10-20 min, the animals received 3-10 mg of lac-Fab. Histologic examination of the liver sections showed the uptake of diI-LDL in the parenchymal cells, as compared to diI-acetyl-LDL which was localized in sinusoidal cells. More than 85% of human LDL disappeared within 2.5 hr after lac-Fab injection, reducing plasma cholesterol from 133.0 +/- 12.6 mg/dl to 66.4 +/- 8.0 mg/dl, the basal value in the rat. In control rats, only about 20% of radioactivity and cholesterol disappeared at 2.5 hr. HDL levels were unaffected. The authors conclude that lac-Fab is a specific carrier of LDL to hepatocytes and can lower plasma LDL-cholesterol in vivo. Lac-Fab specific for other antigens may act as specific carriers of molecule or macromolecules to hepatocytes.

  10. Single-reagent one-step procedures for the purification of ovine IgG, F(ab')2 and Fab antivenoms by caprylic acid. (United States)

    Al-Abdulla, Ibrahim; Casewell, Nicholas R; Landon, John


    Antivenoms are typically produced in horses or sheep and often purified using salt precipitation of immunoglobulins or F(ab')2 fragments. Caprylic (octanoic) acid fractionation of antiserum has the advantage of not precipitating the desired antibodies, thereby avoiding potential degradation that can lead to the formation of aggregates, which may be the cause of some adverse reactions to antivenoms. Here we report that when optimising the purification of immunoglobulins from ovine antiserum raised against snake venom, caprylic acid was found to have no effect on the activity of the enzymes pepsin and papain, which are employed in antivenom manufacturing to digest immunoglobulins to obtain F(ab')2 and Fab fragments, respectively. A "single-reagent" method was developed for the production of F(ab')2 antivenom whereby whole ovine antiserum was mixed with both caprylic acid and pepsin and incubated for 4h at 37°C. For ovine Fab antivenom production from whole antiserum, the "single reagent" comprised of caprylic acid, papain and l-cysteine; after incubation at 37°C for 18-20h, iodoacetamide was added to stop the reaction. Caprylic acid facilitated the precipitation of albumin, resulting in a reduced protein load presented to the digestion enzymes, culminating in substantial reductions in processing time. The ovine IgG, F(ab')2 and Fab products obtained using these novel caprylic acid methods were comparable in terms of yield, purity and specific activity to those obtained by multi-step conventional salt fractionation with sodium sulphate.

  11. Dynamics of ovarian maturation during the reproductive cycle of Metynnis maculatus, a reservoir invasive fish species (Teleostei: Characiformes

    Directory of Open Access Journals (Sweden)

    Thiago Scremin Boscolo Pereira

    Full Text Available In this study, we evaluated the dynamics of ovarian maturation and the spawning processes during the reproductive cycle of Metynnis maculatus. Adult females (n = 36 were collected bimonthly between April 2010 and March 2011. The mean gonadosomatic index (GSI was determined, ovarian and blood samples were submitted for morphometric evaluation and the steroid plasma concentration was determined by ELISA. This species demonstrated asynchronous ovarian development with multiple spawns. This study revealed that, although defined as a multiple spawning species, the ovaries of M. maculatus have a pattern of development with a predominance of vitellogenesis between April and August and have an intensification in spawning in September; in October, a drop in the mean GSI values occurred, and the highest frequencies of post-ovulatory follicles (POFs were observed. We observed a positive correlation between the POF and the levels of 17α-hydroxyprogesterone. Metynnis maculatus has the potential to be used as a source of pituitary tissue for the preparation of crude extracts for hormonal induction; the theoretical period for use is from September to December, but specific studies to determine the feasibility of this approach must be conducted.

  12. Stomach contents of the catfishes Osteogeneiosus militaris (Linnaeus, 1758 and Arius maculatus (Thunberg, 1792 in the Songkhla Lake

    Directory of Open Access Journals (Sweden)

    Angsupanich, S.


    Full Text Available The catfishes, Osteogeneiosus militaris and Arius maculatus, were collected from Songkhla Lake, southern Thailand at two sites that varied in the extent to which they were exposed to higher salinity (the Outer Songkhla Lake and which were exposed to lower salinity (the Inner Songkhla Lake. Both fish species were carnivores feeding mainly on benthic invertebrates with regard to its major components of Tanaidacea, Amphipoda, Bivalvia, Polychaeta and Isopoda. Tanaidacean (Apseudes sapensis clearly dominated the diets of O. militaris in both sites ranging from 33.3 to 96.7% by frequency of occurrence and 23.5 to 90.2% by number. Amphipods were the most dominant prey of A. maculatus ranging from 40.0 to 93.3% by frequency of occurrence and 7.0-60.2% by number. Although Polychaeta were less numerically abundantly eaten by catfishes (0.4-6.4%, the frequency of occurrence was high (10.0-86.7% in almost all seasons. Some other large (Alpheidae and fish larvae and fast mobile animals (Mysidacea were much more commonly found in A. maculatus than in O. militaris. This study implies that these catfish species feed opportunistically on a variety of prey in their environment coupled with preferential feeding.

  13. Contrasting Genetic Structure and Diversity of Galaxias maculatus (Jenyns, 1848) Along the Chilean Coast: Stock Identification for Fishery Management. (United States)

    González-Wevar, Claudio; Salinas, Pilar; Hüne, Mathias; Segovia, Nicolás; Vargas-Chacoff, Luis; Oda, Esteban; Poulin, Elie


    Galaxias maculatus (Pisces: Galaxiidae) commonly known as "puye" has a disjunct distribution along the Southern Hemisphere including landlocked and migratory populations at latitudes over 30°S in South America, Australia, Tasmania, and New Zealand. Chilean artisanal fishery of G. maculatus has become less important as a resource due to multiple factors including overexploitation, pollution, introduction of predators, and competitors. At the same time, the current conservation status of the species in Chile is still uncertain. Here, we used mtDNA control region sequences (925bp) to investigate main patterns of genetic diversity and structure in populations from 2 biogeographic areas along the Chilean coast. Extremely high levels of genetic diversity characterize the species, suggesting a low amount of influence of the last glacial cycle over its demography compared with other studies in freshwater and marine South American fishes. However, we recognized contrasting genetic patterns between the Intermediate Area (between 30°S and 42°S) and the Magellanic Province (between 42°S and 56°S). On the one hand, over a narrow geographical range (stock units for G. maculatus and this information should be integrated in future management strategies and aquaculture programs for this species. © The American Genetic Association 2015. All rights reserved. For permissions, please e-mail:

  14. Intraspecific and interspecific competition in Callosobruchus maculatus (F.) and Callosobruchus subinnotatus (Pic) on stored bambara groundnut, Vigna subterranea (L.) Verdcourt. (United States)

    Lale, N E.S.; Vidal, S


    Intraspecific competition was studied in Callosobruchus maculatus and Callosobruchus subinnotatus. Interspecific competition between the two bruchids was also studied to determine which of these species is likely to cause more damage to stored bambara groundnuts, Vigna subterranea in cases of joint infestation. Results showed that increasing the adult density up to 8 females per 10g of bambara groundnut seeds did not significantly reduce the mean number of eggs laid per female, the number of eggs developing to the adult stage, or the weight of emerged adults of either species. The developmental period of the two species was also not significantly affected. The adult emergence curve of C. maculatus was similar to that of C. subinnotatus and was of the scramble type. C. maculatus performed better than C. subinnotatus in interspecific competition and it achieved this through a higher egg-laying ability and a higher rate of progeny production coupled with a shorter life-cycle. The implications of these findings with respect to damage and possible loss of stored bambara groundnut are discussed.

  15. A parallel panning scheme used for selection of a GluA4-specific Fab targeting the ligand-binding domain

    DEFF Research Database (Denmark)

    Clausen, Rasmus P; Mohr, Andreas Ø; Riise, Erik


    molecule was used for immunization. A Fab-phage library was constructed and a parallel panning approach enabled selection of murine Fab fragments towards either intact ectodomain or the isolated LBD of the GluA4 receptor. One LBD-Fab (FabL9) showed exclusive selectivity for the GluA4 LBD, over a panel...

  16. 20 CFR 30.318 - Can the FAB consider objections to HHS's reconstruction of a radiation dose or to the guidelines... (United States)


    ... at 42 CFR part 82, is binding on the FAB. The FAB reviewer may determine, however, that objections... CFR part 81, is also binding on the FAB (see § 30.213). However, since OWCP applies this methodology... 20 Employees' Benefits 1 2010-04-01 2010-04-01 false Can the FAB consider objections to...

  17. Meleagrin, a new FabI inhibitor from Penicillium chryosogenum with at least one additional mode of action.

    Directory of Open Access Journals (Sweden)

    Chang Ji Zheng

    Full Text Available Bacterial enoyl-acyl carrier protein reductase (FabI is a promising novel antibacterial target. We isolated a new class of FabI inhibitor from Penicillium chrysogenum, which produces various antibiotics, the mechanisms of some of them are unknown. The isolated FabI inhibitor was determined to be meleagrin by mass spectroscopy and nuclear magnetic resonance spectral analyses, and its more active and inactive derivatives were chemically prepared. Consistent with their selective inhibition of Staphylococcus aureus FabI, meleagrin and its more active derivatives directly bound to S. aureus FabI in a fluorescence quenching assay, inhibited intracellular fatty acid biosynthesis and growth of S. aureus, and increased the minimum inhibitory concentration for fabI-overexpressing S. aureus. The compounds that were not effective against the FabK isoform, however, inhibited the growth of Streptococcus pneumoniae that contained only the FabK isoform. Additionally no resistant mutant to the compounds was obtained. Importantly, fabK-overexpressing Escherichia coli was not resistant to these compounds, but was resistant to triclosan. These results demonstrate that the compounds inhibited another target in addition to FabI. Thus, meleagrin is a new class of FabI inhibitor with at least one additional mode of action that could have potential for treating multidrug-resistant bacteria.

  18. Site-specific fab fragment biotinylation at the conserved nucleotide binding site for enhanced Ebola detection. (United States)

    Mustafaoglu, Nur; Alves, Nathan J; Bilgicer, Basar


    The nucleotide binding site (NBS) is a highly conserved region between the variable light and heavy chains at the Fab domains of all antibodies, and a small molecule that we identified, indole-3-butyric acid (IBA), binds specifically to this site. Fab fragment, with its small size and simple production methods compared to intact antibody, is good candidate for use in miniaturized diagnostic devices and targeted therapeutic applications. However, commonly used modification techniques are not well suited for Fab fragments as they are often more delicate than intact antibodies. Fab fragments are of particular interest for sensor surface functionalization but immobilization results in damage to the antigen binding site and greatly reduced activity due to their truncated size that allows only a small area that can bind to surfaces without impeding antigen binding. In this study, we describe an NBS-UV photocrosslinking functionalization method (UV-NBS(Biotin) in which a Fab fragment is site-specifically biotinylated with an IBA-EG11-Biotin linker via UV energy exposure (1 J/cm(2)) without affecting its antigen binding activity. This study demonstrates successful immobilization of biotinylated Ebola detecting Fab fragment (KZ52 Fab fragment) via the UV-NBS(Biotin) method yielding 1031-fold and 2-fold better antigen detection sensitivity compared to commonly used immobilization methods: direct physical adsorption and NHS-Biotin functionalization, respectively. Utilization of the UV-NBS(Biotin) method for site-specific conjugation to Fab fragment represents a proof of concept use of Fab fragment for various diagnostic and therapeutic applications with numerous fluorescent probes, affinity molecules and peptides.

  19. Direct-write scanning probe lithography: towards a desktop fab (United States)

    Giam, Louise R.; Senesi, Andrew J.; Liao, Xing; Wong, Lu Shin; Chai, Jinan; Eichelsdoerfer, Daniel J.; Shim, Wooyoung; Rasin, Boris; He, Shu; Mirkin, Chad A.


    Massively parallel scanning-probe based methods have been used to address the challenges of nanometer to millimeter scale printing for a variety of materials and mark a step towards the realization of a "desktop fab." Such tools enable simple, flexible, high-throughput, and low-cost nano- and microscale patterning, which allow researchers to rapidly synthesize and study systems ranging from nanoparticle synthesis to biological processes. We have developed a novel scanning probe-based cantilever-free printing method termed polymer pen lithography (PPL), which uses an array of elastomeric tips to transfer materials (e.g. alkanethiols, proteins, polymers) in a direct-write manner onto a variety of surfaces. This technique takes the best attributes of dip-pen nanolithography (DPN) and eliminates many of the disadvantages of contact printing. Various related techniques such as beam pen lithography (BPL), scanning probe block copolymer lithography (SPBCL), and hard-tip, soft spring lithography (HSL) are also discussed.

  20. Integrated fab process for metal oxide EUV photoresist (United States)

    Grenville, Andrew; Anderson, Jeremy T.; Clark, Benjamin L.; De Schepper, Peter; Edson, Joseph; Greer, Michael; Jiang, Kai; Kocsis, Michael; Meyers, Stephen T.; Stowers, Jason K.; Telecky, Alan J.; De Simone, Danilo; Vandenberghe, Geert


    Inpria is developing directly patternable, metal oxide hardmasks as robust, high-resolution photoresists for EUV lithography. Targeted formulations have achieved 13nm half-pitch at 35 mJ/cm2 on an ASML's NXE:3300B scanner. Inpria's second-generation materials have an absorbance of 20/μm, thereby enabling an equivalent photon shot noise compared to conventional resists at a dose lower by a factor of 4X. These photoresists have ~40:1 etch selectivity into a typical carbon underlayer, so ultrathin 20nm films are possible, mitigating pattern collapse. In addition to lithographic performance, we review progress in parallel advances required to enable the transition from lab to fab for such a metal oxide photoresist. This includes considerations and data related to: solvent compatibility, metals cross-contamination, coat uniformity, stability, outgassing, and rework.

  1. Blood cytology of the common jollytail (Galaxias maculatus) (Jenyns, 1842) (Osmeriformes: Galaxiidae) at postlarval and adult stages Estudio de la citología sanguínea del puye (Galaxias maculatus) (Jenyns, 1842) (Osmeriformes: Galaxiidae) en estado postlarval y adulto


    I Valdebenito; Busse, K; N. Jaramillo; Hernández, A.


    “Puye” (Galaxias maculatus) is a small freshwater fish species of great interest to Chilean aquaculture diversification, because of the high commercial value reached by its transparent larvae or “cristalino”. This study presents the first data with regard to blood cytology in larvae and adults of this specie. Blood smears stained with May Grünwald-Giemsa were analysed through optical microscopy. The results show that in G. maculatus larvae the flowing blood has a trans...

  2. Transcriptome analysis of female and male Xiphophorus maculatus Jp 163 A.

    Directory of Open Access Journals (Sweden)

    Ziping Zhang

    Full Text Available BACKGROUND: Xiphophorus models are important for melanoma, sex determination and differentiation, ovoviviparity and evolution. To gain a global view of the molecular mechanism(s whereby gene expression may influence sexual dimorphism in Xiphophorus and to develop a database for future studies, we performed a large-scale transcriptome study. METHODOLOGY/PRINCIPAL FINDINGS: The 454-FLX massively parallel DNA sequencing platform was employed to obtain 742,771 and 721,543 reads from 2 normalized cDNA libraries generated from whole adult female and male X. maculatus Jp 163 A, respectively. The reads assembled into 45,538 contigs (here, a "contig" is a set of contiguous sequences, of which, 11,918 shared homology to existing protein sequences. These numbers estimate that the contigs may cover 53% of the total number of Xiphophorus transcriptome. Putative translations were obtained for 11,918 cDNA contigs, of which, 3,049 amino acid sequences contain Pfam domains and 11,064 contigs encode secretory proteins. A total of 3,898 contigs were associated with 2,781 InterPro (IPR entries and 5,411 contigs with 132 KEGG (Kyoto Encyclopedia of Genes and Genomes pathways. There were 10,446 contigs annotated with 69,778 gene ontology (GO terms and the three corresponding organizing principles. Fifty-four potential sex differentially expressed genes have been identified from these contigs. Eight and nine of these contigs were confirmed by real-time PCR as female and male predominantly expressed genes respectively. Based on annotation results, 34 contigs were predicted to be differentially expressed in male and female and 17 of them were also confirmed by real-time PCR. CONCLUSIONS/SIGNIFICANCE: This is the first report of an annotated overview of the transcriptome of X. maculatus and identification of sex differentially expressed genes. These data will be of interest to researchers using the Xiphophorus model. This work also provides an archive for future studies

  3. SpiderFab: Architecture for On-Orbit Construction of Kilometer-Scale Apertures Project (United States)

    National Aeronautics and Space Administration — The SpiderFab effort has investigated the value proposition and technical feasibility of radically changing the way we build and deploy spacecraft in order to escape...

  4. Preparation and Purification of IgY and Fab' Against Human Rotavirus%抗轮状病毒IgY和Fab'的分离与纯化

    Institute of Scientific and Technical Information of China (English)

    孙淑清; 段春燕; 胡彦涛; 孟岩



  5. Fab glycosylation of immunoglobulin G does not associate with improvement of rheumatoid arthritis during pregnancy


    Bondt, Albert; Wuhrer, Manfred; Kuijper, Martijn; Hazes, Mieke; Dolhain, Radboud


    Background Changes in immunoglobulin G (IgG) constant domain (Fc) glycosylation are associated with changes in rheumatoid arthritis (RA) disease activity in response to pregnancy. Here, we sought to determine whether the same holds true for variable domain (Fab) glycosylation. Methods IgGs were captured from RA and control sera obtained before (RA only), during and after pregnancy, followed by Fc and Fab separation, glycan release, and mass spectrometric detection. In parallel, glycans from i...

  6. Anti-tumor Effect and Mechanism of SEA-Fab' Coupled Protein on Gastric Tumor

    Institute of Scientific and Technical Information of China (English)


    The anti-tumor effect and mechanism of SEA-Fab' coupled protein on gastric tumor was studied. The target cell Walker-256 was treated with SEA-Fab' synthesized chemically or SEA respectively for 24 h, 36 h or 72 h. PBMC+Walke-256 cells served as controls. The apoptotic index of SEA-Fab' against effector cells was detected. In the mouse gastric cancer models (n=60), SEAFab', SEA and normal saline was injected in experimental group, SEA group and control group respectively. The occurrence and weight of tumor was observed. The results showed that the apoptotic index was significantly higher in the SEA-Fab' (34.6 %-68.9 %) and SEA group (15.5 %-31.9 %) than in PBMC+Walker-256 group (5.5 %-12.8 %) with the difference being significant (P<0.01). And there was significant difference between SEA-Fab' group and SEA group (P <0. 01). The tumor weight in SEA-Fab', SEA and control groups was 3. 64±0. 53 g, 0. 78±0.26 g and 0.49 ±0.17 g respectively with the difference being statistically significant between the SEAFab' group, SEA group and the control group (P<0.01). In the SEA-Fab's and SEA groups,there were CD4+ T and CD8+ T cell infiltrates, but in the cotnrol group, no or few T lymphocytes were seen in the mouse tumor tissue. It was concluded that SEA-Fab' was more effective to activate T lymphocytes to kill the tumor cells than SEA used alone. It was feasibility by using the monoclonal antibody as carrier to perform the targeted immunotherapy of gatric tumor.

  7. Promoter engineering to optimize recombinant periplasmic Fab' fragment production in Escherichia coli. (United States)

    Schofield, Desmond M; Templar, Alex; Newton, Joseph; Nesbeth, Darren N


    Fab' fragments have become an established class of biotherapeutic over the last two decades. Likewise, developments in synthetic biology are providing ever more powerful techniques for designing bacterial genes, gene networks and entire genomes that can be used to improve industrial performance of cells used for production of biotherapeutics. We have previously observed significant leakage of an exogenous therapeutic Fab' fragment into the growth medium during high cell density cultivation of an Escherichia coli production strain. In this study we sought to apply a promoter engineering strategy to address the issue of Fab' fragment leakage and its consequent bioprocess challenges. We used site directed mutagenesis to convert the Ptac promoter, present in the plasmid, pTTOD-A33 Fab', to a Ptic promoter which has been shown by others to direct expression at a 35% reduced rate compared to Ptac . We characterized the resultant production trains in which either Ptic or Ptac promoters direct Fab' fragment expression. The Ptic promoter strain showed a 25-30% reduction in Fab' expression relative to the original Ptac strain. Reduced Fab' leakage and increased viability over the course of a fed-batch fermentation were also observed for the Ptic promoter strain. We conclude that cell design steps such as the Ptac to Ptic promoter conversion reported here, can yield significant process benefit and understanding with respect to periplasmic Fab' fragment production. It remains an open question as to whether the influence of transgene expression on periplasmic retention is mediated by global metabolic burden effects or periplasm overcapacity. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:840-847, 2016.

  8. Generation and selection of immunized Fab phage display library against human B cell lymphoma

    Institute of Scientific and Technical Information of China (English)

    Yongmei Shen; Xiaochun Yang; Ningzheng Dong; Xiaofang Xie; Xia Bai; Yizhen Shi


    The approval of using monoclonal antibodies as a targeted therapy in the management of patients with B cell lymphoma has led to new treatment options for this group of patients. Production of monoclonal antibodies by the traditional hybridoma technology is costly, and the resulting murine antibodies often have the disadvantage of triggering human anti-mouse antibody (HAMA) response. Therefore recombinant Fab antibodies generated by the phage display technology can be a suitable alternative in managing B cell lymphoma. In this study, we extracted total RNA from spleen cells of BALB/c mice immunized with human B lymphoma cells, and used RT-PCR to amplify cDNAs coding for the K light chains and Fd fragments of heavy chains. After appropriate restriction digests, these cDNA fragments were successively inserted into the phagemid vector pComb3H-SS to construct an immunized Fab phage display library. The diversity of the constructed library was approximately 1.94×107. Following five rounds of biopanning, soluble Fab antibodies were produced from positive clones identified by ELISA. From eight positive clones, FabC06, FabC21, FabC43 and FabC59 were selected for sequence analysis. At the level of amino acid sequences, the variable heavy domains (VH) and variable light domains (VL) were found to share 88-92% and 89-94% homology with sequences coded by the corresponding murine germline genes respectively. Furthermore, reactivity with membrane proteins of the B cell lymphoma was demonstrated by immunohistochemistry and western blotting. These immunized Fab antibodies may provide a valuable tool for further study of B cell lymphoma and could also contribute to the improvement of disease therapy.

  9. Immunoglobulin fragments, F(ab')2, that are cytotoxic to enzyme-treated cells. (United States)

    Holtgrewe, E M; Killion, J J


    Bivalent immunoglobulin fragments of IgG, F(ab')2, prepared from normal murine sera were found to be cytotoxic to neuraminidase-treated cells. The fragments were cytotoxic to both allogenic and syngeneic targets (with respect to the source of the sera), suggesting that the antigen bound by the F(ab')2 is not related to the major histocompatibility locus of mice (H-2).

  10. Functional characterization of triclosan-resistant enoyl-acyl-carrier protein reductase (FabV in Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Yong-Heng Huang


    Full Text Available Pseudomonas aeruginosa is extremely resistant to triclosan. Previous studies have shown that P. aeruginosa encodes a triclosan-resistant enoyl-acyl-carrier protein reductase (ENR, FabV, and that deletion of fabV causes P. aeruginosa to be extremely sensitive to triclosan. In this report, we complemented a P. aeruginosa fabV deletion strain with several triclosan-resistant ENR encoding genes, including Vibrio cholera fabV, Bacillus subtilis fabL and Enterococcus faecalis fabK. All complemented strains restored triclosan resistance to the level of the wild-type strain, which confirmed that triclosan-resistant ENR allows P. aeruginosa to be extremely resistant to triclosan. Moreover, fabV exhibits pleiotropic effects. Deletion of fabV led P. aeruginosa to show attenuated swarming motility, decreased rhamnolipid, pyoverdine and acylhomoserine lactones (AHLs production. Complementation of the fabV mutant with any one ENR encoding gene could restore these features to some extent, in comparison with the wild-type strain. Furthermore, we found that addition of exogenous AHLs could restore to the fabV mutant strain the ability to swarm on semisolid plates and to produce more virulence factors than the fabV mutant strain. These findings indicate that deletion of fabV reduced the activity of ENR in P. aeruginosa, decreased fatty acid synthesis, and subsequently depressed the production of AHLs and other virulence factors, which finally may led to a reduction in the pathogenicity of P. aeruginosa. Therefore, fabV should be an ideal target for the control of P. aeruginosa infectivity.

  11. Functional Characterization of Triclosan-Resistant Enoyl-acyl-carrier Protein Reductase (FabV) in Pseudomonas aeruginosa (United States)

    Huang, Yong-Heng; Lin, Jin-Shui; Ma, Jin-Cheng; Wang, Hai-Hong


    Pseudomonas aeruginosa is extremely resistant to triclosan. Previous studies have shown that P. aeruginosa encodes a triclosan-resistant enoyl-acyl-carrier protein reductase (ENR), FabV, and that deletion of fabV causes P. aeruginosa to be extremely sensitive to triclosan. In this report, we complemented a P. aeruginosa fabV deletion strain with several triclosan-resistant ENR encoding genes, including Vibrio cholerae fabV, Bacillus subtilis fabL and Enterococcus faecalis fabK. All complemented strains restored triclosan resistance to the level of the wild-type strain, which confirmed that triclosan-resistant ENR allows P. aeruginosa to be extremely resistant to triclosan. Moreover, fabV exhibits pleiotropic effects. Deletion of fabV led P. aeruginosa to show attenuated swarming motility, decreased rhamnolipid, pyoverdine and acyl-homoserine lactones (AHLs) production. Complementation of the fabV mutant with any one ENR encoding gene could restore these features to some extent, in comparison with the wild-type strain. Furthermore, we found that addition of exogenous AHLs could restore the fabV mutant strain to swarm on semisolid plates and to produce more virulence factors than the fabV mutant strain. These findings indicate that deletion of fabV reduced the activity of ENR in P. aeruginosa, decreased fatty acid synthesis, and subsequently depressed the production of AHLs and other virulence factors, which finally may led to a reduction in the pathogenicity of P. aeruginosa. Therefore, fabV should be an ideal target for the control of P. aeruginosa infectivity. PMID:27965638

  12. Structural characterisation of the fatty acid biosynthesis enzyme FabF from the pathogen Listeria monocytogenes (United States)

    Soares da Costa, Tatiana P.; Nanson, Jeffrey D.; Forwood, Jade K.


    Development of new antimicrobial agents is required against the causative agent for listeriosis, Listeria monocytogenes, as the number of drug resistant strains continues to increase. A promising target is the β-ketoacyl-acyl carrier protein synthase FabF, which participates in the catalysis of fatty acid synthesis and elongation, and is required for the production of phospholipid membranes, lipoproteins, and lipopolysaccharides. In this study, we report the 1.35 Å crystal structure of FabF from L. monocytogenes, providing an excellent platform for the rational design of novel inhibitors. By comparing the structure of L. monocytogenes FabF with other published bacterial FabF structures in complex with known inhibitors and substrates, we highlight conformational changes within the active site, which will need to be accounted for during drug design and virtual screening studies. This high-resolution structure of FabF represents an important step in the development of new classes of antimicrobial agents targeting FabF for the treatment of listeriosis. PMID:28045020

  13. Investigation of protein selectivity in multimodal chromatography using in silico designed Fab fragment variants. (United States)

    Karkov, Hanne Sophie; Krogh, Berit Olsen; Woo, James; Parimal, Siddharth; Ahmadian, Haleh; Cramer, Steven M


    In this study, a unique set of antibody Fab fragments was designed in silico and produced to examine the relationship between protein surface properties and selectivity in multimodal chromatographic systems. We hypothesized that multimodal ligands containing both hydrophobic and charged moieties would interact strongly with protein surface regions where charged groups and hydrophobic patches were in close spatial proximity. Protein surface property characterization tools were employed to identify the potential multimodal ligand binding regions on the Fab fragment of a humanized antibody and to evaluate the impact of mutations on surface charge and hydrophobicity. Twenty Fab variants were generated by site-directed mutagenesis, recombinant expression, and affinity purification. Column gradient experiments were carried out with the Fab variants in multimodal, cation-exchange, and hydrophobic interaction chromatographic systems. The results clearly indicated that selectivity in the multimodal system was different from the other chromatographic modes examined. Column retention data for the reduced charge Fab variants identified a binding site comprising light chain CDR1 as the main electrostatic interaction site for the multimodal and cation-exchange ligands. Furthermore, the multimodal ligand binding was enhanced by additional hydrophobic contributions as evident from the results obtained with hydrophobic Fab variants. The use of in silico protein surface property analyses combined with molecular biology techniques, protein expression, and chromatographic evaluations represents a previously undescribed and powerful approach for investigating multimodal selectivity with complex biomolecules.

  14. Functional Requirements for Fab-7 Boundary Activity in the Bithorax Complex. (United States)

    Wolle, Daniel; Cleard, Fabienne; Aoki, Tsutomu; Deshpande, Girish; Schedl, Paul; Karch, Francois


    Chromatin boundaries are architectural elements that determine the three-dimensional folding of the chromatin fiber and organize the chromosome into independent units of genetic activity. The Fab-7 boundary from the Drosophila bithorax complex (BX-C) is required for the parasegment-specific expression of the Abd-B gene. We have used a replacement strategy to identify sequences that are necessary and sufficient for Fab-7 boundary function in the BX-C. Fab-7 boundary activity is known to depend on factors that are stage specific, and we describe a novel ∼700-kDa complex, the late boundary complex (LBC), that binds to Fab-7 sequences that have insulator functions in late embryos and adults. We show that the LBC is enriched in nuclear extracts from late, but not early, embryos and that it contains three insulator proteins, GAF, Mod(mdg4), and E(y)2. Its DNA binding properties are unusual in that it requires a minimal sequence of >65 bp; however, other than a GAGA motif, the three Fab-7 LBC recognition elements display few sequence similarities. Finally, we show that mutations which abrogate LBC binding in vitro inactivate the Fab-7 boundary in the BX-C.

  15. Near-Atomic Resolution Structure of a Highly Neutralizing Fab Bound to Canine Parvovirus. (United States)

    Organtini, Lindsey J; Lee, Hyunwook; Iketani, Sho; Huang, Kai; Ashley, Robert E; Makhov, Alexander M; Conway, James F; Parrish, Colin R; Hafenstein, Susan


    Canine parvovirus (CPV) is a highly contagious pathogen that causes severe disease in dogs and wildlife. Previously, a panel of neutralizing monoclonal antibodies (MAb) raised against CPV was characterized. An antibody fragment (Fab) of MAb E was found to neutralize the virus at low molar ratios. Using recent advances in cryo-electron microscopy (cryo-EM), we determined the structure of CPV in complex with Fab E to 4.1 Å resolution, which allowed de novo building of the Fab structure. The footprint identified was significantly different from the footprint obtained previously from models fitted into lower-resolution maps. Using single-chain variable fragments, we tested antibody residues that control capsid binding. The near-atomic structure also revealed that Fab binding had caused capsid destabilization in regions containing key residues conferring receptor binding and tropism, which suggests a mechanism for efficient virus neutralization by antibody. Furthermore, a general technical approach to solving the structures of small molecules is demonstrated, as binding the Fab to the capsid allowed us to determine the 50-kDa Fab structure by cryo-EM.

  16. Heat stress but not inbreeding affects offensive sperm competitiveness in Callosobruchus maculatus. (United States)

    Lieshout, Emile; Tomkins, Joseph L; Simmons, Leigh W


    Environmental and genetic stress have well-known detrimental effects on ejaculate quality, but their concomitant effect on male fitness remains poorly understood. We used competitive fertilization assays to expose the effects of stress on offensive sperm competitive ability in the beetle Callosobruchus maculatus, a species where ejaculates make up more than 5% of male body mass. To examine the effects of environmental and genetic stress, males derived from outcrosses or sib matings were heat shocked at 50°C for 50 min during the pupal stage, while their siblings were maintained at a standard rearing temperature of 28°C. Heat-shocked males achieved only half the offensive paternity success of their siblings. While this population exhibited inbreeding depression in body size, sperm competitiveness was unaffected by inbreeding, nor did the effect of heat shock stress on sperm competitiveness depend on inbreeding status. In contrast, pupal emergence success was increased by 34% among heat-stressed individuals, regardless of their inbreeding status. Heat-shocked males' ejaculate size was 19% reduced, but they exhibited 25% increased mating duration in single mating trials. Our results highlight both the importance of stress in postcopulatory sexual selection, and the variability among stressors in affecting male fitness.

  17. Phylogeography in Galaxias maculatus (Jenyns, 1848) along Two Biogeographical Provinces in the Chilean Coast (United States)

    González-Wevar, Claudio A.; Salinas, Pilar; Hüne, Mathias; Segovia, Nicolás I.; Vargas-Chacoff, Luis; Astorga, Marcela; Cañete, Juan I.; Poulin, Elie


    Major geologic and climatic changes during the Quaternary exerted a major role in shaping past and contemporary distribution of genetic diversity and structure of aquatic organisms in southern South America. In fact, the northern glacial limit along the Pacific coast, an area of major environmental changes in terms of topography, currents, and water salinity, represents a major biogeographic transition for marine and freshwater species. We used mitochondrial DNA sequences (D-loop) to investigate the consequences of Quaternary glacial cycles over the pattern of genetic diversity and structure of G. maculatus (Pisces: Galaxiidae) along two biogeographical provinces in the Chilean coast. Extreme levels of genetic diversity and strong phylogeographic structure characterize the species suggesting a low amount of influence of the last glacial cycle over its demography. However, we recognized contrasting patterns of genetic diversity and structure between main biogeographical areas here analyzed. Along the Intermediate Area (38°–41° S) each estuarine population constitutes a different unit. In contrast, Magellanic populations (43°–53° S) exhibited low levels of genetic differentiation. Contrasting patterns of genetic diversity and structure recorded in the species between the analyzed biogeographic areas are consistent with the marked differences in abiotic factors (i.e., different coastal configurations, Quaternary glacial histories, and oceanographic regimes) and to inherent characteristics of the species (i.e., salt-tolerance, physiology, and reproductive behavior). PMID:26161896

  18. Molecular genetic response to varied wavelengths of light in Xiphophorus maculatus skin. (United States)

    Chang, Jordan; Lu, Yuan; Boswell, William T; Boswell, Mikki; Caballero, Kaela L; Walter, Ronald B


    Xiphophorus fishes represent a model often utilized to study UVB induced tumorigenesis. Recently, varied genetic responses to UVB exposure have been documented in the skin of female and male Xiphophorus, as have differences in UVB response in the skin of different parental species and for interspecies hybrids produced from crossing them. Additionally, it has been shown that exposure to "cool white" fluorescent light induces a shift in the genetic profiles of Xiphophorus skin that is nearly as robust as the UVB response, but involves a fundamentally different set of genes. Given these results and the use of Xiphophorus interspecies hybrids as an experimental model for UVB inducible melanoma, it is of interest to characterize genes that may be transcriptionally modulated in a wavelength specific manner. The global molecular genetic response of skin upon exposure of the intact animal to specific wavelengths of light has not been investigated. Herein, we report results of RNA-Seq experiments from the skin of male Xiphophorus maculatus Jp 163 B following exposure to varied 50nm wavelengths of light ranging from 300-600nm. We identify two specific wavelength regions, 350-400nm (88 genes) and 500-550nm (276 genes), that exhibit transcriptional modulation of a significantly greater number of transcripts than any of the other 50nm regions in the 300-600nm range. Observed functional sets of genes modulated within these two transcriptionally active light regions suggest different mechanisms of gene modulation.

  19. Effects of dietary antioxidant of tomato extract and lycopene on Carassius auratus and Xiphophorus maculatus

    Directory of Open Access Journals (Sweden)

    Cynthia Montoya M.


    Full Text Available Objective. Evaluate the effect on tegument pigmentation, survival, growth and antioxidant capacity in diets supplemented with tomato extract and lycopene as additives in experimental feed for Carassius auratus and Xiphophorus maculatus. Materials and methods. The additives were added in different concentrations to a basic diet. We performed beginning and an ending biometrics for 100% of the population in each bioassay. The growth and survival of organisms were evaluated. The antioxidant capacity was analyzed by ABTS assay, both in the tomato extract sample as well as in foods used in different bioassays. The concentration of lycopene was determined in food and liver and muscle samples of fish fed with it. Acquired pigmentation of fish was assessed through photographs analyzed with Adobe Photoshop®. The results were evaluated by analysis of variance, and when differences were found (p0.05 on pigmentation and growth of the organisms under the established experimental conditions was obtained. Significant differences in antioxidant capacity (p<0.05 were obtained in foods with added lycopene. Conclusions. The inclusion of lycopene or tomato extract in food for the organisms used is not recommended to improve pigmentation, but further studies are needed to demonstrate antioxidant effect.

  20. Can preference for oviposition sites initiate reproductive isolation in Callosobruchus maculatus?

    Directory of Open Access Journals (Sweden)

    Emma Rova

    Full Text Available Theory has identified a variety of evolutionary processes that may lead to speciation. Our study includes selection experiments using different host plants and test key predictions concerning models of speciation based on host plant choice, such as the evolution of host use (preference and performance and assortative mating. This study shows that after only ten generations of selection on different resources/hosts in allopatry, strains of the seed beetle Callosobruchus maculatus develop new resource preferences and show resource-dependent assortative mating when given the possibility to choose mates and resources during secondary contact. The resulting reduced gene flow between the different strains remained for two generations after contact before being overrun by disassortative mating. We show that reduced gene flow can evolve in a population due to a link between host preference and assortative mating, although this result was not found in all lines. However, consistent with models of speciation, assortative mating alone is not sufficient to maintain reproductive isolation when individuals disperse freely between hosts. We conclude that the evolution of reproductive isolation in this system cannot proceed without selection against hybrids. Other possible factors facilitating the evolution of isolation would be longer periods of allopatry, the build up of local adaptation or reduced migration upon secondary contact.

  1. Pyramiding of insecticidal compounds for control of the cowpea bruchid (Callosobruchus maculatus F.). (United States)

    Tarver, Matthew R; Shade, Richard E; Shukle, Richard H; Moar, William J; Muir, William M; Murdock, Larry M; Pittendrigh, Barry R


    The cowpea bruchid (Callosobruchus maculatus F.) (Chrysomelidae: Bruchini) is a major pest of stored cowpea grain. With limited available technologies for controlling the bruchid, transgenic cowpeas with bruchid resistance genes engineered into them could become the next management tools. An investigation was made of two different sets of potential transgenic insecticidal compounds using an artificial seed system: (i) CIP-PH-BT-J and recombinant egg white avidin, and (ii) avidin and wheat alpha-amylase inhibitor. CIP-PH-BT-J (0.1%; 1000 mg kg(-1)) and recombinant egg white avidin (0.006%; 60 mg kg(-1)) incorporated separately into artificial seeds caused 98.2 and 99% larval mortality rates respectively. Combining CIP-PH-BT-J and avidin in the same artificial seed provided additional mortality compared with each factor incorporated singly; no insects survived in seeds with the combined toxins. Similarly, when avidin and wheat alpha-amylase inhibitor (alphaAI) (1%; 10 g kg(-1)) were incorporated separately into artificial seeds, this caused 99.8 and 98% mortality respectively. However, in combination, avidin and alphaAI did not increase mortality, but they did cause a significant increase in developmental time of the cowpea bruchids. These results emphasize that the joint action of potential insecticidal compounds cannot be predicted from results obtained separately for each compound, and they suggest potential transgenes for further consideration.

  2. Effects of cholinergic and noradrenergic agents on locomotion in the mudpuppy (Necturus maculatus). (United States)

    Fok, M; Stein, R B


    Some neurotransmitters act consistently on the central pattern generator (CPG) for locomotion in a wide range of vertebrates. In contrast, acetylcholine (ACh) and noradrenaline (NA) have various effects on locomotion in different preparations. The roles of ACh and NA have not been studied in amphibian walking, so we examined their effects in an isolated spinal cord preparation of the mudpuppy ( Necturus maculatus). This preparation contains a CPG that produces locomotor activity when N-methyl- D-aspartic acid (NMDA), an excitatory amino acid agonist, is added to the bath. The addition of carbachol, a long acting ACh agonist, to the bath disrupted the walking rhythm induced by NMDA, while not changing the level of activity in flexor and extensor motoneurons. Adding clonidine, an alpha(2)-noradrenergic agonist, had no effect on the NMDA-induced walking rhythm. Physostigmine, an ACh-esterase inhibitor, disrupted the walking rhythm, presumably by potentiating the effects of endogenously released ACh. Atropine, an ACh antagonist that binds to muscarinic ACh receptors, blocked the effects of carbachol, indicating that the action is mediated, at least in part, by muscarinic receptors. In the absence of carbachol, atropine had no effect. Locomotion was not induced by carbachol, atropine or clonidine in a resting spinal cord preparation. Cholinergic actions do not seem to be essential to the CPG for walking in the mudpuppy, but ACh may convert a rhythmic walking state to a more tonic state with occasional bursts of EMG activity for postural adjustments.

  3. Lipid Status of the Two High Latitude Fish Species, Leptoclinus maculatus and Lumpenus fabricii

    Directory of Open Access Journals (Sweden)

    Nina N. Nemova


    Full Text Available A comparative study of the lipid status (i.e., the total lipid and phospholipid concentrations and the percentage of fatty acids of the total lipids of adult specimens of daubed shanny (Leptoclinus maculatus from Svalbard waters (Isfjord and slender eel blenny (Lumpenus fabricii from the White Sea (Onega Bay and Tersky shore was performed to study the metabolism and functions of lipids of these fishes in ontogeny and under various ecological conditions. Slender eel blenny from both areas of the White Sea were distinguished by a high level of sphingomyelin compared with the daubed shanny from Svalbard, and the amount of total phospholipids was higher in slender eel blenny from Onega Bay than in slender eel blenny from the Tersky shore. The extent of saturation and the signature of polyenic fatty acids varied according to the specific species of the Stichaeidae family under study. These results demonstrate the differences in the trophoecological and hydrobiological conditions of habitations of these species and highlighted the importance of considering certain trends in the lipid profiles of these fishes as specific features of the organization of the ecological and biochemical mechanisms of adaptation.

  4. Seasonal dynamics of the flower head infestation of Smallanthus maculatus by two nonfrugivorous tephritids. (United States)

    Dzul-Cauich, José F; Hernández-Ortiz, Vicente; Parra-Tabla, Victor; Rico-Gray, Victor


    Seasonal dynamics of the capitula infested by Dictyotrypeta sp. and Rhynencina spilogaster (Steyskal) (Diptera: Tephritidae) was evaluated throughout the flowering cycle of their host plant the sunflower, Smallanthus maculatus (Cavanilles) Robinson (Asterales: Asteraceae). In central Veracruz, Mexico, along 16 consecutive weeks, a total of 1,017 mature capitula were collected, recording the presence and abundance of immature stages (larvae and pupae) and their related parasitoids. Both fly species were present throughout the entire season, with overall infestation of 51.5% of the capitula examined. However, Dictyotrypeta sp. infested 11.3%, representing about one-fifth of them, and R. spilogaster was most abundant infesting four times as many capitula (42.9%), whereas both species were found together in only 2.6% of the capitula examined. Based on the temporal occurrence of larvae and pupae into flower heads as well as their associated parasitoids and times of emergence, Dictyotrypeta sp. had two yearly generations, and it seems that the second generation could enter a seasonal diapause; in contrast, R. spilogaster was a univoltine species that entered diapause that lasted until the next year. © The Author 2014. Published by Oxford University Press on behalf of the Entomological Society of America.

  5. Evaluation of the dust and methanol extracts of Garcinia kolae for the control of Callosobruchus maculatus (F.) and Sitophilus zeamais (Mots)

    Institute of Scientific and Technical Information of China (English)


    Insecticidal effects of different doses of the dust and methanol extracts ofGarcinia kolae on Callosobruchus maculatus and Sitophilus zeamais were tested. The dust had no significant effect on the two insects; none of them died even at 3 d after treatment. The methanol extracts, however, had rapid lethal effects on both C. maculatus and S. zeamais. The mortality of C.maculatus by the lowest concentration of methanol extracts ranged from 95%~100% whereas in S. zeamais, the mortality ranged from 87.5%~100% and 70% ~100% in concentrations of 1 g extract+3 ml methanol and 1 g extract+5 ml methanol, respectively,from 24 to 48 h. The least concentration of 1 g extract+15 ml methanol had no significant lethal effect on Sitophilus zeamais.

  6. Coleoptera Associated with Decaying Wood in a Tropical Deciduous Forest. (United States)

    Muñoz-López, N Z; Andrés-Hernández, A R; Carrillo-Ruiz, H; Rivas-Arancibia, S P


    Coleoptera is the largest and diverse group of organisms, but few studies are dedicated to determine the diversity and feeding guilds of saproxylic Coleoptera. We demonstrate the diversity, abundance, feeding guilds, and succession process of Coleoptera associated with decaying wood in a tropical deciduous forest in the Mixteca Poblana, Mexico. Decaying wood was sampled and classified into four stages of decay, and the associated Coleoptera. The wood was identified according to their anatomy. Diversity was estimated using the Simpson index, while abundance was estimated using a Kruskal-Wallis test; the association of Coleoptera with wood species and decay was assessed using canonical correspondence analysis. Decay wood stage I is the most abundant (51%), followed by stage III (21%). We collected 93 Coleoptera belonging to 14 families, 41 genera, and 44 species. The family Cerambycidae was the most abundant, with 29% of individuals, followed by Tenebrionidae with 27% and Carabidae with 13%. We recognized six feeding guilds. The greatest diversity of Coleoptera was recorded in decaying Acacia farnesiana and Bursera linanoe. Kruskal-Wallis analysis indicated that the abundance of Coleoptera varied according to the species and stage of decay of the wood. The canonical analysis showed that the species and stage of decay of wood determined the composition and community structure of Coleoptera.


    Institute of Scientific and Technical Information of China (English)

    刘小云; 刘秀均; 李毅; 王维刚; 甄永苏


    目的研制一种以单抗Fab'片段为基础的抗肿瘤导向药物.方法制备单抗3A5 Fab'片段及其与平阳霉素(PYM)偶联物Fab'-PYM后,测定Fab'-PYM与肿瘤细胞的免疫反应性、偶联物中PYM的抑菌活性、对肿瘤细胞的杀伤作用和体内抑瘤作用.结果 Fab'及Fab'-PYM保持了与靶细胞C26的免疫反应性;偶联物中PYM的抑菌活性为游离PYM的15%;Fab'-PYM对C26细胞的杀伤作用强于PYM;对非靶细胞KB的杀伤作用与PYM相似;ip和iv给药,Fab'-PYM对小鼠皮下接种的肠癌26生长抑制作用均强于3A5-PYM和PYM.结论 Fab'-PYM具有比PYM及3A5-PYM更强的体内外抗肿瘤作用.

  8. Fab Four: When John and George play gravitation and cosmology

    CERN Document Server

    Bruneton, Jean-Philippe; Kanfon, Antonin; Hees, Aurélien; Schlögel, Sandrine; Füzfa, André


    Scalar-tensor theories of gravitation have recently regained a great interest after the discovery of the Chameleon mechanism and of the Galileon models. The former allows, in principle, to reconcile the presence of cosmological scalar fields with the constraints from experiments at the Solar System scale. The latter open up the possibility of building inflationary models that, among other things, do not need ad hoc potentials. Further generalizations have finally led to the most general tensor-scalar theory, recently dubbed the "Fab Four", with only first and second order derivatives of the fields in the equations of motion and that self-tune to a vanishing cosmological constant. This model has a very rich phenomenology that needs to be explored and confronted with experimental data in order to constrain a very large parameter space. In this paper, we present some results regarding a subset of the theory named "John", which corresponds to a non-minimal derivative coupling between the scalar field and the Eins...

  9. Fab Four: When John and George Play Gravitation and Cosmology

    Directory of Open Access Journals (Sweden)

    J.-P. Bruneton


    Full Text Available Scalar-tensor theories of gravitation attract again a great interest since the discovery of the Chameleon mechanism and of the Galileon models. The former allows reconciling the presence of a scalar field with the constraints from Solar System experiments. The latter leads to inflationary models that do not need ad hoc potentials. Further generalizations lead to a tensor-scalar theory, dubbed the “Fab Four,” with only first and second order derivatives of the fields in the equations of motion that self-tune to a vanishing cosmological constant. This model needs to be confronted with experimental data in order to constrain its large parameter space. We present some results regarding a subset of this theory named “John,” which corresponds to a nonminimal derivative coupling between the scalar field and the Einstein tensor in the action. We show that this coupling gives rise to an inflationary model with very unnatural initial conditions. Thus, we include the term named “George,” namely, a nonminimal, but nonderivative, coupling between the scalar field and Ricci scalar. We find a more natural inflationary model, and, by performing a post-Newtonian analysis, we derive the set of equations that constrain the parameter space with data from experiments in the Solar System.

  10. Triclosan Resistance in a Bacterial Fish Pathogen, Aeromonas salmonicida subsp. salmonicida, is Mediated by an Enoyl Reductase, FabV. (United States)

    Khan, Raees; Lee, Myung Hwan; Joo, Hae-Jin; Jung, Yong-Hoon; Ahmad, Shabir; Choi, Jin-Hee; Lee, Seon-Woo


    Triclosan, the widely used biocide, specifically targets enoyl-acyl carrier protein reductase (ENR) in the bacterial fatty acid synthesis system. Although the fish pathogen Aeromonas salmonicida subsp. salmonicida exhibits triclosan resistance, the nature of this resistance has not been elucidated. Here, we aimed to characterize the triclosan resistance of A. salmonicida subsp. salmonicida causing furunculosis. The fosmid library of triclosan-resistant A. salmonicida subsp. salmonicida was constructed to select a fosmid clone showing triclosan resistance. With the fosmid clone showing triclosan resistance, a subsequent secondary library search resulted in the selection of subclone pTSR-1. DNA sequence analysis of pTSR-1 revealed the presence of a chromosomal-borne fabV-encoding ENR homolog. The ENR of A. salmonicida (FabVas) exhibited significant homology with previously known FabV, including the catalytic domain YX(8)K. fabVas introduction into E. coli dramatically increased its resistance to triclosan. Heterologous expression of FabVas might functionally replace the triclosan-sensitive FabI in vivo to confer E. coli with triclosan resistance. A genome-wide search for fabVas homologs revealed the presence of an additional fabV gene (fabVas2) paralog in A. salmonicida strains and the fabVas orthologs from other gram-negative fish pathogens. Both of the potential FabV ENRs expressed similarly with or without triclosan supplement. This is the first report about the presence of two potential FabV ENRs in a single pathogenic bacterium. Our result suggests that triclosan-resistant ENRs are widely distributed in various bacteria in nature, and the wide use of this biocide can spread these triclosan-tolerant ENRs among fish pathogens and other pathogenic bacteria.

  11. RetroFab: A Design Tool for Retrofitting Physical Interfaces using Actuators, Sensors and 3D Printing



    We present RetroFab, an end-to-end design and fabrication environment that allows non-experts to retrofit physical interfaces. Our approach allows for changing the layout and behavior of physical interfaces. Unlike customizing software interfaces, physical interfaces are often challenging to adapt because of their rigidity. With RetroFab, a new physical interface is designed that serves as a proxy interface for the legacy controls that are now operated by actuators. RetroFab makes this concep...

  12. Effects of Fab' fragments of specific egg yolk antibody (IgY-Fab') against Shewanella putrefaciens on the preservation of refrigerated turbot. (United States)

    Zhang, Qian; Lin, Hong; Sui, Jianxin; Wang, Jingxue; Cao, Limin


    In our previous studies the specific egg yolk antibody (IgY) against Shewanella putrefaciens (one of the specific spoilage organisms for marine products during aerobic chilling storage) demonstrated significant activity to prolong the shelf life of refrigerated fish. The exploitation of the antigen-binding fragment plus the hinge region (IgY-Fab') is now considered a promising method for improving the efficiency of such natural antimicrobial agents. The antimicrobial activity of IgY-Fab' against S. putrefaciens was investigated using refrigerated turbot as samples. By microbial, chemical and sensory tests, it was shown to be able to effectively inhibit bacterial growth and prolong the shelf life of samples, with an efficiency evaluated significantly higher than that of whole IgY with the same molarity. The interaction between IgY agents and S. putrefaciens cells was also investigated, and the IgY-Fab' showed a much greater ability to damage cell membranes than the whole IgY. Compared to whole IgY with the same molarity, IgY-Fab' demonstrated higher and more durable antimicrobial efficiency. Such a result was assumed to be closely related to its structural properties (such as the much lower molecular weight), which may enhance its ability to influence physiological activities of antigen bacteria, especially the property or/and structure of cell membranes. © 2014 Society of Chemical Industry.

  13. 3-Substituted Indole Inhibitors Against Francisella tularensis FabI Identified by Structure-Based Virtual Screening (United States)


    FabI, but share low sequence identity and are poorly inhibited by triclosan.25,26 S. pneumoniae and P. aeruginosa contain FabK,24 and Vibrio cholerae ,27...Massengo-Tiasse, R. P.; Cronan, J. E. Vibrio cholerae FabV defines a new class of enoyl-acyl carrier protein reductase. J. Biol. Chem. 2008, 283, 1308...of enoyl- (acyl-carrier protein) reductase, FabV, from Vibrio fischeri. Acta Crystallogr., Sect. F: Struct. Biol. Cryst. Commun. 2012, 68, 78−80. (27

  14. Effect of polyethylene glycol conjugation on conformational and colloidal stability of a monoclonal antibody antigen-binding fragment (Fab'). (United States)

    Roque, Cristopher; Sheung, Anthony; Rahman, Nausheen; Ausar, S Fernando


    We have investigated the effects of site specific "hinge" polyethylene glycol conjugation (PEGylation) on thermal, pH, and colloidal stability of a monoclonal antibody antigen-binding fragment (Fab') using a variety of biophysical techniques. The results obtained by circular dichroism (CD), ultraviolet (UV) absorbance, and fluorescence spectroscopy suggested that the physical stability of the Fab' is maximized at pH 6-7 with no apparent differences due to PEGylation. Temperature-induced aggregation experiments revealed that PEGylation was able to increase the transition temperature, as well as prevent the formation of visible and subvisible aggregates. Statistical comparison of the three-index empirical phase diagram (EPD) revealed significant differences in thermal and pH stability signatures between Fab' and PEG-Fab'. Upon mechanical stress, micro-flow imaging (MFI) and measurement of the optical density at 360 nm showed that the PEG-Fab' had significantly higher resistance to surface-induced aggregation compared to the Fab'. Analysis of the interaction parameter, kD, indicated repulsive intermolecular forces for PEG-Fab' and attractive forces for Fab'. In conclusion, PEGylation appears to protect Fab' against thermal and mechanical stress-induced aggregation, likely due to a steric hindrance mechanism.

  15. Management of Tissue Loss After Agkistrodon Snakebite: Appropriate Use of Crotalidae-Fab Antivenin. (United States)

    Larson, Kenneth W; Schaefer, Keith R; Austin, Cindy; Norton, Rhy; Finley, Phillip J


    Although initially created for the treatment of rattlesnake (genus: Crotalus) bites, Crotalidae-Fab antivenin is used to treat many different pit viper envenomations. However, the efficacy of Crotalidae-Fab in preventing tissue loss from copperhead (Agkistrodon contortrix) or cottonmouth (Agkistrodon piscivorus) snakebites remains unclear. Recent reports show that Agkistrodon-related bites rarely require treatment beyond simple observation and pain control. The purpose of this study was to examine the amount of tissue loss in patients who received Crotalidae-Fab compared with those who did not after an Agkistrodon bite. After institutional review board approval, a retrospective study was completed at a Level 1 trauma center. Between 2009 and 2013, a total of 57 snakebites were identified. Of the 57 bites, the snake species was documented in 36 cases including 31 copperheads, 1 cottonmouth, and 4 rattlesnakes. The other 21 bites were from unknown or nonvenomous species. Of the 32 Agkistrodon-related bites, 15 patients received Crotalidae-Fab (average of 3 vials administered) and 17 did not receive Crotalidae-Fab. None of the 32 patients, regardless of treatment option, had tissue loss or required surgical interventions. Only 1 patient received Crotalidae-Fab and debridement of a vesicle associated with the bite. No clinically significant differences were observed between the groups. These findings support previous literature that failed to show added benefit of Crotalidae-Fab treatment for Agkistrodon bites beyond patient comfort and pain control. Evaluation of current protocols for Agkistrodon envenomations is warranted. Snakebite wound education in trauma physicians and nurses may decrease unnecessary use of antivenom medication.

  16. A Substrate Mimic Allows High-Throughput Assay of the FabA Protein and Consequently the Identification of a Novel Inhibitor of Pseudomonas aeruginosa FabA. (United States)

    Moynié, Lucile; Hope, Anthony G; Finzel, Kara; Schmidberger, Jason; Leckie, Stuart M; Schneider, Gunter; Burkart, Michael D; Smith, Andrew D; Gray, David W; Naismith, James H


    Eukaryotes and prokaryotes possess fatty acid synthase (FAS) biosynthetic pathways that comprise iterative chain elongation, reduction, and dehydration reactions. The bacterial FASII pathway differs significantly from human FAS pathways and is a long-standing target for antibiotic development against Gram-negative bacteria due to differences from the human FAS, and several existing antibacterial agents are known to inhibit FASII enzymes. N-Acetylcysteamine (NAC) fatty acid thioesters have been used as mimics of the natural acyl carrier protein pathway intermediates to assay FASII enzymes, and we now report an assay of FabV from Pseudomonas aeruginosa using (E)-2-decenoyl-NAC. In addition, we have converted an existing UV absorbance assay for FabA, the bifunctional dehydration/epimerization enzyme and key target in the FASII pathway, into a high-throughput enzyme coupled fluorescence assay that has been employed to screen a library of diverse small molecules. With this approach, N-(4-chlorobenzyl)-3-(2-furyl)-1H-1,2,4-triazol-5-amine (N42FTA) was found to competitively inhibit (pIC50=5.7±0.2) the processing of 3-hydroxydecanoyl-NAC by P. aeruginosa FabA. N42FTA was shown to be potent in blocking crosslinking of Escherichia coli acyl carrier protein and FabA, a direct mimic of the biological process. The co-complex structure of N42FTA with P. aeruginosa FabA protein rationalises affinity and suggests future design opportunities. Employing NAC fatty acid mimics to develop further high-throughput assays for individual enzymes in the FASII pathway should aid in the discovery of new antimicrobials.

  17. Efecto de la salinidad en la supervivencia embrionaria de puye Galaxias maculatus (Jenyns, 1842

    Directory of Open Access Journals (Sweden)

    Juan Barile


    Full Text Available Se evaluó e l efecto de la salinidad en la supervivencia embrionaria, período embrionario y período de eclosión de puye, Galaxias maculatus. Se utilizaron ocho tratamientos, con salinidades de 0, 4, 8, 12, 16 y 20 obteniéndose supervivencias embrionarias promedios respectivas de 84,9; 84,0; 85,5; 86,1; 82,6 y 75,8%, sin diferencias significativas entre ellas, mientras que con salinidad de 24 se obtuvo sobrevivencia de 10,4%, significativamente diferente al resto de los tratamientos y con salinidad de 28 causó 100% de mortalidad. A diferencia del estado juvenil y adulto, el embrión es incapaz de resistir la salinidad marina y su umbral de tolerancia se encuentra entre salinidades de 24 y 28. Las mayores supervivencias promedios larvales a 10 días de vida fueron con salinidades intermedias de 8, 12 y 16 de 79,3; 80,3 y 74,6% respectivamente, sin diferencias significativas entre ellas, demostrando alta viabilidad del embrión post-eclosión. Los períodos embrionarios promedios (50% más breves fueron a bajas salinidades 0, 4, 8, 12 con 28,7; 28,0; 29,7 y 29,7 días y los más extensos en los tratamientos con salinidades de 16 y 20, con 34,0 y 34,3 días, sin diferencias significativas entre ellos. Los períodos de eclosión promedio (50% con salinidades de 0, 4, 8, 12, 16, 20 y 24 fueron de 5,0; 5,5; 7,5; 8,8; 13,0; 14,3 y 15,8 días respectivamente, sin diferencias significativas entre ellos.

  18. Characterization of appetite-regulating factors in platyfish, Xiphophorus maculatus (Cyprinodontiformes Poeciliidae). (United States)

    Pitts, Paul M; Volkoff, Hélène


    The regulation of energy in fish, like most vertebrates, is a complex process that involves a number of brain and peripheral hormones. These signals include anorexigenic (e.g. cholecystokinin (CCK) and cocaine- and amphetamine-regulated transcript (CART)) as well as orexigenic (e.g. orexin and neuropeptide Y (NPY)) peptides. Platyfish, Xiphophorus maculatus, are freshwater viviparous fish for which little is known about the endocrine mechanisms regulating feeding. In order to elucidate the role of these peptides in the regulation of feeding of platyfish, we examined the effects of peripheral injections of CCK and orexin on feeding behavior and food intake. Injections of CCK decreased both food intake and searching behavior, while injections of orexin increased searching behavior but did not affect food consumption. In order to better characterize these peptides, we examined their mRNA tissue distribution and assessed the effects of a 10-day fast on their brain and intestine expressions in both males and females. CCK, CART, NPY and orexin all show widespread distributions in brain and several peripheral tissues, including intestine and gonads. Fasting induced decreases in both CCK and CART and an increase in orexin mRNA expressions in the brain and a decrease in CCK expression in the intestine, but did not affect either expressions of NPY. There were no significant sex-specific differences in either the behavioral responses to injections or the expression responses to fasting. The widespread distribution and the fasting-induced changes in expression of these peptides suggest that they might have several physiological roles in platyfish, including the regulation of feeding. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Bioindication Potential of the Coleoptera

    Directory of Open Access Journals (Sweden)

    Belitskaya Mariya Nikolaevna


    Full Text Available Different families of Coleoptera ambiguously respond to the pollution of SPZs with industrial emissions. For example, the SPZ of Volgograd aluminum plant has the changing biodiversity of insect communities at different distances from the pollution source. The increasing level of pollution is accompanied by the reduction in species abundance. At a distance of 200 m a special ecological zone with the specific composition of the entomofauna was formed. It is significantly different from other habitats. No Cerambycidae species may survive in the zone of maximum pollution, and the number of Curculionidae species is reduced significantly. The number of Cerambycidae decreases by more than 40 % in the presence of even minimal contamination. The most sensitive bioindicators are represented by such insects as Cerambycidae, Curculionidae and Chrysomelidae. Changes in the indices can be described by the function y = arctan (x, where x is the distance from the pollution source (in meters. The specificity of this function is to identify levels of possible changes of species richness and numerical abundance of communities. On the basis of trigonometric functions describing the changes in the species composition and abundance, the authors offered the method for assessing the quality of the environment in SPZs. The use of three families of insects opens up prospects of differentiation zones of technogenic pressure.

  20. Fast conversion of scFv to Fab antibodies using type IIs restriction enzymes. (United States)

    Sanmark, Hanna; Huovinen, Tuomas; Matikka, Tero; Pettersson, Tiina; Lahti, Maria; Lamminmäki, Urpo


    Single chain variable fragment (scFv) antibody libraries are widely used for developing novel bioaffinity reagents, although Fab or IgG molecules are the preferred antibody formats in many final applications. Therefore, rapid conversion methods for combining multiple DNA fragments are needed to attach constant domains to the scFv derived variable domains. In this study we describe a fast and easy cloning method for the conversion of single framework scFv fragments to Fab fragments using type IIS restriction enzymes. All cloning steps excluding plating of the Fab transformants can be done in 96 well plates and the procedure can be completed in one working day. The concept was tested by converting 69 scFv clones into Fab format on 96 well plates, which resulted in 93% success rate. The method is particularly useful as a high-throughput tool for the conversion of the chosen scFv clones into Fab molecules in order to analyze them as early as possible, as the conversion can significantly affect the binding properties of the chosen clones.

  1. Anti-fouling properties of Fab' fragments immobilized on silane-based adlayers (United States)

    Crivianu-Gaita, Victor; Romaschin, Alexander; Thompson, Michael


    Biosensors require surfaces that are highly specific towards the target analyte and that are minimally fouling. However, surface tuning to minimize fouling is a difficult task. The last decade has seen an increase in the use of immobilized antigen-binding antibody fragments (Fab') in biosensors. One Fab' linker compound S-(11-trichlorosilyl-undecanyl)-benzothiosulfonate (TUBTS) and three spacers were used to create the silane-based adlayers. The ultra-high frequency electromagnetic piezoelectric acoustic sensor (EMPAS) was used to gauge the fouling properties of the various surfaces using bovine serum albumin (BSA), goat IgG, and mouse serum. X-ray photoelectron spectroscopy (XPS), contact angle, and atomic force microscopy (AFM) were employed to characterize the surfaces. It was discovered that immobilized oriented Fab' fragments reduced the fouling levels of surfaces up to 80% compared to the surfaces without fragments. An explanation for this phenomenon is that the antibody fragments increase the hydration of the surfaces and aid in the formation of an anti-fouling water barrier. The anti-fouling effect of the Fab' fragments is at its maximum when there is an even distribution of fragments across the surfaces. Finally, using Fab'-covered surfaces, a cancer biomarker was detected from serum, showing the applicability of this work to the field of biodetection.

  2. Mapping of Fab-1:VEGF Interface Using Carboxyl Group Footprinting Mass Spectrometry (United States)

    Wecksler, Aaron T.; Kalo, Matt S.; Deperalta, Galahad


    A proof-of-concept study was performed to demonstrate that carboxyl group footprinting, a relatively simple, bench-top method, has utility for first-pass analysis to determine epitope regions of therapeutic mAb:antigen complexes. The binding interface of vascular endothelial growth factor (VEGF) and the Fab portion of a neutralizing antibody (Fab-1) was analyzed using carboxyl group footprinting with glycine ethyl ester (GEE) labeling. Tryptic peptides involved in the binding interface between VEGF and Fab-1 were identified by determining the specific GEE-labeled residues that exhibited a reduction in the rate of labeling after complex formation. A significant reduction in the rate of GEE labeling was observed for E93 in the VEGF tryptic peptide V5, and D28 and E57 in the Fab-1 tryptic peptides HC2 and HC4, respectively. Results from the carboxyl group footprinting were compared with the binding interface identified from a previously characterized crystal structure (PDB: 1BJ1). All of these residues are located at the Fab-1:VEGF interface according to the crystal structure, demonstrating the potential utility of carboxyl group footprinting with GEE labeling for mapping epitopes.

  3. High affinity mouse-human chimeric Fab against Hepatitis B surface antigen

    Institute of Scientific and Technical Information of China (English)

    Biplab Bose; Navin Khanna; Subrat K Acharya; Subrata Sinha


    AIM: Passive immunotherapy using antibody against hepatitis B surface antigen (HBsAg) has been advocated in certain cases of Hepatitis B infection. We had earlier reported on the cloning and expression of a high affinity scFv derived from a mouse monoclonal (5S) against HBsAg. However this mouse antibody cannot be used for therapeutic purposes as it may elicit anti-mouse immune responses. Chimerization by replacing mouse constant domains with human ones can reduce the immunogenicity of this antibody.METHODS: We cloned the VH and VL genes of this mouse antibody; and fused them with CH1 domain of human IgG1 and CL domain of human kappa chain respectively. These chimeric genes were cloned into a phagemid vector. After initial screening using the phage display system, the chimeric Fab was expressed in soluble form in E. Coli.RESULTS: The chimeric Fab was purified from the bacterial periplasmic extract. We characterized the chimeric Fab using several in vitro techniques and it was observed that the chimeric molecule retained the high affinity and specificity of the original mouse monoclonal.This chimeric antibody fragment was further expressed in different strains of E> coli to increase the yield.CONCLUSION: We have generated a mouse-human chimeric Fab against HBsAg without any significant loss in binding and epitope specificity. This chimeric Fab fragment can be further modified to generate a fulllength chimeric antibody for therapeutic uses.

  4. Cyclization strategies of meditopes: affinity and diffraction studies of meditope–Fab complexes

    Energy Technology Data Exchange (ETDEWEB)

    Bzymek, Krzysztof P.; Ma, Yuelong; Avery, Kendra A.; Horne, David A.; Williams, John C., E-mail: [Beckman Research Institute of City of Hope, 1710 Flower Street, Duarte, CA 91010 (United States)


    An overview of cyclization strategies of a Fab-binding peptide to maximize affinity. Recently, a unique binding site for a cyclic 12-residue peptide was discovered within a cavity formed by the light and heavy chains of the cetuximab Fab domain. In order to better understand the interactions that drive this unique complex, a number of variants including the residues within the meditope peptide and the antibody, as well as the cyclization region of the meditope peptide, were created. Here, multiple crystal structures of meditope peptides incorporating different cyclization strategies bound to the central cavity of the cetuximab Fab domain are presented. The affinity of each cyclic derivative for the Fab was determined by surface plasmon resonance and correlated to structural differences. Overall, it was observed that the disulfide bond used to cyclize the peptide favorably packs against a hydrophobic ‘pocket’ and that amidation and acetylation of the original disulfide meditope increased the overall affinity ∼2.3-fold. Conversely, replacing the terminal cysteines with serines and thus creating a linear peptide reduced the affinity over 50-fold, with much of this difference being reflected in a decrease in the on-rate. Other cyclization methods, including the formation of a lactam, reduced the affinity but not to the extent of the linear peptide. Collectively, the structural and kinetic data presented here indicate that small perturbations introduced by different cyclization strategies can significantly affect the affinity of the meditope–Fab complex.

  5. Reorienting the Fab domains of trastuzumab results in potent HER2 activators.

    Directory of Open Access Journals (Sweden)

    Justin M Scheer

    Full Text Available The structure of the Fab region of antibodies is critical to their function. By introducing single cysteine substitutions into various positions of the heavy and light chains of the Fab region of trastuzumab, a potent antagonist of HER2, and using thiol chemistry to link the different Fabs together, we produced a variety of monospecific F(ab'(2-like molecules with activities spanning from activation to inhibition of breast tumor cell growth. These isomers (or bis-Fabs of trastuzumab, with varying relative spatial arrangements between the Fv-regions, were able to either promote or inhibit cell-signaling activities through the PI3K/AKT and MAPK pathways. A quantitative phosphorylation mapping of HER2 indicated that the agonistic isomers produced a distinct phosphorylation pattern associated with activation. This study suggests that antibody geometric isomers, found both in nature and during synthetic antibody development, can have profoundly different biological activities independent of their affinities for their target molecules.

  6. Contribution of Antibody Hydrodynamic Size to Vitreal Clearance Revealed through Rabbit Studies Using a Species-Matched Fab. (United States)

    Shatz, Whitney; Hass, Philip E; Mathieu, Mary; Kim, Hok Seon; Leach, Kim; Zhou, Michelle; Crawford, Yongping; Shen, Amy; Wang, Kathryn; Chang, Debby P; Maia, Mauricio; Crowell, Susan R; Dickmann, Leslie; Scheer, Justin M; Kelley, Robert F


    We have developed a tool Fab fragment of a rabbit monoclonal antibody that is useful for early evaluation in rabbit models of technologies for long acting delivery (LAD) of proteins to the eye. Using this Fab we show that vitreal clearance can be slowed through increased hydrodynamic size. Fab (G10rabFab) and Fab' (G10rabFab') fragments of a rabbit monoclonal antibody (G10rabIgG) were expressed in Chinese hamster ovary (CHO) cells and purified using antigen-based affinity chromatography. G10rabFab retains antigen-binding upon thermal stress (37 °C) for 8 weeks in phosphate-buffered saline (PBS) and can be detected in rabbit tissues using an antigen-based ELISA. Hydrodynamic radius, measured using quasi-elastic light scattering (QELS), was increased through site-specific modification of the G10rabFab' free cysteine with linear methoxy-polyethylene glycol(PEG)-maleimide of 20000 or 40000 molecular weight. Pharmacokinetic studies upon intravitreal dosing in New Zealand white rabbits were conducted on the G10rabFab and PEGylated G10rabFab'. Results of single and multidose pharmacokinetic experiments yield reproducible results and a vitreal half-life for G10rabFab of 3.2 days. Clearance from the eye is slowed through increased hydrodynamic size, with vitreal half-life showing a linear dependence on hydrodynamic radius (RH). A linear dependence of vitreal half-life on RH suggests that molecule diffusivity makes an important contribution to vitreal clearance. A method for prediction of vitreal half-life from RH measurements is proposed.

  7. Structural Characterisation of FabG from Yersinia pestis, a Key Component of Bacterial Fatty Acid Synthesis. (United States)

    Nanson, Jeffrey D; Forwood, Jade K


    Ketoacyl-acyl carrier protein reductases (FabG) are ubiquitously expressed enzymes that catalyse the reduction of acyl carrier protein (ACP) linked thioesters within the bacterial type II fatty acid synthesis (FASII) pathway. The products of these enzymes, saturated and unsaturated fatty acids, are essential components of the bacterial cell envelope. The FASII reductase enoyl-ACP reductase (FabI) has been the focus of numerous drug discovery efforts, some of which have led to clinical trials, yet few studies have focused on FabG. Like FabI, FabG appears to be essential for survival in many bacteria, similarly indicating the potential of this enzyme as a drug target. FabG enzymes are members of the short-chain alcohol dehydrogenase/reductase (SDR) family, and like other SDRs, exhibit highly conserved secondary and tertiary structures, and contain a number of conserved sequence motifs. Here we describe the crystal structures of FabG from Yersinia pestis (YpFabG), the causative agent of bubonic, pneumonic, and septicaemic plague, and three human pandemics. Y. pestis remains endemic in many parts of North America, South America, Southeast Asia, and Africa, and a threat to human health. YpFabG shares a high degree of structural similarity with bacterial homologues, and the ketoreductase domain of the mammalian fatty acid synthase from both Homo sapiens and Sus scrofa. Structural characterisation of YpFabG, and comparison with other bacterial FabGs and the mammalian fatty acid synthase, provides a strong platform for virtual screening of potential inhibitors, rational drug design, and the development of new antimicrobial agents to combat Y. pestis infections.

  8. Structural Characterisation of FabG from Yersinia pestis, a Key Component of Bacterial Fatty Acid Synthesis.

    Directory of Open Access Journals (Sweden)

    Jeffrey D Nanson

    Full Text Available Ketoacyl-acyl carrier protein reductases (FabG are ubiquitously expressed enzymes that catalyse the reduction of acyl carrier protein (ACP linked thioesters within the bacterial type II fatty acid synthesis (FASII pathway. The products of these enzymes, saturated and unsaturated fatty acids, are essential components of the bacterial cell envelope. The FASII reductase enoyl-ACP reductase (FabI has been the focus of numerous drug discovery efforts, some of which have led to clinical trials, yet few studies have focused on FabG. Like FabI, FabG appears to be essential for survival in many bacteria, similarly indicating the potential of this enzyme as a drug target. FabG enzymes are members of the short-chain alcohol dehydrogenase/reductase (SDR family, and like other SDRs, exhibit highly conserved secondary and tertiary structures, and contain a number of conserved sequence motifs. Here we describe the crystal structures of FabG from Yersinia pestis (YpFabG, the causative agent of bubonic, pneumonic, and septicaemic plague, and three human pandemics. Y. pestis remains endemic in many parts of North America, South America, Southeast Asia, and Africa, and a threat to human health. YpFabG shares a high degree of structural similarity with bacterial homologues, and the ketoreductase domain of the mammalian fatty acid synthase from both Homo sapiens and Sus scrofa. Structural characterisation of YpFabG, and comparison with other bacterial FabGs and the mammalian fatty acid synthase, provides a strong platform for virtual screening of potential inhibitors, rational drug design, and the development of new antimicrobial agents to combat Y. pestis infections.

  9. Evidence of iteroparity in the widely distributed diadromous fish inanga Galaxias maculatus and potential implications for reproductive output. (United States)

    Stevens, J C B; Hickford, M J H; Schiel, D R


    Gaps in understanding variability among populations of inanga Galaxias maculatus in the timing of reproduction were addressed in southern New Zealand (NZ), where G. maculatus constitutes a declining fishery. Reproductive activity was delayed by 1 month on the west coast compared with the east coast and the west coast spawning season was prolonged into winter. The evidence for post-spawning survival of some fish was unequivocal from histological studies. These older and larger fish contributed disproportionately to egg production. Estimates of fecundity were considerably lower than those previously calculated for NZ populations. The importance of quality habitats being available during critical life history periods are highlighted. It was apparent that some streams supported fish that were larger and in better condition and that this translated into greatly increased fecundity. Future research should focus on whether this is a legacy of these fish experiencing better pre-settlement marine habitat as larvae, or higher quality instream habitat enhancing the growth and development of adults.

  10. Potential of the Lectin/Inhibitor Isolated from Crataeva tapia Bark (CrataBL) for Controlling Callosobruchus maculatus Larva Development. (United States)

    Nunes, Natalia N S; Ferreira, Rodrigo S; Silva-Lucca, Rosemeire A; de Sá, Leonardo F R; de Oliveira, Antônia Elenir A; Correia, Maria Tereza dos S; Paiva, Patrícia Maria G; Wlodawer, Alexander; Oliva, Maria Luiza V


    Callosobruchus maculatus is an important predator of cowpeas. Due to infestation during storage, this insect affects the quality of seed and crop yield. This study aimed to investigate the effects of CrataBL, a multifunction protein isolated from Crataeva tapia bark, on C. maculatus larva development. The protein, which is stable even in extreme pH conditions, showed toxic activity, reducing the larval mass 45 and 70% at concentrations of 0.25 and 1.0% (w/w), respectively. Acting as an inhibitor, CrataBL decreased by 39% the activity of cysteine proteinases from larval gut. Conversely, the activity of serine proteinases was increased about 8-fold. The toxic properties of CrataBL may also be attributed to its capacity of binding to glycoproteins or glycosaminoglycans. Such binding interferes with larval metabolism, because CrataBL-FITC was found in the fat body, Malpighian tubules, and feces of larvae. These results demonstrate the potential of this protein for controlling larva development.

  11. Host preference of Callosobruchus maculatus: a comparison of life history characteristics for three strains of beetles on two varieties of cowpea

    NARCIS (Netherlands)

    Boeke, S.J.; Loon, van J.J.A.; Huis, van A.; Dicke, M.


    The reproductive success of Callosobruchus maculatus Fabricius, the main insect pest of stored cowpea, may vary between strains of this beetle and between varieties of the host seeds. Life history parameters of beetle strains from three different origins in West Africa were compared on two susceptib

  12. Targeting human prostate cancer with (111) In-labeled D2B IgG, F(ab')2 and Fab fragments in nude mice with PSMA-expressing xenografts

    NARCIS (Netherlands)

    Lutje, S.; Rij, C.M. van; Franssen, G.M.; Fracasso, G.; Helfrich, W.; Eek, A.; Oyen, W.J.G.; Colombatti, M.; Boerman, O.C.


    D2B is a new monoclonal antibody directed against an extracellular domain of prostate-specific membrane antigen (PSMA), which is overexpressed in prostate cancer. The potential of D2B IgG, and F(ab')2 and Fab fragments of this antibody for targeting prostate cancer was determined in mice bearing sub

  13. Targeting human prostate cancer with In-111-labeled D2B IgG, F(ab ')(2) and Fab fragments in nude mice with PSMA-expressing xenografts

    NARCIS (Netherlands)

    Lutje, Susanne; van Rij, Catharina M.; Franssen, Gerben M.; Fracasso, Giulio; Helfrich, Wijnand; Eek, Annemarie; Oyen, Wim J.; Colombatti, Marco; Boerman, Otto C.


    D2B is a new monoclonal antibody directed against an extracellular domain of prostate-specific membrane antigen (PSMA), which is overexpressed in prostate cancer. The potential of D2B IgG, and F(ab)(2) and Fab fragments of this antibody for targeting prostate cancer was determined in mice bearing su

  14. Treatment of digitalis intoxication with emphasis on the clinical use of digoxin immune Fab. (United States)

    Allen, N M; Dunham, G D


    Many studies and cases of digitalis intoxication have been reported since the time of William Withering's first publication in 1785. Recognition and management of digitalis toxicity is challenging. Before digoxin immune Fab was commercially available, treatment consisted of managing the signs and symptoms of toxicity until the digitalis was eliminated. Digoxin immune Fab offers a safe, effective, and specific method of quickly reversing digitalis toxicity. Factors that must be considered with the clinical use of this agent include the dosage calculation, administration technique, postdose monitoring, pharmacokinetics, mechanism of action, interference with commercially available digoxin assays, partial neutralizing dosing, rebound of free digoxin, and indications for use. For severe, life-threatening toxicity, digoxin immune Fab is the treatment of choice.

  15. Structure of Rotavirus Outer-Layer Protein VP7 Bound with a Neutralizing Fab

    Energy Technology Data Exchange (ETDEWEB)

    Aoki, Scott T.; Settembre, Ethan C.; Trask, Shane D.; Greenberg, Harry B.; Harrison, Stephen C.; Dormitzer, Philip R.; (Stanford-MED); (CH-Boston)


    Rotavirus outer-layer protein VP7 is a principal target of protective antibodies. Removal of free calcium ions (Ca{sup 2+}) dissociates VP7 trimers into monomers, releasing VP7 from the virion, and initiates penetration-inducing conformational changes in the other outer-layer protein, VP4. We report the crystal structure at 3.4 angstrom resolution of VP7 bound with the Fab fragment of a neutralizing monoclonal antibody. The Fab binds across the outer surface of the intersubunit contact, which contains two Ca{sup 2+} sites. Mutations that escape neutralization by other antibodies suggest that the same region bears the epitopes of most neutralizing antibodies. The monovalent Fab is sufficient to neutralize infectivity. We propose that neutralizing antibodies against VP7 act by stabilizing the trimer, thereby inhibiting the uncoating trigger for VP4 rearrangement. A disulfide-linked trimer is a potential subunit immunogen.

  16. Anti-inflammatory activity of human IgG4 antibodies by dynamic Fab arm exchange. (United States)

    van der Neut Kolfschoten, Marijn; Schuurman, Janine; Losen, Mario; Bleeker, Wim K; Martínez-Martínez, Pilar; Vermeulen, Ellen; den Bleker, Tamara H; Wiegman, Luus; Vink, Tom; Aarden, Lucien A; De Baets, Marc H; van de Winkel, Jan G J; Aalberse, Rob C; Parren, Paul W H I


    Antibodies play a central role in immunity by forming an interface with the innate immune system and, typically, mediate proinflammatory activity. We describe a novel posttranslational modification that leads to anti-inflammatory activity of antibodies of immunoglobulin G, isotype 4 (IgG4). IgG4 antibodies are dynamic molecules that exchange Fab arms by swapping a heavy chain and attached light chain (half-molecule) with a heavy-light chain pair from another molecule, which results in bispecific antibodies. Mutagenesis studies revealed that the third constant domain is critical for this activity. The impact of IgG4 Fab arm exchange was confirmed in vivo in a rhesus monkey model with experimental autoimmune myasthenia gravis. IgG4 Fab arm exchange is suggested to be an important biological mechanism that provides the basis for the anti-inflammatory activity attributed to IgG4 antibodies.

  17. 3D printing in social education: Eki-Fab and student PBL (United States)

    Makino, Masato; Saito, Azusa; Kodama, Mai; Takamatsu, Kyuuichiro; Tamate, Hideaki; Sakai, Kazuyuki; Wada, Masato; Khosla, Ajit; Kawakami, Masaru; Furukawa, Hidemitsu


    Additive manufacturing or 3D printer is one of the most innovative material processing methods. We are considering that human resources for 3D printing would be needed in the future. To educate the abilities of the digital fabrication, we have the public digital fabrication space "Eki-Fab" for junior and high school students and Project Based Learning (PBL) class for undergraduate students. Eki-Fab is held on every Saturday at the Yonezawa train station. In the "Eki-Fab", anybody can study the utilizing of 3D printer and modeling technics under the instruction of staff in Yamagata University. In the PBL class, we have the class every Thursday. The students get the techniques of the digital fabrication through the PBL.

  18. Human combinatorial Fab library yielding specific and functional antibodies against the human fibroblast growth factor receptor 3. (United States)

    Rauchenberger, Robert; Borges, Eric; Thomassen-Wolf, Elisabeth; Rom, Eran; Adar, Rivka; Yaniv, Yael; Malka, Michael; Chumakov, Irina; Kotzer, Sarit; Resnitzky, Dalia; Knappik, Achim; Reiffert, Silke; Prassler, Josef; Jury, Karin; Waldherr, Dirk; Bauer, Susanne; Kretzschmar, Titus; Yayon, Avner; Rothe, Christine


    The human combinatorial antibody library Fab 1 (HuCAL-Fab 1) was generated by transferring the heavy and light chain variable regions from the previously constructed single-chain Fv library (Knappik, A., Ge, L., Honegger, A., Pack, P., Fischer, M., Wellnhofer, G., Hoess, A., Wölle, J., Plückthun, A., and Virnekäs, B. (2000) J. Mol. Biol. 296, 57-86), diversified in both complementarity-determining regions 3 into a novel Fab display vector, yielding 2.1 x 10(10) different antibody fragments. The modularity has been retained in the Fab display and screening plasmids, ensuring rapid conversion into various antibody formats as well as antibody optimization using prebuilt maturation cassettes. HuCAL-Fab 1 was challenged against the human fibroblast growth factor receptor 3, a potential therapeutic antibody target, against which, to the best of our knowledge, no functional antibodies could be generated so far. A unique screening mode was designed utilizing recombinant functional proteins and cell lines differentially expressing fibroblast growth factor receptor isoforms diversified in expression and receptor dependence. Specific Fab fragments with subnanomolar affinities were isolated by selection without any maturation steps as determined by fluorescence flow cytometry. Some of the selected Fab fragments completely inhibit target-mediated cell proliferation, rendering them the first monoclonal antibodies against fibroblast growth factor receptors having significant function blocking activity. This study validates HuCAL-Fab 1 as a valuable source for the generation of target-specific antibodies for therapeutic applications.

  19. Enoyl-Acyl Carrier Protein Reductase I (FabI) Is Essential for the Intracellular Growth of Listeria monocytogenes (United States)

    Ericson, Megan E.; Frank, Matthew W.


    Enoyl-acyl carrier protein reductase catalyzes the last step in each elongation cycle of type II bacterial fatty acid synthesis and is a key regulatory protein in bacterial fatty acid synthesis. Genes of the facultative intracellular pathogen Listeria monocytogenes encode two functional enoyl-acyl carrier protein isoforms based on their ability to complement the temperature-sensitive growth phenotype of Escherichia coli strain JP1111 [fabI(Ts)]. The FabI isoform was inactivated by the FabI selective inhibitor AFN-1252, but the FabK isoform was not affected by the drug, as expected. Inhibition of FabI by AFN-1252 decreased endogenous fatty acid synthesis by 80% and lowered the growth rate of L. monocytogenes in laboratory medium. Robust exogenous fatty acid incorporation was not detected in L. monocytogenes unless the pathway was partially inactivated by AFN-1252 treatment. However, supplementation with exogenous fatty acids did not restore normal growth in the presence of AFN-1252. FabI inactivation prevented the intracellular growth of L. monocytogenes, showing that neither FabK nor the incorporation of host cellular fatty acids was sufficient to support the intracellular growth of L. monocytogenes. Our results show that FabI is the primary enoyl-acyl carrier protein reductase of type II bacterial fatty acid synthesis and is essential for the intracellular growth of L. monocytogenes. PMID:27736774

  20. 20 CFR 30.908 - How will the FAB evaluate new medical evidence submitted to challenge the impairment... (United States)


    ... 20 Employees' Benefits 1 2010-04-01 2010-04-01 false How will the FAB evaluate new medical... Medical Evidence of Impairment § 30.908 How will the FAB evaluate new medical evidence submitted to... impairment evaluation that differs from the impairment evaluation relied upon by the district office, the...

  1. The unfolding/denaturation of immunogammaglobulin of isotype 2b and its F-ab and F-c fragments

    NARCIS (Netherlands)

    Vermeer, AWP; Norde, W; van Amerongen, A


    The unfolding and further denaturation of IgG and its F-ab and F-c fragments were studied both on a macroscopic and molecular level, using differential scanning calorimetry and circular dichroism spectroscopy, respectively. It was shown that the structural integrity of the F-ab and F-c units was ret

  2. Enoyl-Acyl Carrier Protein Reductase I (FabI) Is Essential for the Intracellular Growth of Listeria monocytogenes. (United States)

    Yao, Jiangwei; Ericson, Megan E; Frank, Matthew W; Rock, Charles O


    Enoyl-acyl carrier protein reductase catalyzes the last step in each elongation cycle of type II bacterial fatty acid synthesis and is a key regulatory protein in bacterial fatty acid synthesis. Genes of the facultative intracellular pathogen Listeria monocytogenes encode two functional enoyl-acyl carrier protein isoforms based on their ability to complement the temperature-sensitive growth phenotype of Escherichia coli strain JP1111 [fabI(Ts)]. The FabI isoform was inactivated by the FabI selective inhibitor AFN-1252, but the FabK isoform was not affected by the drug, as expected. Inhibition of FabI by AFN-1252 decreased endogenous fatty acid synthesis by 80% and lowered the growth rate of L. monocytogenes in laboratory medium. Robust exogenous fatty acid incorporation was not detected in L. monocytogenes unless the pathway was partially inactivated by AFN-1252 treatment. However, supplementation with exogenous fatty acids did not restore normal growth in the presence of AFN-1252. FabI inactivation prevented the intracellular growth of L. monocytogenes, showing that neither FabK nor the incorporation of host cellular fatty acids was sufficient to support the intracellular growth of L. monocytogenes Our results show that FabI is the primary enoyl-acyl carrier protein reductase of type II bacterial fatty acid synthesis and is essential for the intracellular growth of L. monocytogenes. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  3. Prokaryotic expression and renaturation of engineering chimeric Fab antibody against human hepatoma

    Institute of Scientific and Technical Information of China (English)

    Jin-Liang Xing; Xiang-Min Yang; Xi-Ying Yao; Fei Song; Zhi-Nan Chen


    AIM: To express chimeric Fd (cFd) and chimeric light chain (cL) in E.coli respectively and refold them into chimeric Fab (cFab) antibody.METHODS: cFd and cL genes were respectively inserted into the prokaryotic expression vector pET32a to construct recombinant vectors pET32a/cFd and pET32a/cL. Then,the competent E. colicells were transformed by the recombinant vectors and induced by IPTG. Moreover, a large quantity of cFd and cL expression products were prepared and mixed with equal molar to refold into cFab by gradient dialysis. The refolded products were identified and analyzed by sodium SDS-PAGE, Western blotting,ELISA and HPLC.RESULTS: High efficient prokaryotic expressions of both cFd and cL in the form of non-fusion protein were obtained with the expression levels of 28.3% and 32.3% of total bacteria proteins, respectively. Their relative molecular masses were all 24 ku or so, and both of them mainly existed in the form of inclusion bodies. In addition, cFd and cL were successfully refolded into cFab by gradient dialysis, with about 59.45% of recovery when the starting total protein concentration was 100 μg/mL. The renatured cFab could specifically bind to related antigen with high affinity.CONCLUSION: The cFab antibody against human hepatoma was highly and efficiently expressed and refolded, which laid a solid foundation for studying its application in the treatment of hepatoma.

  4. Effects of Initial Infestation Levels ofCallosobruchus maculatus(F.) (Coleoptera:Chrysomelidae) on Cowpea and Use ofNicotiana tabacum L. Aqueous Extract as Grain Protectant

    Institute of Scientific and Technical Information of China (English)

    Musa A K; Odunayo A; Adeyeye O E


    This study determined the effects of initial infestation of cowpea seeds (Ife brown variety) with different insect densities (0, 2, 4 and 6 pairs per 50 g seeds) ofCallosobruchus maculatus(F.) and evaluated the effects of aqueous leaf extract of Nicotiana tabacum L. onC. maculatus in the laboratory. It was observed that adult beetle population increased significantly (p<0.05) with increase in insect density. The increase in population of beetles and corresponding weight loss of the seeds in different levels of infestation showed that the cowpea variety was susceptible to beetle infestation, emergence and survival of progeny. Significantly more adults emerged on higher infestation compared to lower and no infestation. In Nigeria,Nicotiana tabacum L. is a locally available plant, with known insecticidal properties. The plant leaf extract was easily extracted with water and confirmed its effectiveness as a protective agent for stored cowpea seeds. Experiment was conducted to assess the effects of aqueous extracts of N. tabacumat 0, 0.1, 0.2 and 0.3 mL•50 g-1 cowpea seeds onC. maculatus. Data was recorded and showed varying levels of effectiveness againstC. maculatus. Result showed that seed appearance was dependent on levels of insect population, while N. tabacum aqueous extract exerted effects on survival ofC. maculatus. Aqueous leaf extract ofN. tabacum probably contained some insecticidal properties which might have significantly conferred beetle mortality and reduced beetle emergence leading to a decrease in seed weight loss.

  5. High contrast tumor imaging with radio-labeled antibody Fab fragments tailored for optimized pharmacokinetics via PASylation. (United States)

    Mendler, Claudia T; Friedrich, Lars; Laitinen, Iina; Schlapschy, Martin; Schwaiger, Markus; Wester, Hans-Jürgen; Skerra, Arne


    Although antigen-binding fragments (Fabs) of antibodies constitute established tracers for in vivo radiodiagnostics, their functionality is hampered by a very short circulation half-life. PASylation, the genetic fusion with a long, conformationally disordered amino acid chain comprising Pro, Ala and Ser, provides a convenient way to expand protein size and, consequently, retard renal filtration. Humanized αHER2 and αCD20 Fabs were systematically fused with 100 to 600 PAS residues and produced in E. coli. Cytofluorimetric titration analysis on tumor cell lines confirmed that antigen-binding activities of the parental antibodies were retained. The radio-iodinated PASylated Fabs were studied by positron emission tomography (PET) imaging and biodistribution analysis in mouse tumor xenograft models. While the unmodified αHER2 and αCD20 Fabs showed weak tumor uptake (0.8% and 0.2% ID/g, respectively; 24 h p.i.) tumor-associated radioactivity was boosted with increasing PAS length (up to 9 and 26-fold, respectively), approaching an optimum for Fab-PAS400. Remarkably, 6- and 5-fold higher tumor-to-blood ratios compared with the unmodified Fabs were measured in the biodistribution analysis (48 h p.i.) for αHER2 Fab-PAS100 and Fab-PAS200, respectively. These findings were confirmed by PET studies, showing high imaging contrast in line with tumor-to-blood ratios of 12.2 and 5.7 (24 h p.i.) for αHER2 Fab-PAS100 and Fab-PAS200. Even stronger tumor signals were obtained with the corresponding αCD20 Fabs, both in PET imaging and biodistribution analysis, with an uptake of 2.8% ID/g for Fab-PAS100 vs. 0.24% ID/g for the unmodified Fab. Hence, by engineering Fabs via PASylation, plasma half-life can be tailored to significantly improve tracer uptake and tumor contrast, thus optimally matching reagent/target interactions.

  6. Research advances on Fabs antibodies%Fab类抗体的研究进展

    Institute of Scientific and Technical Information of China (English)

    刘美君; 高向东; 徐晨


    In genetic engineering antibodies,Fabs antibodies have many advantages,such as lower relative molecular mass, specific tissue distribution and lower immunogenicity, which have made Fabs antibodies the research hotspot for the past few decades. Besides the widely used enzymatic generation, multiple expression systems which include E. coli. and insects are used in the generation of Fabs antibodies. E. coli. expression system is most thoroughly explored and has been used in the preparation of clinical Fabs drugs. There are six Fab drugs that have been approved by FDA, such as certolizumabpegol and many Fabs are still in clinical research. This review discusses the recent research progress of Fabs drugs, including Fabs′ structure and characteristics, the generation and expression of Fabs, strategies to increase production and clinical applications, with the emphasis on generation, expression and clinical applications.%在基因工程抗体中,Fab类(Fabs)抗体有许多优势,如相对分子质量小、组织分布特异性强和免疫原性低等,使Fab类抗体成为近几十年来的研究热点。除了应用较广泛的体外酶消化法外,多种表达系统(如大肠杆菌和昆虫等)皆用于Fab类抗体的获得,其中大肠杆菌表达的系统研究最为彻底,且已经应用于制备Fab类临床药物。目前已经有赛妥珠单抗等6种Fab类抗体药物通过了美国FDA申请并上市,并有多种Fab类药物正处于临床试验阶段。本文分别对Fab类抗体的基本结构及特点、Fab类抗体的生成、表达和提高产量的策略、临床应用的研究进展进行介绍。

  7. International Foot and Ankle Biomechanics Community (i-FAB: past, present and beyond

    Directory of Open Access Journals (Sweden)

    Rosenbaum Dieter


    Full Text Available Abstract The International Foot and Ankle Biomechanics Community (i-FAB is an international collaborative activity which will have an important impact on the foot and ankle biomechanics community. It was launched on July 2nd 2007 at the foot and ankle session of the International Society of Biomechanics (ISB meeting in Taipei, Taiwan. i-FAB is driven by the desire to improve our understanding of foot and ankle biomechanics as it applies to health, disease, and the design, development and evaluation of foot and ankle surgery, and interventions such as footwear, insoles and surfaces.

  8. High quality mask storage in an advanced Logic-Fab (United States)

    Jähnert, Carmen; Fritsche, Silvio


    High efficient mask logistics as well as safe and high quality mask storage are essential requirements within an advanced lithography area of a modern logic waferfab. Fast operational availability of the required masks at the exposure tool with excellent mask condition requires a safe mask handling, safeguarding of high mask quality over the whole mask usage time without any quality degradation and an intelligent mask logistics. One big challenge is the prevention of haze on high advanced phase shift masks used in a high volume production line for some thousands of 248nm or 193nm exposures. In 2008 Infineon Dresden qualified a customer specific developed semi-bare mask storage system from DMSDynamic Micro Systems in combination with a high advanced mask handling and an interconnected complex logistic system. This high-capacity mask storage system DMS M1900.22 for more than 3000 masks with fully automated mask and box handling as well as full-blown XCDA purge has been developed and adapted to the Infineon Lithotoollandscape using Nikon and SMIF reticle cases. Advanced features for ESD safety and mask security, mask tracking via RFID and interactions with the exposure tools were developed and implemented. The stocker is remote controlled by the iCADA-RSM system, ordering of the requested mask directly from the affected exposure tool allows fast access. This paper discusses the advantages and challenges for this approach as well as the practical experience gained during the implementation of the new system which improves the fab performance with respect to mask quality, security and throughput. Especially the realization of an extremely low and stable humidity level in addition with a well controlled air flow at each mask surface, preventing masks from haze degradation and particle contamination, turns out to be a notable technical achievement. The longterm stability of haze critical masks has been improved significantly. Relevant environmental parameters like

  9. AGILE integration into APC for high mix logic fab (United States)

    Gatefait, M.; Lam, A.; Le Gratiet, B.; Mikolajczak, M.; Morin, V.; Chojnowski, N.; Kocsis, Z.; Smith, I.; Decaunes, J.; Ostrovsky, A.; Monget, C.


    mix logic Fab) in term of product and technology portfolio AGILE corrects for up to 120nm of product topography error on process layer with less than 50nm depth of focus Based on tool functionalities delivered by ASML and on high volume manufacturing requirement, AGILE integration is a real challenge. Regarding ST requirements "Automatic AGILE" functionality developed by ASML was not a turnkey solution and a dedicated functionality was needed. A "ST homemade AGILE integration" has been fully developed and implemented within ASML and ST constraints. This paper describes this integration in our Advanced Process Control platform (APC).

  10. 20 CFR 30.317 - Can the FAB request a further response from the claimant or return a claim to the district office? (United States)


    ... 20 Employees' Benefits 1 2010-04-01 2010-04-01 false Can the FAB request a further response from....317 Can the FAB request a further response from the claimant or return a claim to the district office? At any time before the issuance of its final decision, the FAB may request that the claimant...

  11. 20 CFR 30.312 - What will the FAB do if the claimant objects to the recommended decision but does not request a... (United States)


    ... 20 Employees' Benefits 1 2010-04-01 2010-04-01 false What will the FAB do if the claimant objects....312 What will the FAB do if the claimant objects to the recommended decision but does not request a... period of time allotted in § 30.310 but does not request a hearing, the FAB will consider any...

  12. Structural analysis of the Pimelodus maculatus (Lacépède, 1803 embryogenesis (Siluriformes: Pimelodidae

    Directory of Open Access Journals (Sweden)

    Hellen Buzollo

    Full Text Available The fish embryonic development comprises the events between the egg fertilization up to larvae hatching, being useful for the identification of viable eggs in productivity and survival studies as well as in raising experiments of several species. The goal of the present study was to characterize the embryonic development of Pimelodus maculatus (Siluriformes; Pimelodidae. The embryogenesis was typical of teleosteans, but with differences in relation to other species such as duration of development, type of blastocoel, moment of somite segmentation among others. Six stages of embryonic development were defined: zygote, cleavage, blastula, gastrula, organogenesis (divided in phases: early segmentation and late segmentation and hatching with a period of incubation equal to 13 hours at 29 ºC and 17 hours at 25 ºC. The extruded oocytes presented a mean diameter of 812 µm before and 1066 µm after hydration. When fertilized, they presented a yellowish coloration and a gelatinous layer surrounding the chorion. The cleavage pattern is described as: 2; 4; 8 (4x2; 16 (4x4; 32 (4x8 and 64 (2x4x8 blastomeres up to morula phase (+64 cells. It was also possible to observe at this phase, the beginning of the formation of the yolk syncyctial layer (YSL. Afterwards, the blastula and gastrula stages followed. The end of gastrula was characterized by the formation of the yolk plug. Subsequently, the differentiation between cephalic and caudal regions began, along with the embryo elongation, structuring of optic, Kupffer's and otic vesicles besides a previously unidentified structure in the yolk syncyctial layer. The end of this stage is typified by the tail detachment. The late segmentation phase was distinguished by a free tail, presence of more than 30 somites, optic and otic vesicles, development of posterior intestine, pigmentation of cephalic and caudal regions of yolk sac and embryo growth. The recently-hatched larvae presented a primordial digestive tract

  13. Heavy metals in carabids (Coleoptera, Carabidae

    Directory of Open Access Journals (Sweden)

    Ruslan Butovsky


    Full Text Available Carabid beetles (Coleoptera, Carabidae are one of the most studied soil groups in relation to heavy metal (HM accumulation and use for bioindication of environmental pollution. Accumulation of Zn and Cu in carabid beetles was species-, sex- and trophic group-specific. No differences were found in HM contents between omnivorous and carnivorous species. The use of carabid beetles as indicators of HM accumulation appears to be rather limited.

  14. New longhorn beetles (Coleoptera: Cerambycidae from Serbia

    Directory of Open Access Journals (Sweden)

    Pil Nataša


    Full Text Available The most recent data (Ilić, 2005 indicate the presence of 245 longhorn beetle species (Coleoptera: Cerambycidae in Serbia. Not included in the mentioned publication, the following five species should be added to the list: Cortodera discolor Fairmaire, 1866; Stenopterus similatus Holzschuh 1979; Chlorophorus aegyptiacus (Fabricius, 1775; Agapanthia osmanlis (Reiche, 1858; Agapanthia maculicornis (Gyllenhal, 1817 (Pil and Stojanović in press. A total number of 250 species are presently known for the Serbian longhorn beetle fauna.

  15. Ostrea puelchana (D'Orbigny 1842: a new host of Tumidotheres maculatus (Say, 1818 in northern Patagonia, Argentina Ostrea puelchana D’Orbigny 1842 nuevo hospedador de Tumidotheres maculatus (Say, 1818 en el norte de Patagonia, Argentina

    Directory of Open Access Journals (Sweden)

    María del Socorro Doldan


    Full Text Available Tumidotheres maculatus has been reported as a commensal of Mytilus platensis and Atrina seminuda and suggested as a parasite of Aequipecten tehuelchus in San Matías Gulf (SMG. The native flat oyster, Ostrea puelchana, has been studied in SMG for decades, and no crab-oyster association has been reported. In autumn-spring 2009 and 2010, oysters were collected from four banks in SMG, and the occurrence of T. maculatus inside the pallial cavity of O. puelchana was recorded. The prevalence varied between sites, with 0-12.5% at El Sótano, 0-16.7% at Caleta de los Loros, and no crabs found in the samples from Las Grutas and Puerto Lobos. Infected oysters hosted a single crab. Oysters were infested by either one male (47.7% or one female (15.1%. A positive but low correlation was found for male crab size and oyster size. The presence of these crabs inside the oysters could be due to the overlap of the distribution areas of O. puelchana and other bivalve hosts. Our findings may also be a consequence of local fishing. Commercial extraction of traditional hosts may have caused crabs to look for new hosts.En el Golfo San Matías (GSM, Tumidotheres maculatus ha sido registrado como comensal de Mytilus platensis y Atrina seminuda, y se lo ha indicado como parásito de Aequipecten tehuelchus. La ostra plana nativa Ostrea puelchana ha sido objeto de estudio por décadas en el GSM y no existen precedentes sobre una asociación entre las ostras y el cangrejo pinotérido. Durante otoño-primavera de 2009 y de 2010 se recolectaron ostras de cuatro bancos del GSM, registrándose la ocurrencia de T. maculatus en la cavidad paleal de O. puelchana. La prevalencia varió entre los sitios: 0-12,5% en El Sótano, 0-16,7% en Caleta de los Loros; no se encontraron cangrejos en los muestreos en Las Grutas y en Puerto Lobos. Se encontró un único cangrejo por hospedador; las ostras estuvieron infestadas por cangrejos macho (47,7%, o hembras (15,1%. Se encontró una

  16. Estolomimus (Coleoptera, Cerambycidae, Desmiphorini: novas espécies e chave Estolomimus (Coleoptera, Cerambycidae, Desmiphorini: new species and key

    Directory of Open Access Journals (Sweden)

    Ubirajara R. Martins

    Full Text Available New species described from Brazil: Estolomimus maculatus (Minas Gerais and São Paulo, E. transversus (Espírito Santo, E. lichenophorus (São Paulo, and from Bolivia: E. abjunctus (Beni. A key to the species is added.

  17. Efeitos de extratos alcoólicos de plantas sobre o caruncho do feijão vigna (Callosobruchus maculatus Effect of alcoholic extract of plants on weevil of cowpea (Callosobruchus maculatus

    Directory of Open Access Journals (Sweden)

    Francisco de A. C. Almeida


    Full Text Available Através de três métodos, extratos vegetais foram aplicados, ao Callosobruchus maculatus na fase adulta, inoculados ou não em uma massa de sementes, e na fase imatura (ovo com o objetivo de se controlar esta praga do feijão armazenado. Utilizaram-se flores, folhas, frutos e caule secos de oito espécies vegetais na obtenção dos extratos, em percolador, com solvente álcool etílico (70%. O delineamento experimental utilizado foi o inteiramente ao acaso, com os tratamentos distribuídos em esquema fatorial, cujos fatores quantitativos foram revelados pela regressão na análise de variância. Mediante os resultados obtidos, concluiu-se que a mortalidade dos insetos está relacionada com o tipo de extrato, os métodos de aplicação e com a dosagem aplicada, sendo os extratos de Callopogonium caeruleum e Piper nigrum os mais eficientes no controle do caruncho de feijão.Vegetable extracts were applied, through three methods, to the Callosobruchus maculatus in the adult phase, inoculated or not in a mass of seeds, in the immature phase (egg with the objective of controlling this pest of the stored beans. Dry flowers, leaves, fruits and dry stems of eight vegetable species were used to obtain the extracts in an extractor, with ethyl alcohol (70%. A completely randomized statistical design was used with the treatments distributed in a factorial scheme, the quantitative factors were analysed by the regression in the variance analysis. From the results obtained, it was concluded that the mortality of the insects is related to the extract type, the application methods and the applied dose, being the extracts of Callopogonium caeruleum and Piper nigrum the most efficient in the control of the weevil of cowpea.

  18. Structure-activity studies of the inhibition of FabI, the enoyl reductase from Escherichia coli, by triclosan: kinetic analysis of mutant FabIs. (United States)

    Sivaraman, Sharada; Zwahlen, Jacque; Bell, Alasdair F; Hedstrom, Lizbeth; Tonge, Peter J


    Triclosan, a common antibacterial additive used in consumer products, is an inhibitor of FabI, the enoyl reductase enzyme from type II bacterial fatty acid biosynthesis. In agreement with previous studies [Ward, W. H., Holdgate, G. A., Rowsell, S., McLean, E. G., Pauptit, R. A., Clayton, E., Nichols, W. W., Colls, J. G., Minshull, C. A., Jude, D. A., Mistry, A., Timms, D., Camble, R., Hales, N. J., Britton, C. J., and Taylor, I. W. (1999) Biochemistry 38, 12514-12525], we report here that triclosan is a slow, reversible, tight binding inhibitor of the FabI from Escherichia coli. Triclosan binds preferentially to the E.NAD(+) form of the wild-type enzyme with a K(1) value of 23 pM. In agreement with genetic selection experiments [McMurry, L. M., Oethinger, M., and Levy, S. B. (1998) Nature 394, 531-532], the affinity of triclosan for the FabI mutants G93V, M159T, and F203L is substantially reduced, binding preferentially to the E.NAD(+) forms of G93V, M159T, and F203L with K(1) values of 0.2 microM, 4 nM, and 0.9 nM, respectively. Triclosan binding to the E.NADH form of F203L can also be detected and is defined by a K(2) value of 51 nM. We have also characterized the Y156F and A197M mutants to compare and contrast the binding of triclosan to InhA, the homologous enoyl reductase from Mycobacterium tuberculosis. As observed for InhA, Y156F FabI has a decreased affinity for triclosan and the inhibitor binds to both E.NAD(+) and E.NADH forms of the enzyme with K(1) and K(2) values of 3 and 30 nM, respectively. The replacement of A197 with Met has no impact on triclosan affinity, indicating that differences in the sequence of the conserved active site loop cannot explain the 10000-fold difference in affinities of FabI and InhA for triclosan.

  19. Quench-condensing superconducting thin films using the Fab on a Chip approach (United States)

    Han, Han; Imboden, Matthias; Del Corro, Pablo; Stark, Thomas; Lally, Richard; Pardo, Flavio; Bolle, Cristian; Bishop, David

    Micro-electromechanical systems (MEMS) being manufactured in a macroscopic fab inspires the idea of getting the process further down to fabricate even smaller structures, namely nano-structures, using MEMS. The Fab on a Chip concept was proposed based on such ideas. By implementing the final-step, additive fabrication approach, manufacturing, characterization and experiments of nano-structures are integrated in-situ. Due to the miniature size of MEMS, the thickness precision is significantly improved while the power consumption is significantly depressed, making the quench-condensation of very thin films well controlled and easily achievable. Among various types of nano-structures, quench-condensed superconducting thin films are of great interest for physicists. Here we present such experiments done on superconducting thin films quench-condensed using the Fab on a Chip. We show that we are able to fabricate very thin films with its thickness precisely controlled, and the base temperature kept under ~3K during the process. The resistivity data demonstrates the high purity and uniformity of the film, as well as the annealing effect when cycling to higher temperatures. Based on the tremendous results obtained from the superconducting thin films, more complex nano-circuits can be fabricated and investigated using the Fab on a Chip, enabling a new approach for novel condensed matter physics experiments. This research is funded by the NSF through their CMMI division. This research is funded by the NSF through their CMMI division.

  20. Integrated MEMS mass sensor and atom source for a ``Fab on a Chip'' (United States)

    Han, Han; Imboden, Matthias; Stark, Thomas; Bishop, David


    ``Fab on a Chip'' is a new concept suggesting that the semiconductor fabrication facility can be integrated into a single silicon chip for nano-manufacturing. Such a chip contains various MEMS devices which can work together, operating in a similar way as a conventional fab does, to fabricate nano-structures. Here we present two crucial ``Fab on a chip'' components: the MEMS mass sensor and atomic evaporation source. The mass sensor is essentially a parallel plate capacitor with one suspended plate. When incident atoms deposit on the suspended plate, the mass change of the plate can be measured by detecting the resonant frequency shift. Using the mass sensor, a mass resolution of 3 fg is achieved. The MEMS evaporation source consists of a polysilicon plate suspended by two electrical leads with constrictions. By resistively heating the plate, this device works as a tunable atom flux source. By arranging many of these devices into an array, one can build a multi-element atom evaporator. The mass sensor and atom source are integrated so that the mass sensor is used to monitor and characterize the atomic flux. A material source and a sensor to monitor the fabrication are two integral components for our ``Fab on a Chip.''

  1. 75 FR 9438 - Samsung Austin Semiconductor, LLC, DRAM Fab 1, a Subsidiary of Samsung Electronics Corporation... (United States)


    ... Employment and Training Administration Samsung Austin Semiconductor, LLC, DRAM Fab 1, a Subsidiary of Samsung..., applicable to workers of Samsung Austin Semiconductor, LLC, a subsidiary of Samsung Electronics Corporation... Systems, Inc. were employed on-site at the Austin, Texas location of Samsung Austin Semiconductor, LLC,...

  2. Fab glycosylation of immunoglobulin G does not associate with improvement of rheumatoid arthritis during pregnancy

    NARCIS (Netherlands)

    A. Bondt (Albert); M. Wuhrer (Manfred); T.M. Kuijper (Martijn); J.M.W. Hazes (Mieke); R.J.E.M. Dolhain (Radboud)


    textabstractBackground: Changes in immunoglobulin G (IgG) constant domain (Fc) glycosylation are associated with changes in rheumatoid arthritis (RA) disease activity in response to pregnancy. Here, we sought to determine whether the same holds true for variable domain (Fab) glycosylation. Methods:

  3. Part I, FAB evaluation & application trials AFUE measurements: Part II, Integrated heating system (IHS) development

    Energy Technology Data Exchange (ETDEWEB)

    Leigh, R.W. [Brookhaven National Lab., Upton, NY (United States); Fisher, L. [BNL Consultant, Colrain, MA (United States)


    An oil burner/boiler efficiency test stand has been set up in the BNL oil heat laboratory which can measure the Annual Fuel Utilization Efficiency (AFUE) of burner/boiler combinations in accordance with ASHRAE and DOE standards. Measurements include both steady state efficiencies and heat-up and cool-down characteristics so that cycling effects can be included in an estimate of seasonal average performance. In addition to AFUE measurements, the direct conversion of fuel energy content to enthalpy increase in the boiler water is monitored. The system is largely automated, with most control functions under computer control and data taken electronically and permanently recorded on disks for future reference. To date, a retention-head burner and a fan atomized burner (FAB) have been tested in a steel boiler, the latter operating at two different fuel flow rates. The results are presented below, and verify that the very tight construction of the FAB`s fan results in a significant decrease in off-cycle sensible heat losses. Tests were also performed on a center-flue water heater fired with a conventional retention-head burner and with an FAB. The tests conformed to DOE standard procedures for hot water heaters, and the results are discussed below.

  4. Novel Schiff-base-derived FabH inhibitors with dioxygenated rings as antibiotic agents. (United States)

    Zhou, Yang; Du, Qian-Ru; Sun, Jian; Li, Jing-Ran; Fang, Fei; Li, Dong-Dong; Qian, Yong; Gong, Hai-Bin; Zhao, Jing; Zhu, Hai-Liang


    Fatty acid biosynthesis plays a vital role in bacterial survival and several key enzymes involved in this biosynthetic pathway have been identified as attractive targets for the development of new antibacterial agents. Of these promising targets, β-ketoacyl-acyl carrier protein (ACP) synthase III (FabH) is the most attractive target that could trigger the initiation of fatty acid biosynthesis and is highly conserved among Gram-positive and -negative bacteria. Designing small molecules with FabH inhibitory activity displays great significance for developing antibiotic agents, which should be highly selective, nontoxic and broad-spectrum. In this manuscript, a series of novel Schiff base compounds were designed and synthesized, and their biological activities were evaluated as potential inhibitors. Among these 21 new compounds, (E)-N-((3,4-dihydro-2H-benzo[b][1,4]dioxepin-7-yl)methylene)hexadecan-1-amine (10) showed the most potent antibacterial activity with a MIC value of 3.89-7.81 μM(-1) against the tested bacterial strains and exhibited the most potent E. coli FabH inhibitory activity with an IC(50) value of 1.6 μM. Docking simulation was performed to position compound 10 into the E. coli FabH active site to determine the probable binding conformation.

  5. Cyclization strategies of meditopes: affinity and diffraction studies of meditope-Fab complexes. (United States)

    Bzymek, Krzysztof P; Ma, Yuelong; Avery, Kendra A; Horne, David A; Williams, John C


    Recently, a unique binding site for a cyclic 12-residue peptide was discovered within a cavity formed by the light and heavy chains of the cetuximab Fab domain. In order to better understand the interactions that drive this unique complex, a number of variants including the residues within the meditope peptide and the antibody, as well as the cyclization region of the meditope peptide, were created. Here, multiple crystal structures of meditope peptides incorporating different cyclization strategies bound to the central cavity of the cetuximab Fab domain are presented. The affinity of each cyclic derivative for the Fab was determined by surface plasmon resonance and correlated to structural differences. Overall, it was observed that the disulfide bond used to cyclize the peptide favorably packs against a hydrophobic `pocket' and that amidation and acetylation of the original disulfide meditope increased the overall affinity ∼2.3-fold. Conversely, replacing the terminal cysteines with serines and thus creating a linear peptide reduced the affinity over 50-fold, with much of this difference being reflected in a decrease in the on-rate. Other cyclization methods, including the formation of a lactam, reduced the affinity but not to the extent of the linear peptide. Collectively, the structural and kinetic data presented here indicate that small perturbations introduced by different cyclization strategies can significantly affect the affinity of the meditope-Fab complex.

  6. In vitro Fab display: a cell-free system for IgG discovery. (United States)

    Stafford, Ryan L; Matsumoto, Marissa L; Yin, Gang; Cai, Qi; Fung, Juan Jose; Stephenson, Heather; Gill, Avinash; You, Monica; Lin, Shwu-Hwa; Wang, Willie D; Masikat, Mary Rose; Li, Xiaofan; Penta, Kalyani; Steiner, Alex R; Baliga, Ramesh; Murray, Christopher J; Thanos, Christopher D; Hallam, Trevor J; Sato, Aaron K


    Selection technologies such as ribosome display enable the rapid discovery of novel antibody fragments entirely in vitro. It has been assumed that the open nature of the cell-free reactions used in these technologies limits selections to single-chain protein fragments. We present a simple approach for the selection of multi-chain proteins, such as antibody Fab fragments, using ribosome display. Specifically, we show that a two-chain trastuzumab (Herceptin) Fab domain can be displayed in a format which tethers either the heavy or light chain to the ribosome while retaining functional antigen binding. Then, we constructed synthetic Fab HC and LC libraries and performed test selections against carcinoembryonic antigen (CEA) and vascular endothelial growth factor (VEGF). The Fab selection output was reformatted into full-length immunoglobulin Gs (IgGs) and directly expressed at high levels in an optimized cell-free system for immediate screening, purification and characterization. Several novel IgGs were identified using this cell-free platform that bind to purified CEA, CEA positive cells and VEGF.

  7. Orienterende Fast Atom Bombardment (FAB) experimenten met de VG-70-SQ massaspectrometer

    NARCIS (Netherlands)

    Hove GJ ten; Boer AC den; Burgers PC; Jong APJM de


    Eerste orienterende metingen met fast atom bombardment (FAB) ionisatietechniek zijn uitgevoerd. De techniek werd toegepast bij de analyse van korte-keten polypeptiden (n=2-5), cyclosporine, NADP en microperoxidase. Onderzocht werd de invloed van de aard van de matrix (glycerol, thioglycerol) op

  8. Crystal Structure of the Fab Fragment of an Anti-factor IX Antibody 10C12

    Institute of Scientific and Technical Information of China (English)

    SHI Xiao-Li; ZENG Tu; HUANG Ming-Dong


    10C12 is an anticoagulant antibody identified from a phage display single-chain Fv human antibody library. It can be directed at the calcium-stabilized Gla domain of Factor-IX, an important coagulation factor in intrinsic pathway of blood coagulation cascade, and interfere with membrane anchoring of Factor IX, thus inhibiting blood coagulation function. 10C12 has been demonstrated as an effective anti-coagulant in attenuating thrombosis in several different animal models. Here, we report the crystal structure of the Fab fragment of 10C12. The crystal contains two Fab molecules in the asymmetric unit with identical conformation, forming a lattice with large cavities. In addition, comparison of this free Fab with the antigen-bound structure of 10C12 shows no change in CDR conformations and the relative disposition of the variable subunits of H and L chains, suggesting the rigid conformation of this 10C12 Fab and a lock-and-key mechanism of antibody-antigen recognition for 10C 12.

  9. Impact of Fab Lab Tulsa on Student Self-Efficacy toward STEM Education (United States)

    Dubriwny, Nicholas; Pritchett, Nathan; Hardesty, Michelle; Hellman, Chan M.


    Student self-confidence is important to any attempt to increase interest and achievement in Science, Technology, Engineering, and Math (STEM) education. This study presents a longitudinal examination of Fab Lab Tulsa's impact on attitude and self-efficacy toward STEM education among middle-school aged students. Paired samples t-test showed a…

  10. Various Energy-Saving Approaches to a TFT-LCD Panel Fab

    Directory of Open Access Journals (Sweden)

    Cheng-Kuang Chang


    Full Text Available This study employs the developed simulation software for the energy use of the high-tech fabrication plant (hereafter referred as a fab to examine six energy-saving approaches for the make-up air unit (MAU of a TFT-LCD (thin-film transistor liquid-crystal display fab. The studied approaches include: (1 Approach 1: adjust the set point of dry bulb temperature and relative humidity in the cleanroom; (2 Approach 2: lower the flow rate of supply air volume in the MAU; (3 Approach 3: use a draw-through type instead of push through type MAU; (4 Approach 4: combine the two stage cooling coils in MAU to a single stage coil; (5 Approach 5: reduce the original MAU exit temperature from 16.5 °C to 14.5 °C; and (6 Approach 6: avoid an excessive increase in pressure drop over the filter by replacing the HEPA filter more frequently. The simulated results are further compared to the measured data of the studied TFT-LCD fab in Taiwan. The simulated results showed that Approach 1 exhibits more significant influence on annual power consumption than the other approaches. The advantage/disadvantage of each approach is elaborated. The impact of the six approaches on the annual power consumption of the fab is also discussed.

  11. Bathydorus laniger and Docosaccus maculatus (Lyssacinosida; Hexactinellida): Two new species of glass sponge from the abyssal eastern North Pacific Ocean. (United States)

    Kahn, Amanda S; Geller, Jonathan B; Reiswig, Henry M; Smith, Kenneth L


    Two new species of glass sponge were discovered from the abyssal plain 200 km west of the coast of California (Station M). The sponges have similar gross morphology--an unusual plate-like form with basalia stilling the body above soft abyssal sediments. Bathydorus laniger sp. n. differs from its congeners by the presence of dermal and atrial stauractins; it is also supported by smooth hypodermal pentactins and hypoatrial hexactins. Microscleres include oxyhexasters and oxyhemihexasters. Docosaccus maculatus sp. n. contains large hexactins (>1 cm), characteristic of the genus. Megascleres include dermal hexactins, atrial pentactins, and choanosomal hexactins and diactins. Microscleres include oxytipped hemihexasters and floricomes. Several features serve to differentiate this species from its only known congener.


    Directory of Open Access Journals (Sweden)

    Blondine Ch. P.


    Full Text Available A study using Vectobac 12 AS ( Bt H-14 and Bacillus thuringiensis H-14 local strain was conducted at laboratory Vector and Reservoir Control Research Unit and breeding ponds of Anopheles maculatus in Kokap regency and Kulon Progo district. The objectives of this study are : (1. to detect the efficacy of B. thuringiensis H-14 local strain toward An. maculatus larvae at the laboratory. (2. to measure the effectiveness of B. thuringiensis H-14 local strain dosages 1 x LC95. 5 x LC95 and 10 x LC95 toward An. maculatus at the field. The efficacy test of B. thuringiensis H-14 local strain toward An. maculatus based on to the method proposed by WHO in order to determine the LC50 and LC90 which is computed using the probit analysis at the laboratory. The methods used R thuringiensis H-14 local strain dosages of 2.145 ppm (1 x LC95, 10.724 ppm (5 x LC95 and 21.448 ppm (10 x LC95 respectively were applied 8 ponds with the width of ponds ranging from 0.08 to 0.45 m2, 0.29 to 0.64 m2 and from 0.08-0.79 m2.The results showed, the dosages after 24 hours were 10.22 ppm (LC50, 27.11 ppm (LC90 and 35.75 ppm (LC95. After 48 hours the dosages were needed 7.74 ppm (LC50, 17.06 ppm (LC90 and 21.34 ppm (LC95, The effectiveness o/R thuringiensis H-14 local strain dosages of 2.145ppm (1 x LC95 toward An. maculatus larvae until 50 % survive the same time (7.35 days as B. thuringiensis H-14 (8.14 days dosages of 10.724 ppm (5 x LC95. B. thuringiensis H-14 local strain dosages of 21.448 ppm (10 x LC95 toward An. maculatus larvae until 50 % survive longer time (16.21 days than B. thuringiensis H-14 local strain 1 x LC95 and 5 x LC95 The B. thuringiensis H-14 local strain is effective for controlling mosquitoes larvae

  13. Construction and selection of the natural immune Fab antibody phage display library from patients with colorectal cancer

    Institute of Scientific and Technical Information of China (English)

    Bao-Ping Wu; Bing Xiao; Tian-Mo Wan; Ya-Li Zhang; Zhen-Shu Zhang; Dian-Yuan Zhou; Zhuo-Sheng Lai; Chun-Fang Gao


    AIM: To construct the natural immune Fab antibody phage display libraries of colorectal cancer and to select antibodies related with colorectal cancer. METHODS: Extract total RNA from tissue of local cancer metastasis lymph nodes of patients with colorectal cancer.RT-PCR was used to amplify the heavy chain Fd and light chain к and the amplification products were inserted successively into the vector pComb3 to construct the human libraries of Fab antibodies. They were then panned by phage display technology. By means of Dot immunoblotting and ELISA, the libraries were identified and the Fab phage antibodies binding with antigens of colorectal cancer were selected. RESULTS: The amplified fragments of Fd and к gained by RT-PCR were about 650bp. Fd and к PCR products were subsequently inserted into the vector pComb3, resulting in a recombination rate of 40% and the volume of Fab phage display library reached 1.48 x 106. The libraries were enriched about 120-fold by 3 cycles of adsorption-elution- multiplication (panning). Dot immunoblotting showed Fab expressions on the phage libraries and ELISA showed 5clones of Fab phage antibodies which had binding activities with antigens of colorectal cancer. CONCLUSION: The natural immune Fab antibody phage display libraries of colorectal cancer were constructed. They could be used to select the relative antibodies of colorectal cancer.

  14. Effectiveness of Alpha-toxin Fab Monoclonal Antibody Therapy in Limiting the Pathology of Staphylococcus aureus Keratitis. (United States)

    Caballero, A; Foletti, D; Bierdeman, M; Tang, A; Arana, A; Hasa-Moreno, A; Sangalang, E; O'Callaghan, R J


    Abstract Purpose: To investigate the effectiveness of a high-affinity human monoclonal antibody Fab fragment to Staphylococcus aureus alpha-toxin (LTM14 Fab) as therapy for S. aureus keratitis. Methods: A single topical drop of the LTM14 Fab antibody to alpha-toxin alone, or in 0.006% benzalkonium chloride (BAK), was applied every 30 min to S. aureus-infected rabbit corneas from 9 to 14 hours post-infection. Erosions and pathology were measured at 15 h post-infection. Results: LTM14 Fab with BAK limited corneal erosions better than LTM14 Fab alone (p = 0.036), and both limited erosions compared to untreated eyes (p ≤ 0.0001). Overall pathology was similar in all groups (p ≥ 0.070), but iritis and chemosis were reduced by treatment (p ≤ 0.036). Conclusions: The high-affinity human monoclonal Fab fragment antibody (LTM14 Fab) to S. aureus alpha-toxin was effective in reducing corneal damage during S. aureus keratitis.

  15. The tumor-inhibitory effectiveness of a novel anti-Trop2 Fab conjugate in pancreatic cancer. (United States)

    Mao, Yuan; Wang, Xiaoying; Zheng, Feng; Wang, Changjun; Tang, Qi; Tang, Xiaojun; Xu, Ning; Zhang, Huiling; Zhang, Dawei; Xiong, Lin; Liang, Jie; Zhu, Jin


    Human trophoblastic cell surface antigen 2 (Trop2) has been reported to act oncogenically. In this study, one-step quantitative real-time polymerase chain reaction (qPCR) test and immunohistochemistry (IHC) analysis with were employed to evaluate the relationship between Trop2 expression and the clinicopathological features of patients with PC. Then a novel anti-Trop2 Fab antibody was conjugated with Doxorubicin (DOX) to form Trop2Fab-DOX, an antibody-drug conjugate. This Trop2Fab-DOX conjugate was characterized by cell ELISA and immunofluorescence assay. MTT and wound healing analyses were used to evaluate the inhibitory effect of Trop2Fab-DOX on PC cell growth in vitro, while xenograft nude mice model was established to examine the tumor-inhibitory effects of PC in vivo. High Trop2 expression was observed in PC tissues and Trop2 expression was associated with several malignant attributes of PC patients, including overall survival. Trop2Fab-DOX can bind to the Trop2-expressing PC cells and provide an improved releasing type of DOX. In addition, Trop2Fab-DOX inhibited the proliferation and suppressed the migration of PC cells in a dose-dependent manner in vitro, while inhibited the growth of PC xenografts in vivo. Trop2 is a specific marker for PC, and a novel Trop2Fab-DOX ADC has a potent antitumor activity.

  16. Variant vicilins from a resistant Vigna unguiculata lineage (IT81D-1053) accumulate inside Callosobruchus maculatus larval midgut epithelium. (United States)

    Oliveira, Gabriel B; Kunz, Daniele; Peres, Tanara V; Leal, Rodrigo B; Uchôa, Adriana F; Samuels, Richard I; Macedo, Maria Lígia R; Carlini, Célia R; Ribeiro, Alberto F; Grangeiro, Thalles B; Terra, Walter R; Xavier-Filho, José; Silva, Carlos P


    It has been demonstrated that variant vicilins are the main resistance factor of cowpea seeds (Vigna unguiculata) against attack by the cowpea beetle Callosobruchus maculatus. There is evidence that the toxic properties of these storage proteins may be related to their interaction with glycoproteins and other microvillar membrane constituents along the digestive tract of the larvae. New findings have shown that following interaction with the microvilli, the vicilins are absorbed across the intestinal epithelium and thus reach the internal environment of the larvae. In the present paper we studied the insecticidal activity of the variant vicilins purified from a resistant cowpea variety (IT81D-1053). Bioassays showed that the seeds of this genotype affected larval growth, causing developmental retardation and 100% mortality. By feeding C. maculatus larvae on susceptible and IT81D-1053 derived vicilins (FITC labelled or unlabelled), followed by fluorescence and immunogold cytolocalization, we were able to demonstrate that both susceptible and variant forms are internalized in the midgut cells and migrate inside vesicular structures from the apex to the basal portion of the enterocytes. However, when larvae were fed with the labelled vicilins for 24h and then returned to a control diet, the concentration of the variant form remained relatively high, suggesting that variant vicilins are not removed from the cells at the same rate as the non-variant vicilins. We suggest that the toxic effects of variant vicilins on midgut cells involve the binding of these proteins to the cell surface followed by internalization and interference with the normal physiology of the enterocytes, thereby affecting larval development in vivo.

  17. Comparative toxicity and micronuclei formation in Tribolium castaneum, Callosobruchus maculatus and Sitophilus oryzae exposed to high doses of gamma radiation. (United States)

    Ahmadi, Mehrdad; Mozdarani, Hossein; Abd-Alla, Adly M M


    The effects of gamma radiation on mortality and micronucleus formation in Tribolium castaneum Herbst, Callosobruchus maculatus (F.) and Sitophilus oryzae (L.) genital cells were evaluated. Two groups of healthy and active adult insects 1-3 and 8-10 days old were irradiated with various doses (50-200 Gy) gamma ray. Seven days post-irradiation; mortality rates and micronucleus formation were assessed in genital cells of the irradiated insects. The results show that with increasing gamma doses, the mortality rate of each species increased and T. castaneum and S. oryzae showed the low and high sensitivity respectively. It was shown that the micronucleus appearance in the tested insects had correlation with amount and intensity of radiation doses. Moreover our results indicate different levels in the genotoxicity of gamma radiation among the insects' genital cells under study. The frequency of micronuclei in genital cells of 1-3 days old insects exposed to 50 and 200 Gy were 12.6 and 38.8 Mn/1000 cells in T. castaneum, 20.8 and 46.8 Mn/1000 cells in C. maculatus and 16.8 and 57.2 Mn/1000 cells in S. oryzae respectively. A high sensitivity of the genital cells to irradiation exposure was seen in S. oryzae correlated with its high mortality rate compared with the other two species. These results might be indicative of inflicting chromosomal damage expressed as micronucleus in high mortality rates observed in the pest population; an indication of genotoxic effects of radiation on the studied species.

  18. Docking and molecular dynamics studies on triclosan derivatives binding to FabI. (United States)

    Yang, Xuyun; Lu, Junrui; Ying, Ming; Mu, Jiangbei; Li, Peichun; Liu, Yue


    FabI, enoyl-ACP reductase (ENR), is the rate-limiting enzyme in the last step for fatty acids biosynthesis in many bacteria. Triclosan (TCL) is a commercial bactericide, and as a FabI inhibitor, it can depress the substrate (trans-2-enoyl-ACP) binding with FabI to hinder the fatty acid synthesis. The structure-activity relationship between TCL derivatives and FabI protein has already been acknowledged, however, their combination at the molecular level has never been investigated. This paper uses the computer-aided approaches, such as molecular docking, molecular dynamics simulation, and binding free energy calculation based on the molecular mechanics/Poisson-Bolzmann surface area (MM/PBSA) method to illustrate the interaction rules of TCL derivatives with FabI and guide the development of new derivatives. The consistent data of the experiment and corresponding activity demonstrates that electron-withdrawing groups on side chain are better than electron-donating groups. 2-Hydroxyl group on A ring, promoting the formation of hydrogen bond, is vital for bactericidal effect; and the substituents at 4-position of A ring, 2'-position and 4'-position of B ring benefit antibacterial activity due to forming a hydrogen bond or stabilizing the conformation of active pocket residues of receptor. While the substituents at 3'-position and 5'-position of B ring destroy the π-π stacking interaction of A ring and NAD(+) which depresses the antibacterial activity. This study provides a new sight for designing novel TCL derivatives with superior antibacterial activity.

  19. Construction, Expression and in vitro Biological Effects of Idiotype Ig Fab Fragment of B-Chronic Lymphocytic Leukemia

    Institute of Scientific and Technical Information of China (English)

    Feng WANG; Ping LEI; Ping HU; Lijuan ZHU; Huifeng ZHU; Yue ZHANG; Jing YANG; Guanxin SHEN


    Summary: The purpose of this study was to construct expression vectors of idiotype (Id) Smlg in patients with B-chronic lymphocytic leukemia and to express them in E.coli to obtain recombinant Id,and to investigate the effect of the protein on the proliferation and secretion of IL-2 and IFN-γ of stimulated peripheral blood mononuclear cells (PBMC) in vitro. Light chain gene and Fd fragment of heavy chain gene were inserted into fd-tet-DOG2 vector to construct fd-tet-DOG2-Fab. Fab gene was further cloned into expression vector pHEN2 to construct the soluble expression vector pHEN2-Fab. After induction by IPTG, Fab protein was purified by Ni-NTA-chromatography. MTT was used to determine the effects of purified protein on the proliferation of stimulated PBMC in vitro and the concentrations of IL-2 and IFN-γ in the culture supernatants were detected by ELISA. The results showed that recombinant pHEN2-Fab expression vector was constructed successfully. Fab protein was expressed in positive clone after induced by 1PTG and two specific bands at 24-25 kD position were observed by SDS-PAGE electrophoresis. Proliferation of PBMC could be induced by purified Fab and the concentrations of IL-2 and IFN-γ, in culture supernatants were increased significantly after induction. It was suggested that the expression vector of SmIg Fab fragment was constructed successfully, and expressed and secreted from E. Coli. The Fab protein could induce proliferation of PBMC and promote secretion of IL-2 and IFN-γ.

  20. FabSim: facilitating computational research through automation on large-scale and distributed e-infrastructures

    CERN Document Server

    Groen, Derek; Suter, James; Hetherington, James; Zasada, Stefan; Coveney, Peter


    We present FabSim, a toolkit developed to simplify a range of computational tasks for researchers in diverse disciplines. FabSim is flexible, adaptable, and allows users to perform a wide range of tasks with ease. It also provides a systematic way to automate the use of resourcess, including HPC and distributed resources, and to make tasks easier to repeat by recording contextual information. To demonstrate this, we present three use cases where FabSim has enhanced our research productivity. These include simulating cerebrovascular bloodflow, modelling clay-polymer nanocomposites across multiple scales, and calculating ligand-protein binding affinities.

  1. Native MS and ECD Characterization of a Fab-Antigen Complex May Facilitate Crystallization for X-ray Diffraction (United States)

    Zhang, Ying; Cui, Weidong; Wecksler, Aaron T.; Zhang, Hao; Molina, Patricia; Deperalta, Galahad; Gross, Michael L.


    Native mass spectrometry (MS) and top-down electron-capture dissociation (ECD) combine as a powerful approach for characterizing large proteins and protein assemblies. Here, we report their use to study an antibody Fab (Fab-1)-VEGF complex in its near-native state. Native ESI with analysis by FTICR mass spectrometry confirms that VEGF is a dimer in solution and that its complex with Fab-1 has a binding stoichiometry of 2:2. Applying combinations of collisionally activated dissociation (CAD), ECD, and infrared multiphoton dissociation (IRMPD) allows identification of flexible regions of the complex, potentially serving as a guide for crystallization and X-ray diffraction analysis.

  2. A new type of pseudothrombocytopenia: EDTA-mediated agglutination of platelets bearing Fab fragments of a chimaeric antibody. (United States)

    Christopoulos, C G; Machin, S J


    In vitro agglutination of platelets leading to low automated platelet counts was observed in EDTA-anticoagulated blood from human volunteers receiving infusions of Fab fragments of a chimaeric monoclonal antibody to platelet glycoprotein IIb-IIIa. This pseudothrombocytopenia depended on the presence of chimaeric Fab on the platelet surface and was not seen when sodium citrate was used as anticoagulent. Preliminary evidence suggests that this phenomenon might be mediated by immunoglobulin G reactive with the human component of the chimaeric Fab. It is important to exclude pseudothrombocytopenia when low automated platelet counts are reported in association with the administration of chimaeric anti-platelet antibodies.

  3. 双功能螯合剂BAT与鼠IgG1 Fab'片段的点特异性交联方法的研究%Site-Specific Conjugation of Bifunctional Chelator BAT to Mouse IgG1 Fab' Fragment

    Institute of Scientific and Technical Information of China (English)

    李君; 王学浩; 成峰; 王晓明; 陈兆雷; 沈喜平


    目的报道一种用双功能螯合剂BAT与鼠单克隆抗体Fab'片段行点特异性交联的新方法.方法通过位于Fab'铰链区、远离抗原结合位点的活性巯基(-SH)直接与BAT相交联.用简单的两步法制备B43单抗的Fab'片段.首先单抗用胃蛋白酶处理,产生稳定的F(ab')2 片段,然后 F(ab')2经半胱氨酸还原后产生Fab'片段,最后Fab'片段直接与BAT交联.结果每个Fab'分子平均含1.8个-SH基团,74%的Fab'片段可与BAT生成交联物,平均每个Fab'分子携带1.28个BAT.经检测,F(ab')2、Fab' 及 Fab'-BAT 均保持着良好的免疫活性.结论该交联法简单、高效,为双功能螯合剂BAT与鼠单抗Fab'片段的交联提供了一个新途径.

  4. New species of Cerambycinae from the Neotropical Region, and nomen novum for Anelaphus maculatus Galileo, Martins, and Santos-Silva, 2014 (Elaphidiini). (United States)

    Galileo, Maria Helena M; Martins, Ubirajara R; Santos-Silva, Antonio


    Six new species and one new genus are described: Criodion spinosum sp. nov. (Cerambycini), from Bolivia; Eburodacrys wappesi sp. nov. and Eburodacrys skillmani sp. nov. (Eburiini), from Bolivia; Eupempelus rileyorum sp. nov. (Heteropsini) from Panama; Sphalloeme mexicana sp. nov. (Oemini), from Mexico; Wappesoeme camiri sp. nov., new genus (Oemini), from Bolivia. Wappesoeme, Eburodacrys wappesi, E. skillmani, Eupempelus rileyorum, and Criodion spinosum are included in previously published keys. Anelaphus erakyra nomen novum for A. maculatus Galileo et al., 2014 is established.

  5. Assessment of resistance to the attack of bean beetle Callosobruchus maculatus (Fabricius in chickpea genotypes on the basis of various parameters during storage

    Directory of Open Access Journals (Sweden)

    Muhammad Sarwar


    Full Text Available Chickpea (Cicer arietinum L., is an important pulse food. During storage this commodity is severely attacked bybean beetle Callosobruchus maculatus (Fabricius resulting losses in quantity and nutritional quality. Research studies onrelative resistance of 12 chickpea genotypes to the attack of C. maculatus during storage were carried out. The genotypesmost tolerant to bruchids comprised CH-52/02 and B-8/03, whereas, the most susceptible reactions were apparent inCH-86/02 and CC-117/00. The moderate pest incidence was observed in CH-28/02, CH-4/02, CH-32/02, CH-31/02, CH-9/02,CM-772/03, B-8/02 and CM-628/03 genotypes. The tolerant genotypes exhibited hard and wrinkled seed coat, dark browncolour and small size grain. These characteristics demonstrated a significant harmful effect to pest appearance and graindamage. The vulnerable genotypes had soft and smooth seed coat, white seed colour and bigger grain size that causedvulnerability to C. maculatus. Based on the present investigation, chickpea genotypes CH-52/02 and B-8/03 deserve specialconsideration and may be recommended for relatively longer storage to achieve the goal of long term and sustainable pestmanagement strategies.

  6. Effect of Callosobruchus maculatus Infestation on the Nutrient-antinutrient Composition, Phenolic Composition and Antioxidant Activities of Some Varieties of Cowpeas (Vigna unguiculata

    Directory of Open Access Journals (Sweden)

    Sule Ola Salawu


    Full Text Available Cowpea Vigna unguiculata L. Walp. is an important legume crops in most part of the world and is of health benefit in addition to the nutritional potential. However, this important nutrient and disease preventing crop, is prone to post harvest infestation damage, mostly by Callosobruchus maculatus . Therefore, the present study is to evaluate the antioxidant activities, nutrient composition, anti-nutrient content and phenolic composition of Callosobruchus maculatus infested and non-infested cowpeas commonly consumed in Nigeria. The result revealed that infestation brings about an increase in the evaluated anti-nutrients (phytate, oxalate and alkaloids and a marked reduction in protein, carbohydrate content, whereas a marked increase was recorded for the fibre content. The result of the antioxidant activities revealed a significant reduction for total phenol, total flavonoids and reducing power and a slight reduction was recorded for the inhibitory action against lipid oxidation, hydroxyl radical scavenging, 2, 2’-azinobis-3-ethylbenzothiazoline-6-sulfonic acid radical scavenging and 1, 1-diphenyly-2-picrylhydrazyl radical scavenging activities after infestation. The present investigations revealed that Callosobruchus maculatus infestation do not only affect the nutrient composition of the studied cowpeas by reducing the nutritional value, but would also have an adverse effect on their potential to prevent free radical mediated diseases.

  7. Campylobacter jejuni fatty acid synthase II: Structural and functional analysis of [beta]-hydroxyacyl-ACP dehydratase (FabZ)

    Energy Technology Data Exchange (ETDEWEB)

    Kirkpatrick, Andrew S.; Yokoyama, Takeshi; Choi, Kyoung-Jae; Yeo, Hye-Jeong; (Houston)


    Fatty acid biosynthesis is crucial for all living cells. In contrast to higher organisms, bacteria use a type II fatty acid synthase (FAS II) composed of a series of individual proteins, making FAS II enzymes excellent targets for antibiotics discovery. The {beta}-hydroxyacyl-ACP dehydratase (FabZ) catalyzes an essential step in the FAS II pathway. Here, we report the structure of Campylobacter jejuni FabZ (CjFabZ), showing a hexamer both in crystals and solution, with each protomer adopting the characteristic hot dog fold. Together with biochemical analysis of CjFabZ, we define the first functional FAS II enzyme from this pathogen, and provide a framework for investigation on roles of FAS II in C. jejuni virulence

  8. Crystallization of the Fab from a human monoclonal antibody against gp 41 of human immunodeficiency virus type I (United States)

    Casale, Elena; He, Xiao-Min; Snyder, Robert S.; Carter, Daniel C.; Wenisch, Elisabeth; Jungbauer, Alois; Tauer, Christa; Ruker, Florian; Righetti, Pier Giorgio


    A monoclonal IgG antibody directed against gp 41 from the human immunodeficiency virus (HIV-1) has been crystallized in both intact and Fab forms. Crystals of the intact antibody grow as tetragonal-like prisms too small for conventional X-ray analysis. However, the Fab portion of the antibody produces suitable platelike crystals which belong to the space group P2(1)2(1)2(1) with unit cell constants of a = 66.5 A, b = 74.3 A, and c = 105.3 A. There is one molecule of Fab in the asymmetric unit. The Fab crystals show diffraction to d-spacings less than 3.0 A.

  9. Necrobiont Coleoptera North-West Caucasus

    Directory of Open Access Journals (Sweden)

    Sergey Viktorovich Pushkin


    Full Text Available The complex necrobionts of the alpine girdle of Northwest Caucasus is described ecologically. The specific composition of groups is circumscribed. The legitimacies of creation necrobionts of the complex from a type of a landscape and taxonomic of an accessory of a corpse are detected. Studying of regional features ¬of formation necrobionts complexes has, both scientific, and practical -interest. Throughout 20 years we studied fauna and ecological features necrobiont Coleoptera mountain landscapes of Northwest Caucasus and -adjoining areas of Ciscaucasia.

  10. Immatures of Acanthocinini (Coleoptera, Cerambycidae, Lamiinae

    Directory of Open Access Journals (Sweden)

    Sônia A. Casari


    Full Text Available Immatures of Acanthocinini (Coleoptera, Cerambycidae, Lamiinae. Larva and pupa of Eutrypanus dorsalis (Germar, 1928, collected in trunks of Pinus elliottii Engelm., and Paratenthras martinsi Monné, 1998, collected in spathes of Scheelea phalerata (Mart. ex Spreng. Burret, are described and illustrated. Larva and pupa of Lophopoeum timbouvae Lameere, 1884, collected in Hymenaea corbaril L., Enterolobium contortisiliquum (Vell. Morong and Pterogyne nitens Tul., are redescribed and illustrated. A table with all described immatures of Lamiinae, and a comparison among the immatures of Acanthocinini are presented. Biological notes and new records are also included.

  11. Description of the immature stages of four species of Macrodactylini (Coleoptera: Melolonthidae: Melolonthinae). (United States)

    Neita-Moreno, J C; Morón, M A; Zuluaga-Correa, C A


    The third instar of the genera Ceraspis, Clavipalpus, Isonychus and Manopus (Melolonthidae: Melolonthinae: Macrodactylini) are described for the first time. Descriptions are based on the larvae of Ceraspis innotata (Blanchard), Clavipalpus ursinus Blanchard, Isonychus maculatus Waterhouse and Manopus biguttatus Laporte. The pupae of C. ursinus, I. maculatus and M. biguttatus are also described. A key to the larvae of nine genera of Macrodactylini and a list of the species with immature descriptions are provided.

  12. Improving Assessment of Work Related Mental Health Function Using the Work Disability Functional Assessment Battery (WD-FAB). (United States)

    Marfeo, Elizabeth E; Ni, Pengsheng; McDonough, Christine; Peterik, Kara; Marino, Molly; Meterko, Mark; Rasch, Elizabeth K; Chan, Leighton; Brandt, Diane; Jette, Alan M


    Purpose To improve the mental health component of the Work Disability Functional Assessment Battery (WD-FAB), developed for the US Social Security Administration's (SSA) disability determination process. Specifically our goal was to expand the WD-FAB scales of mood & emotions, resilience, social interactions, and behavioral control to improve the depth and breadth of the current scales and expand the content coverage to include aspects of cognition & communication function. Methods Data were collected from a random, stratified sample of 1695 claimants applying for the SSA work disability benefits, and a general population sample of 2025 working age adults. 169 new items were developed to replenish the WD-FAB scales and analyzed using factor analysis and item response theory (IRT) analysis to construct unidimensional scales. We conducted computer adaptive test (CAT) simulations to examine the psychometric properties of the WD-FAB. Results Analyses supported the inclusion of four mental health subdomains: Cognition & Communication (68 items), Self-Regulation (34 items), Resilience & Sociability (29 items) and Mood & Emotions (34 items). All scales yielded acceptable psychometric properties. Conclusions IRT methods were effective in expanding the WD-FAB to assess mental health function. The WD-FAB has the potential to enhance work disability assessment both within the context of the SSA disability programs as well as other clinical and vocational rehabilitation settings.

  13. A novel human Fab antibody for Trop2 inhibits breast cancer growth in vitro and in vivo. (United States)

    Lin, Hong; Zhang, Huiling; Wang, Jun; Lu, Meiping; Zheng, Feng; Wang, Changjun; Tang, Xiaojun; Xu, Ning; Chen, Renjie; Zhang, Dawei; Zhao, Ping; Zhu, Jin; Mao, Yuan; Feng, Zhenqing


    Human trophoblastic cell surface antigen 2 (Trop2) has been suggested as an oncogene, which is associated with the different types of tumors. In this study, a human Fab antibody against Trop2 extracellular domain was isolated from phage library by phage display technology, and characterized by ELISA, FACS, fluorescence staining and Western blotting analysis. MTT, apoptosis assay and wound healing assay were employed to evaluate the inhibitory effects of Trop2 Fab on breast cancer cell growth in vitro, while tumor-xenograft model was employed to evaluate the inhibitory effects on breast cancer growth in vivo. The results showed that Trop2 Fab inhibited the proliferation, induced the apoptosis and suspended the migration of MDA-MB-231 cells in a dose dependent manner. The expression caspase-3 was activated, and the expression of Bcl-2 was reduced while that of Bax was elevated in MDA-MB-231 cells by treating with Trop2 Fab. In addition, Trop2 Fab inhibited the growth of breast cancer xenografts and the expression of Bcl-2 was reduced while that of Bax was elevated in xenografts. Trop2 Fab, which was isolated successfully in this research, is a promising therapeutic agent for the treatment of Trop2 expressing breast cancer.


    Institute of Scientific and Technical Information of China (English)

    Li Li; Fei Qiao; Wu Qidi


    Based on the analysis of collective activities of ant colonies, the typical example of swarm intelligence, a new approach to construct swarm intelligence based multi-agent-system (SMAS) for dynamic real-time scheduling for semiconductor wafer fab is proposed. The relevant algorithm,pheromone-based dynamic real-time scheduling algorithm (PBDR), is given. MIMAC test bed data set mini-fab is used to compare PBDR with FIFO (first in first out), SRPT(shortest remaining processing time) and CR(critical ratio) under three different release rules, i.e. deterministic rule, Poisson rule and CONWIP (constant WIP). It is shown that PBDR is prior to FIFO, SRPT and CR with better performance of cycle time, throughput, and on-time delivery, especially for on-time delivery performance.

  15. The FermiFab Toolbox for Fermionic Many-Particle Quantum Systems

    CERN Document Server

    Mendl, Christian B


    This paper introduces the FermiFab toolbox for many-particle quantum systems. It is mainly concerned with the representation of (symbolic) fermionic wavefunctions and the calculation of corresponding reduced density matrices (RDMs). The toolbox transparently handles the inherent antisymmetrization of wavefunctions and incorporates the creation/annihilation formalism. Thus, it aims at providing a solid base for a broad audience to use fermionic wavefunctions with the same ease as matrices in Matlab, say. Leveraging symbolic computation, the toolbox can greatly simply tedious pen-and-paper calculations for concrete quantum mechanical systems, and serves as "sandbox" for theoretical hypothesis testing. FermiFab (including full source code) is freely available as a plugin for both Matlab and Mathematica.

  16. Development of tools to study personal weight control strategies: OxFAB taxonomy


    Hartmann‐Boyce, Jamie; Aveyard, Paul; Koshiaris, Constantinos; Jebb, Susan A


    Objective To describe the development of the Oxford Food and Activity Behaviors (OxFAB) taxonomy and questionnaire to explore the cognitive and behavioral strategies used by individuals during weight management attempts. Methods The taxonomy was constructed through a qualitative analysis of existing resources and a review of existing behavior change taxonomies and theories. The taxonomy was translated into a questionnaire to identify strategies used by individuals. Think‐aloud interviews were...

  17. Development of tools to study personal weight control strategies: OxFAB taxonomy.


    Hartmann-Boyce, J; Aveyard, P; Koshiaris, C; Jebb, SA


    To describe the development of the Oxford Food and Activity Behaviors (OxFAB) taxonomy and questionnaire to explore the cognitive and behavioral strategies used by individuals during weight management attempts.The taxonomy was constructed through a qualitative analysis of existing resources and a review of existing behavior change taxonomies and theories. The taxonomy was translated into a questionnaire to identify strategies used by individuals. Think-aloud interviews were conducted to test ...

  18. Fab the coming revolution on your desktop : from personal computers to personal fabrication

    CERN Document Server

    Gershenfeld, Neil


    What if you could someday put the manufacturing power of an automobile plant on your desktop? According to Neil Gershenfeld, the renowned MIT scientist and inventor, the next big thing is personal fabrication-the ability to design and produce your own products, in your own home, with a machine that combines consumer electronics and industrial tools. Personal fabricators are about to revolutionize the world just as personal computers did a generation ago, and Fab shows us how.

  19. A status analysis of current digital marketing: a case study of Kauneusstudio FAB


    Tran, Trong


    The subject of this thesis is a small company's current digital marketing status. This study was con-ducted in order for the owners of the beauty and hair salon Kauneusstudio FAB to improve their understanding of their customers’ behavior online and the significance of each digital channel they are using in the present marketing strategy. The goal of this study is to provide information for the company to recognize the strengths and the development points of the current digital marketing stra...

  20. Aerial image measurement technique for automated reticle defect disposition (ARDD) in wafer fabs (United States)

    Zibold, Axel M.; Schmid, Rainer M.; Stegemann, B.; Scheruebl, Thomas; Harnisch, Wolfgang; Kobiyama, Yuji


    The Aerial Image Measurement System (AIMS)* for 193 nm lithography emulation has been brought into operation successfully worldwide. A second generation system comprising 193 nm AIMS capability, mini-environment and SMIF, the AIMS fab 193 plus is currently introduced into the market. By adjustment of numerical aperture (NA), illumination type and partial illumination coherence to match the conditions in 193 nm steppers or scanners, it can emulate the exposure tool for any type of reticles like binary, OPC and PSM down to the 65 nm node. The system allows a rapid prediction of wafer printability of defects or defect repairs, and critical features, like dense patterns or contacts on the masks without the need to perform expensive image qualification consisting of test wafer exposures followed by SEM measurements. Therefore, AIMS is a mask quality verification standard for high-end photo masks and established in mask shops worldwide. The progress on the AIMS technology described in this paper will highlight that besides mask shops there will be a very beneficial use of the AIMS in the wafer fab and we propose an Automated Reticle Defect Disposition (ARDD) process. With smaller nodes, where design rules are 65 nm or less, it is expected that smaller defects on reticles will occur in increasing numbers in the wafer fab. These smaller mask defects will matter more and more and become a serious yield limiting factor. With increasing mask prices and increasing number of defects and severability on reticles it will become cost beneficial to perform defect disposition on the reticles in wafer production. Currently ongoing studies demonstrate AIMS benefits for wafer fab applications. An outlook will be given for extension of 193 nm aerial imaging down to the 45 nm node based on emulation of immersion scanners.

  1. Characterization of a recombinant humanized anti-cocaine monoclonal antibody and its Fab fragment. (United States)

    Kirley, Terence L; Norman, Andrew B


    Variations of post-translational modifications are important for stability and in vivo behavior of therapeutic antibodies. A recombinant humanized anti-cocaine monoclonal antibody (h2E2) was characterized for heterogeneity of N-linked glycosylation and disulfide bonds. In addition, charge heterogeneity, which is partially due to the presence or absence of C-terminal lysine on the heavy chains, was examined. For cocaine overdose therapy, Fab fragments may be therapeutic, and thus, a simplified method of generation, purification, and characterization of the Fab fragment generated by Endoproteinase Lys-C digestion was devised. Both the intact h2E2 antibody and purified Fab fragments were analyzed for their affinities for cocaine and 2 of its metabolites, benzoylecgonine and cocaethylene, by fluorescence quenching of intrinsic antibody tyrosine and tryptophan fluorescence resulting from binding of these drugs. Binding constants obtained from fluorescence quenching measurements are in agreement with recently published radioligand and ELISA binding assays. The dissociation constants determined for the h2E2 monoclonal and its Fab fragment are approximately 1, 5, and 20 nM for cocaethylene, cocaine, and benzoylecgonine, respectively. Tryptophan fluorescence quenching (emission at 330 nm) was measured after either excitation of tyrosine and tryptophan (280 nm) or selective excitation of tryptophan alone (295 nm). More accurate binding constants are obtained using tryptophan selective excitation at 295 nm, likely due to interfering absorption of cocaine and metabolites at 280 nm. These quenching results are consistent with multiple tryptophan and tyrosine residues in or near the predicted binding location of cocaine in a previously published 3-D model of this antibody's variable region.

  2. Fab(nimotuzumab)-HYNIC-99mTc: Antibody Fragmentation for Molecular Imaging Agents. (United States)

    Calzada, Victoria; García, María Fernanda; Alonso-Martínez, Luis Michel; Camachoc, Ximena; Goicochea, Enzo; Fernández, Marcelo; Castillo, Abmel Xiques; Díaz-Miqueli, Arlhee; Iznaga-Escobar, Normando; Montaña, René Leyva; Alonso, Omar; Gambini, Juan Pablo; Cabral, Pablo


    Finally, fast blood clearance nimotuzumab is a humanized monoclonal antibody that recognise, with high specific affinity, the epidermal growth factor receptor (EGF-R) which play an important role in the growth process associated with many solid tumors. In this work, the whole antibody was digested with papain in order to generate a Fab fragment, derivatized with NHS-HYNIC-Tfa and radiolabel with technetium-99m (99mTc) as a potential agent of molecular imaging of cancer. Both, whole and fragment radiolabels were in-vivo and in-vitro characterized. Radiolabeling conditions with Tricine as coligand and quality controls were assessed to confirm the integrity of the labeled fragment. Biodistribution and imaging studies in normal and spontaneous adenocarcinoma mice were performed at different times to determine the in-vivo characteristics of the radiolabel fragment. Tumor localization was visualized by conventional gamma camera imaging studies, and the results were compared with the whole antibody. Also, an immunoreactivity assay was carried out for both. The results showed clearly the integrity of the nimotuzumab fragment and the affinity by the receptor was verified. Fab(nimotuzumab)-HYNIC was obtained with high purity and a simple strategy of radiolabeling was performed. Finally, a fast blood clearance was observed in the biodistribution studies increasing the tumor uptake of Fab(nimotuzumab)- HYNIC-99mTc over time, with tumor/muscle ratios of 3.81 ± 0.50, 5.16 ± 1.97 and 6.32 ± 1.98 at 1 h, 4 h and 24 h post injection. Urinary excretion resulted in 32.89 ± 3.91 %ID eliminated at 24 h. Scintigraphy images showed uptake in the tumor and the activity in non-target organs was consistent with the biodistribution data at the same time points. Hence, these preliminary results showed important further characteristic of Fab(nimotuzumab)-HYNIC-99mTc as a molecular imaging agent of cancer.

  3. Functional capacity of immunoglobulin G preparations and the F(ab')2 split product.


    Steele, R W


    Five immunoglobulin G preparations, including one 5S F(ab')2 split product, were compared for activity against common bacterial, viral, and protozoan pathogens. Standard assays were used to quantitate antibodies to tetanus, diphtheria, cytomegalovirus, herpes simplex virus types 1 and 2, rubella virus, and Toxoplasma gondii. Opsonization and killing of bacteria were examined by chemiluminescence methods using Streptococcus pneumoniae types 5, 12F, and 14 and Staphylococcus aureus. Antibodies ...

  4. Family of Advanced Beyond Line-of-Sight Terminals (FAB-T) (United States)


    Conferencing (PNVC) function; the Telemetry , Tracking & Control for the Milstar and AEHF constellations, for Nuclear Command, Control, & Communications...Coexistences: 4 ​Footnotes: 1/ For FAB-T, access to privileged Tracking Telemetry and Control (TT&C) capabilities and resource controller capabilities is...DIACAP - DoD Information Assurance Certification & Accreditation Process DIRECT - Defense IEMATS Replacement Command and Control Terminal DMU - Dual

  5. [Digitoxin poisoning: reversing ventricular fibrillation with Fab fragments of anti-digoxin antibody]. (United States)

    Domart, Y; Bismuth, C; Schermann, J M; Abuaf, N; Pontal, P G; Baud, F; Bolo, A; Gailliot, M; Fournier, P E


    Purified Fab fragments of ovine anti-digoxin antibodies (Wellcome Foundation) were used to treat a patient who attempted suicide by absorbing 10 mg of digitoxin (serum concentration 265 micrograms/l). The poor prognosis, as assessed clinically and from serum potassium levels (7.5 mEq/l), seemed to warrant such a treatment. The weak (6.85%) cross-reactivity elicited in vitro between the anti-digoxin antibodies and digitoxin was compensated by increasing the doses, but improvement was observed with 3.6 g, i.e. about half the effective dosage initially considered. The criteria of effectiveness were clinical, electrocardiographic (reversal of the ventricular fibrillation), biochemical (simultaneous and opposite changes in extra- and intracellular potassium levels, suggesting that ATPase inhibition by digitalis is a reversible process) and toxicological: there was an increase in digitoxin serum levels suggesting displacement of the drug from tissue sites to plasma and other extracellular compartments where the Fab fragments are distributed, and Fab-bound digitoxin appeared fairly rapidly in the urine, which suggested shunting of the normal hepatic metabolic pathway.

  6. An Efficient and Economical Assay to Screen for Triclosan Binding to FabI. (United States)

    Demissie, Robel D; Kabre, Pauline; Tuntland, Micheal L; Fung, Leslie W-M


    Triclosan is an effective inhibitor for enoyl acyl carrier protein reductase (ENR) in fatty acid biosynthesis. Triclosan-resistant mutants of ENR have emerged. Thus, it is important to detect these triclosan-resistant mutations in ENR. Generally, enzyme activity assays on the mutants are used to determine the effect of triclosan on ENR activity. Since the substrates are linked to acyl carrier protein (ACP), the assays are challenging due to the need to prepare the ACP and link it to the substrates. Non-ACP-linked (coenzyme A [CoA]-linked) substrates can be used in some ENR, but not in all. Consequently, screening for triclosan-resistant mutants is also challenging. We have developed a simple thermal shift assay, which does not use ACP-linked substrates, to determine the binding ability of triclosan to the ENR active site, and thus it can be used for screening for triclosan-resistant mutants. Staphylococcus aureus FabI enzyme and its mutants were used to demonstrate the binding ability of triclosan with NADP(+) to FabI. The direct correlation between the binding ability and enzyme activity was demonstrated with Francisella tularensis FabI. This method may also be applied to select effective triclosan analogues that inhibit ENR activity.

  7. Productivity improvement through automated operation of reticle defect inspection tools in a wafer fab environment (United States)

    Holfeld, Christian; Wagner, Heiko; Tchikoulaeva, Anna; Loebeth, Steffen; Melzig, Stephan; Zhang, Yulin; Tanabe, Shinichi; Katoh, Takenori; Moriizumi, Koichi


    Traditionally, product development for reticle defect inspection mostly addressed operational requirements of the mask shops with highly individualized manufacturing. As a result, limited automation capability was available as compared to the standards in wafer production. Wafer fabs are guided by completely different conditions. Thousands of active reticles exist in a single fab requiring frequent re-inspections without interruption of wafer exposures. This requires high throughput of inspection tools, smart management of tool fleet, sophisticated scheduling and in-time execution of reticle inspections linked to the wafer manufacturing. The paper reports about the successful implementation of fully automated reticle defect inspection in a high-volume advanced logic fab. Automation scenarios - created based on existing SEMI standards - included inspection scheduling, reticle transport and inspection tool operation. A considerable productivity gain for the operation of Lasertec MATRICS X700 series inspection tools was obtained. Based on the learning throughout implementation, the requirements to the automation capability and tool operation as well as adjustments to working procedures are discussed.

  8. Fabricating quench condensed lead thin film circuits using MEMS Fab on a Chip technology (United States)

    Imboden, Matthias; Han, Han; Del Corro, Pablo; Pardo, Flavio; Bolle, Cristian; Bishop, David


    We have developed a MEMS Fab on a Chip consisting of micro-sources, mass sensors, heaters/thermometers, shutters and a dynamic stencil. The fab only occupies a volume of a few cubic millimeters and consumes milliwatts of power, and hence can be operated in a cryostat. Thin film patterns of arbitrary shapes using multiple materials can be manufactured, while strongly suppressing thermal annealing effects. We demonstrate deposition of quench condensed lead films with fractions of a monolayer thickness control. Furthermore, using low deposition rates it is estimated that the surface temperature of the target heats by only 1.7 K. We study the effects of growing quench condensed films with different evaporation rates to demonstrate thermal annealing effects which occur during deposition. We measure the minimum conduction thickness (insulator to metal transition) as well as the superconducting transition temperature as a function of film thickness in order to shed light on growth of amorphous films and the transition to nanocluster formations. The Fab on a Chip will allow us to build nanocircuits made of ultra-thin materials. Annealing and doping is controlled and measurements occur in situ, without exposing the fabricated circuits to thermal fluctuations or foreign contaminants. This enables new types of experiments based on quantum circuits which cannot be fabricated using standard lithography techniques.

  9. Precise construction of oligonucleotide-Fab fragment conjugate for homogeneous immunoassay using HaloTag technology. (United States)

    Päkkilä, Henna; Peltomaa, Riikka; Lamminmäki, Urpo; Soukka, Tero


    The use of oligonucleotide-protein conjugates enables the development of novel types of bioanalytical assays. However, convenient methods for producing covalent and stoichiometric oligonucleotide-protein conjugates are still rare. Here we demonstrate, for the first time, covalent conjugation of DNA oligonucleotide to Fab fragments with a 1:1 ratio using HaloTag self-labeling technology. The oligonucleotide coupling was carried out while the Fab was attached to protein G matrix, thereby enabling straightforward production of covalent conjugates. Furthermore, it allowed convenient purification of the product because the unreacted components were easily removed before the elution of the high-purity conjugate. The prepared conjugate was employed in a homogeneous immunoassay where prostate-specific antigen was used as a model analyte. Switchable lanthanide luminescence was used for detection, and the obtained limit of detection was 0.27 ng/ml. In the future, the developed method for covalent conjugation and successive purification in protein G column could also be applied for introducing other kinds of modifications to Fab fragments in a simple and site-specific manner.

  10. Bases biológicas para el cultivo del puye Galaxias maculatus (Jenyns, 1842: una revisión

    Directory of Open Access Journals (Sweden)

    Rolando Vega


    Full Text Available Galaxias maculatus (Jenyns, 1842 es un pez gourmet de importancia comercial cuyas pesquerías están sobre-explotadas, siendo esencial el estudio de su biología para desarrollar su tecnología de cultivo (galaxicultura, que se encuentra en etapa piloto. Los objetivos de éste trabajo son: 1 entregar una síntesis de la información de la literatura sobre la biología de la especie, y 2 identificar la carencia de conocimiento científico y puntos críticos para el desarrollo de su tecnología de cultivo comercial. G. maculatus es un pequeño pez carnívoro, con poblaciones eurihalinas diferenciadas en diadrómicas y dulceacuícolas. Las poblaciones diadrómicas desovan en los estuarios y la larva migra al mar, retornando a la edad de seis meses para metamorfosearse en adulto. Son escasos los estudios de su sistemática, poblaciones y estado larvario en aguas chilenas, pero los correspondientes a alimentación, reproducción y enfermedades son más numerosos. Las hembras de un año desovan, aproximadamente, 1.200 huevos adhesivos pero un número importante muere después del primer desove (40%. La especie tiene un crecimiento rápido (1,1% dia-1 y alto metabolismo, con un promedio de vida de dos años. Se puede cultivar en cautividad, desovar, incubar sus huevos y obtener larvas; los adultos comen starter pelletizado de salmón y crecen en estanques. El protozoo ciliado Ichtyophthirius multifilis (ich produce altas mortalidades, en ejemplares en cautiverio, que pueden ser controladas con baños de sal. Los problemas de investigación a resolver para una futura piscicultura comercial son: identificar y seleccionar las poblaciones adecuadas para cultivo, aumentar el número de huevos desovados por hembra, desarrollar alimentos para larvas y reproductores, y controlar las enfermedades ectoparasitarias. El punto crítico es la masificación de la producción mediante el mejoramiento de las técnicas de reproducción y larvicultura.

  11. The evolution of asymmetric genitalia in Coleoptera (United States)

    de Jong, Paulien; van Beek, Rick; Hoogenboom, Tamara; zu Schlochtern, Melanie Meijer


    The evolution of asymmetry in male genitalia is a pervasive and recurrent phenomenon across almost the entire animal kingdom. Although in some taxa the asymmetry may be a response to the evolution of one-sided, male-above copulation from a more ancestral female-above condition, in other taxa, such as Mammalia and Coleoptera, this explanation appears insufficient. We carried out an informal assessment of genital asymmetry across the Coleoptera and found that male genital asymmetry is present in 43% of all beetle families, and at all within-family taxonomic levels. In the most diverse group, Cucujiformia, however, genital asymmetry is comparatively rare. We also reconstructed the phylogeny of the leiodid tribe Cholevini, and mapped aspects of genital asymmetry on the tree, revealing that endophallus sclerites, endophallus, median lobe and parameres are, in a nested fashion, increasingly unlikely to have evolved asymmetry. We interpret these results in the light of cryptic female choice versus sexually antagonistic coevolution and advocate further ways in which the phenomenon may be better understood. This article is part of the themed issue ‘Provocative questions in left–right asymmetry’. PMID:27821530

  12. Crystal structure and substrate specificity of the [beta]-ketoacyl-acyl carrier protein synthase III (FabH) from Staphylococcus aureus

    Energy Technology Data Exchange (ETDEWEB)

    Qiu, Xiayang; Choudhry, Anthony E.; Janson, Cheryl A.; Grooms, Michael; Daines, Robert A.; Lonsdale, John T.; Khandekar, Sanjay S. (GSK)


    {beta}-Ketoacyl-ACP synthase III (FabH), an essential enzyme for bacterial viability, catalyzes the initiation of fatty acid elongation by condensing malonyl-ACP with acetyl-CoA. We have determined the crystal structure of FabH from Staphylococcus aureus, a Gram-positive human pathogen, to 2 {angstrom} resolution. Although the overall structure of S. aureus FabH is similar to that of Escherichia coli FabH, the primer binding pocket in S. aureus FabH is significantly larger than that present in E. coli FabH. The structural differences, which agree with kinetic parameters, provide explanation for the observed varying substrate specificity for E. coli and S. aureus FabH. The rank order of activity of S. aureus FabH with various acyl-CoA primers was as follows: isobutyryl- > hexanoyl- > butyryl- > isovaleryl- >> acetyl-CoA. The availability of crystal structure may aid in designing potent, selective inhibitors of S. aureus FabH.

  13. Purificação e caracterização do fragmento Fab anti-digoxina obtido pela técnica de phage display.



    A digoxina é um dos medicamentos indicados para o tratamento de falência cardíaca. Possui janela terapêutica estreita, sendo responsável por casos de intoxicação. O único antídoto disponível para a desintoxicação é o anticorpo policlonal DigiFab®, no formato Fab. O seu uso é eficaz, porém de custo elevado. Clones bacterianos produtores de fragmento Fab monoclonal anti-digoxina foram obtidos previamente pelo nosso grupo, pela técnica de phage display. Neste trabalho as variantes Fab dos 4 clo...

  14. Regulation of fatty acid biosynthesis by the global regulator CcpA and the local regulator FabT in Streptococcus mutans


    Faustoferri, R.C.; Hubbard, C.J.; Santiago, B.; Buckley, A.A.; Seifert, T.B.; Quivey, R.G.


    SMU.1745c, encoding a putative transcriptional regulator of the MarR family, maps to a location proximal to the fab gene cluster in Streptococcus mutans. Deletion of the SMU.1745c (fabTSm) coding region resulted in a membrane fatty acid composition comprised of longer-chained, unsaturated fatty acids (UFA), compared with the parent strain. Previous reports have indicated a role for FabT in regulation of genes in the fab gene cluster in other organisms, through binding to a palindromic DNA seq...

  15. Functional Dissection of the Blocking and Bypass Activities of the Fab-8 Boundary in the Drosophila Bithorax Complex.

    Directory of Open Access Journals (Sweden)

    Olga Kyrchanova


    Full Text Available Functionally autonomous regulatory domains direct the parasegment-specific expression of the Drosophila Bithorax complex (BX-C homeotic genes. Autonomy is conferred by boundary/insulator elements that separate each regulatory domain from its neighbors. For six of the nine parasegment (PS regulatory domains in the complex, at least one boundary is located between the domain and its target homeotic gene. Consequently, BX-C boundaries must not only block adventitious interactions between neighboring regulatory domains, but also be permissive (bypass for regulatory interactions between the domains and their gene targets. To elucidate how the BX-C boundaries combine these two contradictory activities, we have used a boundary replacement strategy. We show that a 337 bp fragment spanning the Fab-8 boundary nuclease hypersensitive site and lacking all but 83 bp of the 625 bp Fab-8 PTS (promoter targeting sequence fully rescues a Fab-7 deletion. It blocks crosstalk between the iab-6 and iab-7 regulatory domains, and has bypass activity that enables the two downstream domains, iab-5 and iab-6, to regulate Abdominal-B (Abd-B transcription in spite of two intervening boundary elements. Fab-8 has two dCTCF sites and we show that they are necessary both for blocking and bypass activity. However, CTCF sites on their own are not sufficient for bypass. While multimerized dCTCF (or Su(Hw sites have blocking activity, they fail to support bypass. Moreover, this bypass defect is not rescued by the full length PTS. Finally, we show that orientation is critical for the proper functioning the Fab-8 replacement. Though the inverted Fab-8 boundary still blocks crosstalk, it disrupts the topology of the Abd-B regulatory domains and does not support bypass. Importantly, altering the orientation of the Fab-8 dCTCF sites is not sufficient to disrupt bypass, indicating that orientation dependence is conferred by other factors.

  16. Oriented Immobilization of Fab Fragments by Site-Specific Biotinylation at the Conserved Nucleotide Binding Site for Enhanced Antigen Detection. (United States)

    Mustafaoglu, Nur; Alves, Nathan J; Bilgicer, Basar


    Oriented immobilization of antibodies and antibody fragments has become increasingly important as a result of the efforts to reduce the size of diagnostic and sensor devices to miniaturized dimensions for improved accessibility to the end-user. Reduced dimensions of sensor devices necessitate the immobilized antibodies to conserve their antigen binding activity for proper operation. Fab fragments are becoming more commonly used in small-scaled diagnostic devices due to their small size and ease of manufacture. In this study, we used the previously described UV-NBS(Biotin) method to functionalize Fab fragments with IBA-EG11-Biotin linker utilizing UV energy to initiate a photo-cross-linking reaction between the nucleotide binding site (NBS) on the Fab fragment and IBA-Biotin molecule. Our results demonstrate that immobilization of biotinylated Fab fragments via UV-NBS(Biotin) method generated the highest level of immobilized Fab on surfaces when compared to other typical immobilization methods while preserving antigen binding activity. UV-NBS(Biotin) method provided 432-fold, 114-fold, and 29-fold improved antigen detection sensitivity than physical adsorption, NHS-Biotin, and ε-NH3(+), methods, respectively. Additionally, the limit of detection (LOD) for PSA utilizing Fab fragments immobilized via UV-NBS(Biotin) method was significantly lower than that of the other immobilization methods, with an LOD of 0.4 pM PSA. In summary, site-specific biotinylation of Fab fragments without structural damage or loss in antigen binding activity provides a wide range of application potential for UV-NBS immobilization technique across numerous diagnostic devices and nanotechnologies.

  17. A Fab-Selective Immunoglobulin-Binding Domain from Streptococcal Protein G with Improved Half-Life Extension Properties.

    Directory of Open Access Journals (Sweden)

    Felix Unverdorben

    Full Text Available Half-life extension strategies have gained increasing interest to improve the pharmacokinetic and pharmacodynamic properties of protein therapeutics. Recently, we established an immunoglobulin-binding domain (IgBD from streptococcal protein G (SpGC3 as module for half-life extension. SpGC3 is capable of binding to the Fc region as well as the CH1 domain of Fab arms under neutral and acidic conditions.Using site-directed mutagenesis, we generated a Fab-selective mutant (SpGC3Fab to avoid possible interference with the FcRn-mediated recycling process and improved its affinity for mouse and human IgG by site-directed mutagenesis and phage display selections. In mice, this affinity-improved mutant (SpGC3FabRR conferred prolonged plasma half-lives compared with SpGC3Fab when fused to small recombinant antibody fragments, such as single-chain Fv (scFv and bispecific single-chain diabody (scDb. Hence, the SpGC3FabRR domain seems to be a suitable fusion partner for the half-life extension of small recombinant therapeutics.The half-life extension properties of SpGC3 can be retained by restricting binding to the Fab fragment of serum immunoglobulins and can be improved by increasing binding activity. The modified SpGC3 module should be suitable to extend the half-life of therapeutic proteins and, thus to improve therapeutic activity.

  18. Cholestatic liver disease after rituximab and adalimumab and the possible role of cross-reacting antibodies to Fab 2 fragments.

    Directory of Open Access Journals (Sweden)

    Joerg Latus

    Full Text Available BACKGROUND: Millions of patients are treated with therapeutic monoclonal antibodies (Tmabs for miscellaneous diseases. We investigated sera from six patients who received immune globulin, from one patient with refractory anti-neutrophil-cytoplasmic antibody (ANCA-associated granulomatosis with polyangiitis (GPA who developed two episodes of acute cholestatic liver disease, one after treatment with rituximab and a second after adalimumab and a healthy control group. METHODS: Three sera from the patient and six sera from patients who received immune globulin were analyzed for antibodies to rituximab and adalimumab by ELISA. Additionally, sera from the patients and from nine healthy blood donors were coated with the Fab fragment of an unrelated humanized monoclonal antibody, with human Fc proteins as well as a mouse IgG globulin. RESULTS: Viral serology for hepatitis A, B, C and autoantibodies specific for autoimmune liver disorders were negative. In all three sera from the patient antibodies to rituximab could be detected, but also antibodies to adalimumab were present even at time points when the patient had not yet received adalimumab, indicating cross reactivity between both substances. Testing against an unrelated human Fab fragment revealed positive results, indicating that the patient had antibodies against human Fab fragments in general. The Fc proteins were negative, and patients' sera did also not react with mouse IgG globulins. Remarkably, 2 out of 5 patients which were treated with immune globulin had antibodies against human Fab fragments in general whereas in none of the samples from healthy controls antibodies to Fab fragment could be detected. CONCLUSION: This is the first study demonstrating cholestatic liver disease induced by two different Tmabs. Cross - reacting antibodies to Fab2 fragments in general are probably involved. Further studies must show if these Fab2 antibodies in general are related with drug-induced side effects

  19. Functional Dissection of the Blocking and Bypass Activities of the Fab-8 Boundary in the Drosophila Bithorax Complex. (United States)

    Kyrchanova, Olga; Mogila, Vladic; Wolle, Daniel; Deshpande, Girish; Parshikov, Alexander; Cléard, Fabienne; Karch, Francois; Schedl, Paul; Georgiev, Pavel


    Functionally autonomous regulatory domains direct the parasegment-specific expression of the Drosophila Bithorax complex (BX-C) homeotic genes. Autonomy is conferred by boundary/insulator elements that separate each regulatory domain from its neighbors. For six of the nine parasegment (PS) regulatory domains in the complex, at least one boundary is located between the domain and its target homeotic gene. Consequently, BX-C boundaries must not only block adventitious interactions between neighboring regulatory domains, but also be permissive (bypass) for regulatory interactions between the domains and their gene targets. To elucidate how the BX-C boundaries combine these two contradictory activities, we have used a boundary replacement strategy. We show that a 337 bp fragment spanning the Fab-8 boundary nuclease hypersensitive site and lacking all but 83 bp of the 625 bp Fab-8 PTS (promoter targeting sequence) fully rescues a Fab-7 deletion. It blocks crosstalk between the iab-6 and iab-7 regulatory domains, and has bypass activity that enables the two downstream domains, iab-5 and iab-6, to regulate Abdominal-B (Abd-B) transcription in spite of two intervening boundary elements. Fab-8 has two dCTCF sites and we show that they are necessary both for blocking and bypass activity. However, CTCF sites on their own are not sufficient for bypass. While multimerized dCTCF (or Su(Hw)) sites have blocking activity, they fail to support bypass. Moreover, this bypass defect is not rescued by the full length PTS. Finally, we show that orientation is critical for the proper functioning the Fab-8 replacement. Though the inverted Fab-8 boundary still blocks crosstalk, it disrupts the topology of the Abd-B regulatory domains and does not support bypass. Importantly, altering the orientation of the Fab-8 dCTCF sites is not sufficient to disrupt bypass, indicating that orientation dependence is conferred by other factors.

  20. A RAD-tag genetic map for the platyfish (Xiphophorus maculatus) reveals mechanisms of karyotype evolution among teleost fish. (United States)

    Amores, Angel; Catchen, Julian; Nanda, Indrajit; Warren, Wesley; Walter, Ron; Schartl, Manfred; Postlethwait, John H


    Mammalian genomes can vary substantially in haploid chromosome number even within a small taxon (e.g., 3-40 among deer alone); in contrast, teleost fish genomes are stable (24-25 in 58% of teleosts), but we do not yet understand the mechanisms that account for differences in karyotype stability. Among perciform teleosts, platyfish (Xiphophorus maculatus) and medaka (Oryzias latipes) both have 24 chromosome pairs, but threespine stickleback (Gasterosteus aculeatus) and green pufferfish (Tetraodon nigroviridis) have just 21 pairs. To understand the evolution of teleost genomes, we made a platyfish meiotic map containing 16,114 mapped markers scored on 267 backcross fish. We tiled genomic contigs along the map to create chromosome-length genome assemblies. Genome-wide comparisons of conserved synteny showed that platyfish and medaka karyotypes remained remarkably similar with few interchromosomal translocations but with numerous intrachromosomal rearrangements (transpositions and inversions) since their lineages diverged ∼120 million years ago. Comparative genomics with platyfish shows how reduced chromosome numbers in stickleback and green pufferfish arose by fusion of pairs of ancestral chromosomes after their lineages diverged from platyfish ∼195 million years ago. Zebrafish and human genomes provide outgroups to root observed changes. These studies identify likely genome assembly errors, characterize chromosome fusion events, distinguish lineage-independent chromosome fusions, show that the teleost genome duplication does not appear to have accelerated the rate of translocations, and reveal the stability of syntenies and gene orders in teleost chromosomes over hundreds of millions of years.

  1. Extended incubation affects larval morphology, hatching success and starvation resistance in a terrestrially spawning fish, Galaxias maculatus (Jenyns 1842). (United States)

    Semmens, D; Swearer, S E


    The effect of extended incubation (delayed hatching) on larval morphology in the terrestrially spawning common galaxias Galaxias maculatus was investigated by inducing larvae to hatch 1 and 2 weeks after the normal 2 week incubation period. After 1 week of extended incubation, larvae were larger (longer in standard length, L(S), and greater in body depth) compared to controls (larvae that experienced normal incubation durations). After 2 weeks of extended incubation, larvae were smaller (shorter in L(S) and smaller in body depth) than larvae that experienced 1 week of extended incubation. Furthermore, eye area increased while yolk-sac size decreased monotonically with increasing incubation duration. These results suggest that larvae experiencing long periods of extended incubation are using somatic tissue to meet their metabolic demands. Larvae that experienced 2 weeks of extended incubation succumbed to starvation sooner than control larvae, but hatching success was not significantly different. Temperature mediated the effect of extended incubation on the morphology of larvae at hatching, most likely, through its effects on developmental rate and efficiency of yolk utilization. This study demonstrates some of the consequences of terrestrial spawning with extended incubation, which will assist in determining why this intriguing behaviour has evolved several times in a diverse range of taxa.

  2. Male-biased sex ratio does not promote increased sperm competitiveness in the seed beetle, Callosobruchus maculatus. (United States)

    McNamara, Kathryn B; Robinson, Stephen P; Rosa, Márta E; Sloan, Nadia S; van Lieshout, Emile; Simmons, Leigh W


    Sperm competition risk and intensity can select for adaptations that increase male fertilisation success. Evolutionary responses are examined typically by generating increased strength of sexual selection via direct manipulation of female mating rates (by enforcing monandry or polyandry) or by alteration of adult sex ratios. Despite being a model species for sexual selection research, the effect of sexual selection intensity via adult sex-ratio manipulation on male investment strategies has not been investigated in the seed beetle, Callosobruchus maculatus. We imposed 32 generations of experimental evolution on 10 populations of beetles by manipulating adult sex ratio. Contrary to predictions, males evolving in male-biased populations did not increase their testes and accessory gland size. This absence of divergence in ejaculate investment was also reflected in the fact that males from male-biased populations were not more successful in either preventing females from remating, or in competing directly for fertilisations. These populations already demonstrate divergence in mating behaviour and immunity, suggesting sufficient generations have passed to allow divergence in physiological and behavioural traits. We propose several explanations for the absence of divergence in sperm competitiveness among our populations and the pitfalls of using sex ratio manipulation to assess evolutionary responses to sexual selection intensity.

  3. Novel seminal fluid proteins in the seed beetle Callosobruchus maculatus identified by a proteomic and transcriptomic approach. (United States)

    Bayram, H; Sayadi, A; Goenaga, J; Immonen, E; Arnqvist, G


    The seed beetle Callosobruchus maculatus is a significant agricultural pest and increasingly studied model of sexual conflict. Males possess genital spines that increase the transfer of seminal fluid proteins (SFPs) into the female body. As SFPs alter female behaviour and physiology, they are likely to modulate reproduction and sexual conflict in this species. Here, we identified SFPs using proteomics combined with a de novo transcriptome. A prior 2D-sodium dodecyl sulphate polyacrylamide gel electrophoresis analysis identified male accessory gland protein spots that were probably transferred to the female at mating. Proteomic analysis of these spots identified 98 proteins, a majority of which were also present within ejaculates collected from females. Standard annotation workflows revealed common functional groups for SFPs, including proteases and metabolic proteins. Transcriptomic analysis found 84 transcripts differentially expressed between the sexes. Notably, genes encoding 15 proteins were highly expressed in male abdomens and only negligibly expressed within females. Most of these sequences corresponded to 'unknown' proteins (nine of 15) and may represent rapidly evolving SFPs novel to seed beetles. Our combined analyses highlight 44 proteins for which there is strong evidence that they are SFPs. These results can inform further investigation, to better understand the molecular mechanisms of sexual conflict in seed beetles.

  4. Selection in a fluctuating environment and the evolution of sexual dimorphism in the seed beetle Callosobruchus maculatus. (United States)

    Hallsson, L R; Björklund, M


    Temperature changes in the environment, which realistically include environmental fluctuations, can create both plastic and evolutionary responses of traits. Sexes might differ in either or both of these responses for homologous traits, which in turn has consequences for sexual dimorphism and its evolution. Here, we investigate both immediate changes in and the evolution of sexual dimorphism in response to a changing environment (with and without fluctuations) using the seed beetle Callosobruchus maculatus. We investigate sex differences in plasticity and also the genetic architecture of body mass and developmental time dimorphism to test two existing hypotheses on sex differences in plasticity (adaptive canalization hypothesis and condition dependence hypothesis). We found a decreased sexual size dimorphism in higher temperature and that females responded more plastically than males, supporting the condition dependence hypothesis. However, selection in a fluctuating environment altered sex-specific patterns of genetic and environmental variation, indicating support for the adaptive canalization hypothesis. Genetic correlations between sexes (r(MF) ) were affected by fluctuating selection, suggesting facilitated independent evolution of the sexes. Thus, the selective past of a population is highly important for the understanding of the evolutionary dynamics of sexual dimorphism.

  5. Cowpea bruchid Callosobruchus maculatus uses a three-component strategy to overcome a plant defensive cysteine protease inhibitor. (United States)

    Zhu-Salzman, K; Koiwa, H; Salzman, R A; Shade, R E; Ahn, J-E


    The soybean cysteine protease inhibitor, soyacystatin N (scN), negatively impacts growth and development of the cowpea bruchid, Callosobruchus maculatus[Koiwa et al. (1998) Plant J 14: 371-379]. However, the developmental delay and feeding inhibition caused by dietary scN occurred only during the early developmental stages (the 1st, 2nd and 3rd instars) of the cowpea bruchid. The 4th instar larvae reared on scN diet (adapted) exhibited rates of feeding and development which were comparable to those feeding on an scN-free diet (unadapted) prior to pupation. Total gut proteolytic capacity at this larval stage significantly increased in the scN-adapted insects. The elevated enzymatic activity was attributed to a differential expression of insect gut cysteine proteases (representing the major digestive enzymes), and of aspartic proteases. scN degradation by the gut extract was observed only in adapted bruchids, and this activity appeared to be a combined effect of scN-induced cysteine and aspartic proteases. Thirty cDNAs encoding cathepsin L-like cysteine proteases were isolated from insect guts, and they were differentially regulated by dietary scN. Our results suggest that the cowpea bruchid adapts to the challenge of scN by qualitative and quantitative remodelling of its digestive protease complement, and by activating scN-degrading protease activity.

  6. Salinity-dependent nickel accumulation and effects on respiration, ion regulation and oxidative stress in the galaxiid fish, Galaxias maculatus. (United States)

    Blewett, Tamzin A; Wood, Chris M; Glover, Chris N


    Inanga (Galaxias maculatus) are a euryhaline and amphidromous Southern hemisphere fish species inhabiting waters highly contaminated in trace elements such as nickel (Ni). Ni is known to exert its toxic effects on aquatic biota via three key mechanisms: inhibition of respiration, impaired ion regulation, and stimulation of oxidative stress. Inanga acclimated to freshwater (FW), 50% seawater (SW) or 100% SW were exposed to 0, 150 or 2000 μg Ni L(-1), and tissue Ni accumulation, metabolic rate, ion regulation (tissue ions, calcium (Ca) ion influx), and oxidative stress (catalase activity, protein carbonylation) were measured after 96 h. Ni accumulation increased with Ni exposure concentration in gill, gut and remaining body, but not in liver. Only in the gill was Ni accumulation affected by exposure salinity, with lower branchial Ni burdens in 100% and 50% SW inanga, relative to FW fish. There were no Ni-dependent effects on respiration, or Ca influx, and the only Ni-dependent effect on tissue ion content was on gill potassium. Catalase activity and protein carbonylation were affected by Ni, primarily in FW, but only at 150 μg Ni L(-1). Salinity therefore offsets the effects of Ni, despite minimal changes in Ni bioavailability. These data suggest only minor effects of Ni in inanga, even at highly elevated environmental Ni concentrations.

  7. Genetic variability and resistance of cultivars of cowpea [Vigna unguiculata (L.) Walp] to cowpea weevil (Callosobruchus maculatus Fabr.). (United States)

    Vila Nova, M X; Leite, N G A; Houllou, L M; Medeiros, L V; Lira Neto, A C; Hsie, B S; Borges-Paluch, L R; Santos, B S; Araujo, C S F; Rocha, A A; Costa, A F


    The cowpea weevil (Callosobruchus maculatus Fabr.) is the most destructive pest of the cowpea bean; it reduces seed quality. To control this pest, resistance testing combined with genetic analysis using molecular markers has been widely applied in research. Among the markers that show reliable results, the inter-simple sequence repeats (ISSRs) (microsatellites) are noteworthy. This study was performed to evaluate the resistance of 27 cultivars of cowpea bean to cowpea weevil. We tested the resistance related to the genetic variability of these cultivars using ISSR markers. To analyze the resistance of cultivars to weevil, a completely randomized test design with 4 replicates and 27 treatments was adopted. Five pairs of the insect were placed in 30 grains per replicate. Analysis of variance showed that the number of eggs and emerged insects were significantly different in the treatments, and the means were compared by statistical tests. The analysis of the large genetic variability in all cultivars resulted in the formation of different groups. The test of resistance showed that the cultivar Inhuma was the most sensitive to both number of eggs and number of emerged adults, while the TE96-290-12-G and MNC99-537-F4 (BRS Tumucumaque) cultivars were the least sensitive to the number of eggs and the number of emerged insects, respectively.

  8. Male-biased sex ratio does not promote increased sperm competitiveness in the seed beetle, Callosobruchus maculatus (United States)

    McNamara, Kathryn B.; Robinson, Stephen P.; Rosa, Márta E.; Sloan, Nadia S.; van Lieshout, Emile; Simmons, Leigh W.


    Sperm competition risk and intensity can select for adaptations that increase male fertilisation success. Evolutionary responses are examined typically by generating increased strength of sexual selection via direct manipulation of female mating rates (by enforcing monandry or polyandry) or by alteration of adult sex ratios. Despite being a model species for sexual selection research, the effect of sexual selection intensity via adult sex-ratio manipulation on male investment strategies has not been investigated in the seed beetle, Callosobruchus maculatus. We imposed 32 generations of experimental evolution on 10 populations of beetles by manipulating adult sex ratio. Contrary to predictions, males evolving in male-biased populations did not increase their testes and accessory gland size. This absence of divergence in ejaculate investment was also reflected in the fact that males from male-biased populations were not more successful in either preventing females from remating, or in competing directly for fertilisations. These populations already demonstrate divergence in mating behaviour and immunity, suggesting sufficient generations have passed to allow divergence in physiological and behavioural traits. We propose several explanations for the absence of divergence in sperm competitiveness among our populations and the pitfalls of using sex ratio manipulation to assess evolutionary responses to sexual selection intensity. PMID:27306351

  9. Targeting human prostate cancer with 111In-labeled D2B IgG, F(ab')2 and Fab fragments in nude mice with PSMA-expressing xenografts. (United States)

    Lütje, Susanne; van Rij, Catharina M; Franssen, Gerben M; Fracasso, Giulio; Helfrich, Wijnand; Eek, Annemarie; Oyen, Wim J; Colombatti, Marco; Boerman, Otto C


    D2B is a new monoclonal antibody directed against an extracellular domain of prostate-specific membrane antigen (PSMA), which is overexpressed in prostate cancer. The potential of D2B IgG, and F(ab')2 and Fab fragments of this antibody for targeting prostate cancer was determined in mice bearing subcutaneous prostate cancer xenografts. The optimal time point for imaging was determined in biodistribution and microSPECT imaging studies with (111)In-D2B IgG, (111)In-capromab pendetide, (111)In-D2B F(ab')2 and (111)In-D2B Fab fragments in mice with PSMA-expressing LNCaP and PSMA-negative PC3 tumors at several time points after injection. All (111)In-labeled antibody formats specifically accumulated in the LNCaP tumors, with highest uptake of (111)In-D2B IgG and (111)In-capromab pendetide at 168 h p.i. (94.8 ± 19.2% injected dose per gram (ID/g) and 16.7 ± 2.2% ID/g, respectively), whereas uptake of (111)In-D2B F(ab')2 and (111)In-D2B Fab fragments peaked at 24 h p.i. (12.1 ± 3.0% ID/g and 15.1 ± 2.9% ID/g, respectively). Maximum LNCaP tumor-to-blood ratios were 13.0 ± 2.3 (168 h p.i.), 6.2 ± 0.7 (24 h p.i.), 23.0 ± 4.0 (24 h p.i.) and 4.5 ± 0.6 (168 h p.i.) for (111)In-D2B IgG, (111)In-F(ab')2, (111)In-Fab and (111)In-capromab pendetide, respectively. LNCaP tumors were clearly visualized with microSPECT with all antibody formats. This study demonstrates the feasibility of D2B IgG, F(ab')2 and Fab fragments for targeting PSMA-expressing prostate cancer xenografts.


    Institute of Scientific and Technical Information of China (English)

    龚振明; 李蓓; 陈鲁勇; 沈明泉


    用硫酸铵沉淀法对兔抗孕酮血清进行粗提后用DEAE Sepharose Fast Fl0W对IgG进行层析纯化,然后用胃蛋白酶切除Fc片段而制备F(ab')2,用β-巯基乙醇还原F(ab')2后得到Fab',再将Fab'与热稳定性较高的大豆过氧化物酶(SBP)偶联制得抗孕酮Fab'-SBP.将抗孕酮Fab'-SBP用于二级竞争酶联免疫法测定孕酮有以下优点:(1)减少因Fc片段与固相载体非特异性吸附而引起的高背景干扰;(2)由于Fab'和SBP在很温和的条件下偶联,可使偶联产物保持较高的免疫活性和酶活性.

  11. Feasibility study of the Fab fragment of a monoclonal antibody against tissue factor as a diagnostic tool. (United States)

    Tsumura, Ryo; Sato, Ryuta; Furuya, Fumiaki; Koga, Yoshikatsu; Yamamoto, Yoshiyuki; Fujiwara, Yuki; Yasunaga, Masahiro; Matsumura, Yasuhiro


    Tissue factor (TF) is expressed strongly in various types of cancer, especially cancers that are often refractory to treatment, such as pancreatic cancer. In this study, we compared the differences in the biophysical and pharmacological properties of whole IgG and the Fab fragment of anti-human TF monoclonal antibody (1849 antibodies), in order to determine their suitability for application in the diagnosis and treatment of cancers. In the biophysical examination, we investigated the characteristics of 1849-whole IgG and 1849-Fab by SPR sensing and confocal fluorescence microscopy analysis using recombinant human TF antigen and TF-overexpressing human pancreatic cancer cell line, BxPC3, respectively. After conjugation with Alexa-Flour-647, in vivo imaging was conducted in mice bearing BxPC3 xenograft tumors. Furthermore, the distribution of the conjugates in tumors and major organs was evaluated by ex vivo study. The in vitro experiments showed that 1849 antibodies had high affinity against TF antigen. In addition, 1849-Fab showed a faster dissociation rate from the antigen than 1849-whole IgG. In mice, 1849-Fab-Alexa-Flour-647 showed rapid renal clearance and faster tumor accumulation, achieving a high contrast signal over nearby normal tissues in the early phase and enhanced tumor penetration after administration. On the other hand, 1849-whole IgG-Alexa-Flour-647 showed slow clearance from the blood and sustained high tumor accumulation. These results suggest that 1849-Fab may be a useful tool for pancreatic cancer diagnosis.

  12. Tumour imaging by the detection of fibrin clots in tumour stroma using an anti-fibrin Fab fragment. (United States)

    Obonai, Toshifumi; Fuchigami, Hirobumi; Furuya, Fumiaki; Kozuka, Naoyuki; Yasunaga, Masahiro; Matsumura, Yasuhiro


    The diagnosis of early and aggressive types of cancer is important for providing effective cancer therapy. Cancer-induced fibrin clots exist only within lesions. Previously, we developed a monoclonal antibody (clone 102-10) that recognizes insoluble fibrin but not fibrinogen or soluble fibrin and confirmed that fibrin clots form continuously in various cancers. Here, we describe the development of a Fab fragment probe of clone 102-10 for tumour imaging. The distribution of 102-10 Fab was investigated in genetically engineered mice bearing pancreatic ductal adenocarcinoma (PDAC), and its effect on blood coagulation was examined. Immunohistochemical and ex vivo imaging revealed that 102-10 Fab was distributed selectively in fibrin clots in PDAC tumours 3 h after injection and that it disappeared from the body after 24 h. 102-10 Fab had no influence on blood coagulation or fibrinolysis. Tumour imaging using anti-fibrin Fab may provide a safe and effective method for the diagnosis of invasive cancers by detecting fibrin clots in tumour stroma.

  13. Deletion of fabN in Enterococcus faecalis results in unsaturated fatty acid auxotrophy and decreased release of inflammatory cytokines. (United States)

    Diederich, Ann-Kristin; Duda, Katarzyna A; Romero-Saavedra, Felipe; Engel, Regina; Holst, Otto; Huebner, Johannes


    The Gram-positive bacterium Enterococcus faecalis can cause life-threatening infections and is resistant to several commonly used antibiotics. The type II fatty acid pathway in bacteria is discussed as a potential target for antimicrobial therapy. However, it was shown that inhibition or deletion of its enzymes can be rescued in Gram-positive bacteria by supplementation with fatty acids. Here we show that by deletion of the fabN gene, which is essential for unsaturated fatty acid (UFA) synthesis in E. faecalis, growth is impaired but can be rescued by supplementation with oleic acid or human serum. Nonetheless, we demonstrate alterations of the UFA profile after supplementation with oleic acid in the ΔfabN mutant using a specific glycolipid. In addition, we demonstrate that cytokine release in vitro is almost abolished after stimulation of mouse macrophages by the mutant in comparison to the wild type. The results indicate that fabN is not a suitable target for antimicrobials as UFA auxotrophy can be overcome. However, deletion of fabN resulted in a decreased inflammatory response indicating that fabN and resulting UFA synthesis are relevant for virulence.

  14. 抗HBs Fab-IFNα融合蛋白的制备与初步鉴定%Preparation and preliminary identification of anti HBs Fab-IFNa fusion protein

    Institute of Scientific and Technical Information of China (English)

    陆慧琦; 宋杰; 叶伟民; 韩焕兴


    目的 构建抗HBsAg的pComb Fab-IFNα载体,原核表达具双重生物活性的抗HBs Fab-IFNα融合蛋白.方法 以pBAD-Fab和pBAD IFNα作为模板,分别扩增Fd、λ和IENα基因,经相应的限制性内切酶酶切后分3次克隆人pComBHss质粒,转化XL-1 Blue大肠埃希菌.限制性酶切和测序鉴定重组质粒,免疫蛋向印迹(Western blotting)和斑点印迹(Dot blotting)鉴定融合蛋白的表达及抗原结合活性.结果 重组载体的酶切、电泳及测序表明抗HBs Fab-IFNα基因克隆正确.表达产物经12%SDS-PAGE电泳、转印,Western blotting显示该融合蛋白分子量约为65kD,Dot blotting显示其与HBsAg具有结合能力.细胞病变抑制法测定IFNα生物学活性为7.8×104~5.1×105U/ml.结论 该原核系统成功表达了抗HBs Fab-IFNα融合蛋白,表明其既具有抗HBsAg结合能力,又具备IFNα的生物活性,为进一步的系统表达和应用研究提供了条件.%Objective To construct the anti HBsAg pComb Fab-IFNe vector, and to express the fusion protein consisting of IFN and anti-HBs Fab in prokaryon. Methods Using pBAD-IFN plasmid and pBAD-Fab plasmid as template, the anti-HBs Fd, and IFN were amplified separately with corresponding endonuclease sites by polymerase chain reaction (PCR). Each PCR product was digested with specific endonuclease and inserted into pComBHss vector, and then transformed into XL-1 Blue. The recombinant plasmid was isolated by miniprep for restriction analysis and sequencing. Fusion protein was identified by Western blotting and Dot blotting. Results The recombinant plasmid was confirmed by restriction electrophoresis and sequencing. Aliquots of human anti-HBs Fab-IFNα were concentrated and size fractionated by 12% SDS-PAGE and stained with Coomassie. The proteins were transferred to nitrocellulose, incubated with HRP- conjugated goat anti-human IgG Fab and Rah anti-human IFN. The molecular weight of the fusion protein was about 65kD. Anti HBs Fab-IFNa fusion

  15. Expression, purification and characterization of enoyl-ACP reductase II, FabK, from Porphyromonas gingivalis

    Energy Technology Data Exchange (ETDEWEB)

    Hevener, Kirk E.; Mehboob, Shahila; Boci, Teuta; Truong, Kent; Santarsiero, Bernard D.; Johnson, Michael E. (UIC)


    The rapid rise in bacterial drug resistance coupled with the low number of novel antimicrobial compounds in the discovery pipeline has led to a critical situation requiring the expedient discovery and characterization of new antimicrobial drug targets. Enzymes in the bacterial fatty acid synthesis pathway, FAS-II, are distinct from their mammalian counterparts, FAS-I, in terms of both structure and mechanism. As such, they represent attractive targets for the design of novel antimicrobial compounds. Enoyl-acyl carrier protein reductase II, FabK, is a key, rate-limiting enzyme in the FAS-II pathway for several bacterial pathogens. The organism, Porphyromonas gingivalis, is a causative agent of chronic periodontitis that affects up to 25% of the US population and incurs a high national burden in terms of cost of treatment. P. gingivalis expresses FabK as the sole enoyl reductase enzyme in its FAS-II cycle, which makes this a particularly appealing target with potential for selective antimicrobial therapy. Herein we report the molecular cloning, expression, purification and characterization of the FabK enzyme from P. gingivalis, only the second organism from which this enzyme has been isolated. Characterization studies have shown that the enzyme is a flavoprotein, the reaction dependent upon FMN and NADPH and proceeding via a Ping-Pong Bi-Bi mechanism to reduce the enoyl substrate. A sensitive assay measuring the fluorescence decrease of NADPH as it is converted to NADP{sup +} during the reaction has been optimized for high-throughput screening. Finally, protein crystallization conditions have been identified which led to protein crystals that diffract x-rays to high resolution.

  16. The FermiFab toolbox for fermionic many-particle quantum systems (United States)

    Mendl, Christian B.


    This paper introduces the FermiFab toolbox for many-particle quantum systems. It is mainly concerned with the representation of (symbolic) fermionic wavefunctions and the calculation of corresponding reduced density matrices (RDMs). The toolbox transparently handles the inherent antisymmetrization of wavefunctions and incorporates the creation/annihilation formalism. Thus, it aims at providing a solid base for a broad audience to use fermionic wavefunctions with the same ease as matrices in Matlab, say. Leveraging symbolic computation, the toolbox can greatly simply tedious pen-and-paper calculations for concrete quantum mechanical systems, and serves as "sandbox" for theoretical hypothesis testing. FermiFab (including full source code) is freely available as a plugin for both Matlab and Mathematica. Program summaryProgram title:FermiFab Catalogue identifier: AEIN_v1_0 Program summary URL: Program obtainable from: CPC Program Library, Queen's University, Belfast, N. Ireland Licensing provisions: Special license provided by the author No. of lines in distributed program, including test data, etc.: 1 165 461 No. of bytes in distributed program, including test data, etc.: 15 557 308 Distribution format: tar.gz Programming language: MATLAB 7.9, Mathematica 7.0, C Computer: PCs, Sun Solaris workstation Operating system: Any platform supporting MATLAB or Mathematica; tested with Windows (32 and 64 bit) and Sun Solaris. RAM: Case dependent Classification: 4.15 Nature of problem: Representation of fermionic wavefunctions, computation of RDMs (reduced density matrices) and handing of the creation/annihilation operator formalism. Solution method: Mapping of Slater determinants to bitfields, implementation of the creation/annihilation and RDM formalism by bit operations. Running time: Depends on the problem size; several seconds for the provided demonstration files.

  17. The life-cycle of the digenetic trematode, Proctoeces maculatus (Looss, 1901) Odhner, 1911 (Syn. P. rubtenuis [Linton, 1907] Hanson, 1950), and description of Cerceria adranocerca n. sp (United States)

    Stunkard, H.W.; Uzmann, J.R.


    The genus Proctoeces was erected by Odhner ( 191 1) to contain Distonium maculatuni Looss, 1901, from Labrus merula and Crenilabrus spp. at Triest. Odhner had found the parasite in Blennius ocellaris at Naples. One adult specimen from Chrysophrys bifasciata and two immature specimens from lulis lunaris taken in the Red Sea, were described as a new species, Proctoeces erythraeus. Dawes (1946) listed P. erythraeus as a synonym of  P. maculatus (Looss) , but the species was recognized by Manter ( 1947) on the basis of six specimens he had collected from Calamus calamus and Calamus bajonado at the biological laboratory of the Carnegie Institution at Dry Tortugas, Florida. Several additional species have been de scribed. Fujita ( 1925) reported a metacercaria from the Japanese oyster, Ostrea gigas, as a new species, Proctoeces ostreae. The paper was translated by R. Ph. Dollfus who noted (p. 57) ,“Il est à souhaiter que des recherches chez les poissons mangers de Lamellibranches, sur les côtes de la préfecture d'Hiroshima, permettent de découvrir des exemplaires complètement adultes de Proctoeces ostreae Fuj., chez lesquels l'extension des vitellogènes et les dimensions des oeufs puissent être observées avec précision; il sera alors possible de savoir définitivement si P. ostreae Fuj. doit ou non tomber en synonymie avec P. maculatus (Looss)." Yamaguti (1934) described P. maculatus from Sparus aries, Sparws macrocephalus, Pagrosomus auratus, and Epinephelus akaara in Japan. Several specimens from Pagrosomus auratus, which differed from P. maculatus in larger size, larger eggs, and trilobed ovary, he described as a new species, Proctoeces major. Yamaguti ( 1938) reported P. nzaculatus from Sensicossyphus reticulatus and described a larva from the liver of the pelecypod mollusk, Brachidontes senhausi, as an unidentified member of the genus Proctoeces. Manter ( 1940) described Proctoeces tnagnorus from a single specimen found in the intestine of Caulolatilus

  18. Preservation of Cowpea (Vigna unguiculata Seeds: Incidence of Ethanolic Extract from Balanites aegyptiaca, Melia azedarach and Ocimum gratissimum Leaves on Callosobruchus maculatus (Coleptera: Bruchidae

    Directory of Open Access Journals (Sweden)

    Ange-Patrice Takoudjou Miafo


    Full Text Available This study aims to evaluate the effect of ethanolic extracts of Balanites aegyptiaca, Melia azedarach and Ocimum gratissimum leaves as bioinsecticide in the preservation of cowpea seeds (Vigna unguiculata against the pest Callosobruchus maculatus. The extracts were obtained by maceration of leaves powdersin ethanol (95%. These extracts were then used to achieve anti-insecticides tests in jars at doses of 10, 15, 25 and 50% compared to a reference compound (Stargrain. The weevils rearing and some biological tests were conducted in laboratory conditions at a temperature of 29.1°C and a relative humidity of 74%. The results showed that these extracts have anti-oviposition activity and ovicidal dose-dependent. Doses 25 and 50% of Melia azedarach and Ocimum gratissimum have completely inhibited the spawning activity of C. maculatus 24 h after treatment. There was significant difference (p<0.05 between oviposition due to the positive control (Stargrain and that due to dose 50% of the three extracts. Compared to the negative control (ethanol 95%, all these plant extracts have significantly reduced (p<0.05 oviposition of C. maculatus female (38 eggs per 100 seeds after 4 days. The dose 10% of B. aegyptiaca and 50% of O. gratissimum showed no weevil emergency. It have been also noticed a reduction in seeds depreciation and rate attack in all the treated settings compared to the control. So the rate attack and the mass loss were proportional to the doses of B. aegyptiaca leaves extract but conversely proportional to doses of Melia azedarach and O. gratissimum leaves extracts. No depreciation of seeds was recorded at doses 10 and 15% of B. aegyptiaca and 50% of O. gratissimum. The different treatments did not affect the germination of seeds; the highest germination rate (93.67% was recorded with seeds treated with the dose 50% of O. gratissimum against only 65% with those treated with the positive control. These ethanolic extracts have shown insecticidal

  19. The distribution of neuropeptide Y and dynorphin immunoreactivity in the brain and pituitary gland of the platyfish, Xiphophorus maculatus, from birth to sexual maturity (United States)

    Cepriano, L. M.; Schreibman, M. P.


    Immunoreactive neuropeptide Y and dynorphin have been localized in the brain and pituitary gland of the platyfish, Xiphophorus maculatus, at different ages and stages of development from birth to sexual maturity. Immunoreactive neuropeptide Y was found in perikarya and tracts of the nucleus olfactoretinalis, telencephalon, ventral tegmentum and in the neurohypophysis and in the three regions of the adenohypophysis. Immunoreactive dynorphin was found in nerve tracts in the olfactory bulb and in cells of the pars intermedia and the rostral pars distalis of the pituitary gland.


    Directory of Open Access Journals (Sweden)

    Blondine Ch.P Blondine Ch.P


    Full Text Available Bacillus thuringiensis H-14 (local strain is a pathogenic bacteria which specijiclly target to mosquito larvae. It is safe for human, mammals and does not cause any environmental pollution. The aims of this study were to measure 1. the efficacy of liquid formula of B.thuringiensis H-14 local strain at several concentration in soybean infusion medium against An. maculatus, 2. the determine optimum concentration of the liquid formula of Bacillus thuringiensis H-14 local strain in soybean infusion medium against An. maculatus larvae in Kokap District Kulon Progo Regency DIY. This research was based on the quasi experiment with the Pre test-Post test Control Group Design. We tested 3 liquid formula of B. thuringiensis H-14 local strain with concentration of 1 x LC90 5 x LC90 and 10 x LC90 in 9 ponds each formula and one control sample. Other 9 ponds were selected as a control group of An. maculatus with 0.20 to 1.20 m2 width. The effectiveness ofthe liquid formula of B.thuringiensis H-14 local strain in soybean infusion medium concentration of 0.059 ml/l (1 x LC90, 0.295 ml/l (5 x LC90 and 0.59 ml/l (10 x LC90 against An. maculatus larvae which reached 70% were survived for 6.75 days, 8.35 days and 12.58 days. There was a significant difference between the effectiveness of the liquid formula of B. thuringiensis H-14 local strain in soybean infusion medium concentration of 0.059 ml/l (1 x LC90 and 0.295 ml/l (5 x LC90 and 0.59 ml/l (10 x LC90 at p<0.05. The liquid formula of B.thuringiensis H-14 local strain in soybean infusion medium could be used as the vector control agent. Keywords: Effectiveness , B. thuringiensis H-14, Soybean infusion

  1. Isolation and purification of a papain inhibitor from Egyptian genotypes of barley seeds and its in vitro and in vivo effects on the cowpea bruchid, Callosobruchus maculatus (F.). (United States)

    Abd El-Latif, Ashraf Oukasha


    The cysteine inhibitors that are known as cystatin have been identified and characterized from several plant species. In the current study, 44 barley (Hordeum vulgare) genotypes including 3 varieties and 41 promising lines were screened for their potential as protease inhibitors. The barley genotypes showed low inhibitory activity against trypsin and chymotrypsin enzymes with a mean of 4.15 TIU/mg protein and 4.40 CIU/mg protein. The barley variety, Giza 123, showed strong papain inhibitory activity of 97.09 PIU/mg proteins and was subjected for further purification studies using ammonium sulfate fractionation and DEAE-Sephadex A-25 column. Barley purified proteins showed two bands on SDS-PAGE corresponding to a molecular mass of 12.4-54.8 kDa. The purified barley PI was found to be stable at a temperature below 80 °C and at a wide range of pH from 2 to 12. Barley PI was found to have higher potential inhibitory activity against papain enzyme compared to the standard papain inhibitor, E-64 with an IC50 value of 21.04 µg/ml and 25.62 µg/ml for barley PI and E-64, respectively. The kinetic analysis revealed a non-competitive type of inhibition with a Ki value of 1.95 × 10(-3 )µM. The antimetabolic effect of barley PI was evaluated against C. maculatus by incorporating the F30-60 protein of the purified inhibitor into the artificial diet using artificial seeds. Barley PI significantly prolonged the development of C. maculatus in proportion to PI concentration. Barley PI significantly increased the mortality of C. maculatus and caused a significant reduction in its fecundity. On the other hand, barley PI seemed to have non-significant effects on the adult longevity and the adult dry weight. The in vitro and in vivo results proved the efficiency of the papain inhibitory protein isolated from barley as a tool for managing the cowpea bruchid, C. maculatus.

  2. Evaluation of vacuum technology to kill larvae of the Asian longhorned beetle, Anoplophora glabripennis (Coleoptera: Cerambycidae), and the emerald ash borer, Agrilus planipennis (Coleoptera: Buprestidae), in wood (United States)

    Zhangjing Chen; Marshall S. White; Melody A. Keena; Therese M. Poland; Erin L. Clark


    The potential for using vacuum technology to kill larvae of the Asian longhorned beetle (ALB), Anoplophora glabripennis (Motschulsky) (Coleoptera: Cerambycidae), and emerald ash borer (EAB), Agrilus planipennis Fairmaire (Coleoptera: Buprestidae), in solid-wood packing materials (SWPM) and other wood products was assessed. Current...

  3. Molecular markers detect cryptic predation on coffee berry borer (Coleoptera: Curculionidae) by silvanid and laemophloeid flat bark beetles (Coleoptera: Silvanidae, Laemophloeidae) in coffee beans (United States)

    The coffee berry borer, Hypothenemus hampei(Coleoptera: Curculionidae)(Ferrari), is a serious pest of coffee worldwide and has been recently introduced in Hawai’i, first detected in the state in 2010. Adult silvanid flat bark beetles, Cathartus quadricollis (Coleoptera: Silvanidae) and adult laemoph...

  4. Use of FabV-Triclosan Plasmid Selection System for Efficient Expression and Production of Recombinant Proteins in Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Syed A Ali

    Full Text Available Maintenance of recombinant plasmid vectors in host bacteria relies on the presence of selection antibiotics in the growth media to suppress plasmid -free segregants. However, presence of antibiotic resistance genes and antibiotics themselves is not acceptable in several applications of biotechnology. Previously, we have shown that FabV-Triclosan selection system can be used to select high and medium copy number plasmid vectors in E. coli. Here, we have extended our previous work and demonstrated that expression vectors containing FabV can be used efficiently to express heterologous recombinant proteins in similar or better amounts in E. coli host when compared with expression vectors containing β-lactamase. Use of small amount of non-antibiotic Triclosan as selection agent in growth medium, enhanced plasmid stability, applicability in various culture media, and compatibility with other selection systems for multiple plasmid maintenance are noteworthy features of FabV-Triclosan selection system.


    Directory of Open Access Journals (Sweden)

    M. Pour-Amir


    Full Text Available In this study we produced and purified a high titer of specific and high affin¬ity Fab fragments of anti-digoxin antibody. Immunization of rabbits with a conju¬gate of the cardiac glycoside digoxin, coupled by a periodate oxidation method to the amino group of lysine in bovine serum albumin resulted in the production of this type of high titer digoxin-specific antibodies with exceptionally high affinity (109 L/mol and specificity in immune response. Increase in titer was found in steps of purification ending up with the highest titer for Fab fragment to be at 1.75 ug of purified Fab (for 50% binding of I25I-digoxin. High specificity for antigenic determinants of the steroid nucleus of digoxin was observed such that much less cross-reaction with digoxin (2.3% and no cross-reaction with ouabaine, estradiol, Cortisol, progesterone and testosterone were detected.

  6. Critical Stage Rule-Based Real Time Dispatch(RTD)System in Highly-Mixed-Products (HMP) FAB

    Institute of Scientific and Technical Information of China (English)

    YUXiao-hua; XIANGYu-qun


    An improving utilization and efficiency of critical equipments in semiconductor wafer fabrication facilities are concerned. Semiconductor manufacturing FAB is one of the most complicated and cost sensitive environments. A good dispatching tool will make big difference in equipment utilization and FAB output as a whole. The equipment in this paper is In-Line DUV Scanner.There are many factors impacting utilization and output on this equipment group. In HMP environment one of the issues is changing of reticule in this area and idle counts due to load unbalance between equipments. Here we'll introduce a rule-based RTD system which aiming at decreasing the number of recipe change and idle counts among a group of scanner equipment in a high-mixedproducts FAB.

  7. Critical Stage Rule-Based Real Time Dispatch(RTD) System in Highly-Mixed-Products (HMP) FAB

    Institute of Scientific and Technical Information of China (English)

    YU Xiao-hua; XIANG Yu-qun


    An improving utilization and efficiency of critical equipments in semiconductor wafer fabrication facilities are concerned. Semiconductor manufacturing FAB is one of the most omplicated and cost sensitive environments. A good dispatching tool will make big difference in equipment utilization and FAB output as a whole. The equipment in this paper is In-Line DUV Scanner.There are many factors impacting utilization and output on this equipment group. In HMP environment one of the issues is changing of reticule in this area and idle counts due to load unbalance between equipments. Here we'll introduce a rule-based RTD system which aiming at decreasing the number of recipe change and idle counts among a group of scanner equipment in a high-mixedproducts FAB.

  8. Antibacterial FabH Inhibitors with Mode of Action Validated in Haemophilus influenzae by in Vitro Resistance Mutation Mapping. (United States)

    McKinney, David C; Eyermann, Charles J; Gu, Rong-Fang; Hu, Jun; Kazmirski, Steven L; Lahiri, Sushmita D; McKenzie, Andrew R; Shapiro, Adam B; Breault, Gloria


    Fatty acid biosynthesis is essential to bacterial growth in Gram-negative pathogens. Several small molecules identified through a combination of high-throughput and fragment screening were cocrystallized with FabH (β-ketoacyl-acyl carrier protein synthase III) from Escherichia coli and Streptococcus pneumoniae. Structure-based drug design was used to merge several scaffolds to provide a new class of inhibitors. After optimization for Gram-negative enzyme inhibitory potency, several compounds demonstrated antimicrobial activity against an efflux-negative strain of Haemophilus influenzae. Mutants resistant to these compounds had mutations in the FabH gene near the catalytic triad, validating FabH as a target for antimicrobial drug discovery.

  9. Blood cytology of the common jollytail (Galaxias maculatus (Jenyns, 1842 (Osmeriformes: Galaxiidae at postlarval and adult stages Estudio de la citología sanguínea del puye (Galaxias maculatus (Jenyns, 1842 (Osmeriformes: Galaxiidae en estado postlarval y adulto

    Directory of Open Access Journals (Sweden)

    I Valdebenito


    Full Text Available “Puye” (Galaxias maculatus is a small freshwater fish species of great interest to Chilean aquaculture diversification, because of the high commercial value reached by its transparent larvae or “cristalino”. This study presents the first data with regard to blood cytology in larvae and adults of this specie. Blood smears stained with May Grünwald-Giemsa were analysed through optical microscopy. The results show that in G. maculatus larvae the flowing blood has a transparent look and mature erythrocytes were not observed, as opposed to the adults of this species where the complete erythrocyte developmental line could be observed. In leucocytes, there was no presence of coarse grain cells such as basophils and eosinophyls. In general, the morphology of the blood cells found in larvae and adults of G. maculatus have the same characteristic traits as described elsewhere in reports of other teleosts.El puye (Galaxias maculatus es un pequeño pez nativo de gran interés para la diversificación de la acuicultura chilena, debido al alto valor comercial de su estado postlarval cristalino. La presente investigación entrega los primeros antecedentes respecto de la citología sanguínea de postlarvas y adultos de esta especie, determinada mediante microscopía óptica utilizando frotis sanguíneos preparados con May Grünwald-Giemsa. Los resultados muestran que en la sangre circulante de G. maculatus en estado de postlarva no se observan eritrocitos maduros y la sangre es de aspecto transparente. Sin embargo, en adultos se pudo observar la línea eritrocitaria completa y en leucocitos no se observó la presencia de células de grano grueso como son los basófilos y eosinófilos. La morfología de las células encontradas en ambos estados del desarrollo del puye mantienen las características propias entregadas en la literatura para peces teleósteos.

  10. A study on EUV reticle surface molecular contamination under different storage conditions in a HVM foundry fab (United States)

    Singh, SherJang; Yatzor, Brett; Taylor, Ron; Wood, Obert; Mangat, Pawitter


    The prospect of EUVL (Extreme Ultraviolet Lithography) insertion into HVM (High Volume Manufacturing) has never been this promising. As technology is prepared for "lab to fab" transition, it becomes important to comprehend challenges associated with integrating EUVL infrastructure within existing high volume chip fabrication processes in a foundry fab. The existing 193nm optical lithography process flow for reticle handling and storage in a fab atmosphere is well established and in-fab reticle contamination concerns are mitigated with the reticle pellicle. However EUVL reticle pellicle is still under development and if available, may only provide protection against particles but not molecular contamination. HVM fab atmosphere is known to be contaminated with trace amounts of AMC's (Atmospheric Molecular Contamination). If such contaminants are organic in nature and get absorbed on the reticle surface, EUV photon cause photo-dissociation resulting into carbon generation which is known to reduce multilayer reflectivity and also degrades exposure uniformity. Chemical diffusion and aggregation of other ions is also reported under the e-beam exposure of a EUV reticle which is known to cause haze issues in optical lithography. Therefore it becomes paramount to mitigate absorbed molecular contaminant concerns on EUVL reticle surface. In this paper, we have studied types of molecular contaminants that are absorbed on an EUVL reticle surface under HVM fab storage and handling conditions. Effect of storage conditions (gas purged vs atmospheric) in different storage pods (Dual pods, Reticle Clamshells) is evaluated. Absorption analysis is done both on ruthenium capping layer as well as TaBN absorber. Ru surface chemistry change as a result of storage is also studied. The efficacy of different reticle cleaning processes to remove absorbed contaminant is evaluated as well.

  11. Rapid adaptation to a novel host in a seed beetle (Callosobruchus maculatus): the role of sexual selection. (United States)

    Fricke, Claudia; Arnqvist, Göran


    Rapid diversification is common among herbivorous insects and is often the result of host shifts, leading to the exploitation of novel food sources. This, in turn, is associated with adaptive evolution of female oviposition behavior and larval feeding biology. Although natural selection is the typical driver of such adaptation, the role of sexual selection is less clear. In theory, sexual selection can either accelerate or impede adaptation. To assess the independent effects of natural and sexual selection on the rate of adaptation, we performed a laboratory natural selection experiment in a herbivorous bruchid beetle (Callosobruchus maculatus). We established replicated selection lines where we varied natural (food type) and sexual (mating system) selection in a 2 x 2 orthogonal design, and propagated our lines for 35 generations. In half of the lines, we induced a host shift whereas the other half was kept on the ancestral host. We experimentally enforced monogamy in half of the lines, whereas the other half remained polygamous. The beetles rapidly adapted to the novel host, which primarily involved increased host acceptance by females and an accelerated rate of larval development. We also found that our mating system treatment affected the rate of adaptation, but that this effect was contingent upon food type. As beetles adapted to the novel host, sexual selection reinforced natural selection whereas populations residing close to their adaptive peak (i.e., those using their ancestral host) exhibited higher fitness in the absence of sexual selection. We discuss our findings in light of current sexual selection theory and suggest that the net evolutionary effect of reproductive competition may critically depend on natural selection. Sexual selection may commonly accelerate adaptation under directional natural selection whereas sexual selection, and the associated load brought by sexual conflict, may tend to depress population fitness under stabilizing natural

  12. Developmental temperature affects the expression of ejaculatory traits and the outcome of sperm competition in Callosobruchus maculatus. (United States)

    Vasudeva, R; Deeming, D C; Eady, P E


    The outcome of post-copulatory sexual selection is determined by a complex set of interactions between the primary reproductive traits of two or more males and their interactions with the reproductive traits of the female. Recently, a number of studies have shown the primary reproductive traits of both males and females express phenotypic plasticity in response to the thermal environment experienced during ontogeny. However, how plasticity in these traits affects the dynamics of sperm competition remains largely unknown. Here, we demonstrate plasticity in testes size, sperm size and sperm number in response to developmental temperature in the bruchid beetle Callosobruchus maculatus. Males reared at the highest temperature eclosed at the smallest body size and had the smallest absolute and relative testes size. Males reared at both the high- and low-temperature extremes produced both fewer and smaller sperm than males reared at intermediate temperatures. In the absence of sperm competition, developmental temperature had no effect on male fertility. However, under conditions of sperm competition, males reared at either temperature extreme were less competitive in terms of sperm offence (P(2)), whereas those reared at the lowest temperature were less competitive in terms of sperm defence (P(1)). This suggests the developmental pathways that regulate the phenotypic expression of these ejaculatory traits are subject to both natural and sexual selection: natural selection in the pre-ejaculatory environment and sexual selection in the post-ejaculatory environment. In nature, thermal heterogeneity during development is commonplace. Therefore, we suggest the interplay between ecology and development represents an important, yet hitherto underestimated component of male fitness via post-copulatory sexual selection.

  13. RNA-Seq analysis of salinity stress–responsive transcriptome in the liver of spotted sea bass (Lateolabrax maculatus) (United States)

    Zhang, Xiaoyan; Wen, Haishen; Wang, Hailiang; Ren, Yuanyuan; Zhao, Ji; Li, Yun


    Salinity is one of the most prominent abiotic factors, which greatly influence reproduction, development, growth, physiological and metabolic activities of fishes. Spotted sea bass (Lateolabrax maculatus), as a euryhaline marine teleost, has extraordinary ability to deal with a wide range of salinity changes. However, this species is devoid of genomic resources, and no study has been conducted at the transcriptomic level to determine genes responsible for salinity regulation, which impedes the understanding of the fundamental mechanism conferring tolerance to salinity fluctuations. Liver, as the major metabolic organ, is the key source supplying energy for iono- and osmoregulation in fish, however, little attention has been paid to its salinity-related functions but which should not be ignored. In this study, we perform RNA-Seq analysis to identify genes involved in salinity adaptation and osmoregulation in liver of spotted sea bass, generating from the fishes exposed to low and high salinity water (5 vs 30ppt). After de novo assembly, annotation and differential gene expression analysis, a total of 455 genes were differentially expressed, including 184 up-regulated and 271 down-regulated transcripts in low salinity-acclimated fish group compared with that in high salinity-acclimated group. A number of genes with a potential role in salinity adaptation for spotted sea bass were classified into five functional categories based on the gene ontology (GO) and enrichment analysis, which include genes involved in metabolites and ion transporters, energy metabolism, signal transduction, immune response and structure reorganization. The candidate genes identified in L. maculates liver provide valuable information to explore new pathways related to fish salinity and osmotic regulation. Besides, the transcriptomic sequencing data supplies significant resources for identification of novel genes and further studying biological questions in spotted sea bass. PMID:28253338

  14. ImmunoPET of tissue factor expression in triple-negative breast cancer with a radiolabeled antibody Fab fragment

    Energy Technology Data Exchange (ETDEWEB)

    Shi, Sixiang [University of Wisconsin, Materials Science Program, Madison, WI (United States); Hong, Hao; Orbay, Hakan; Yang, Yunan; Ohman, Jakob D. [University of Wisconsin, Department of Radiology, Madison, WI (United States); Graves, Stephen A.; Nickles, Robert J. [University of Wisconsin, Department of Medical Physics, Madison, WI (United States); Liu, Bai; Wong, Hing C. [Altor BioScience, Miramar, FL (United States); Cai, Weibo [University of Wisconsin, Materials Science Program, Madison, WI (United States); University of Wisconsin, Department of Radiology, Madison, WI (United States); University of Wisconsin, Department of Medical Physics, Madison, WI (United States); University of Wisconsin Carbone Cancer Center, Madison, WI (United States); University of Wisconsin, Departments of Radiology and Medical Physics, Madison, WI (United States)


    To date, there is no effective therapy for triple-negative breast cancer (TNBC), which has a dismal clinical outcome. Upregulation of tissue factor (TF) expression leads to increased patient morbidity and mortality in many solid tumor types, including TNBC. Our goal was to employ the Fab fragment of ALT-836, a chimeric anti-human TF mAb, for PET imaging of TNBC, which can be used to guide future TNBC therapy. ALT-836-Fab was generated by enzymatic papain digestion. SDS-PAGE and FACS studies were performed to evaluate the integrity and TF binding affinity of ALT-836-Fab before NOTA conjugation and {sup 64}Cu-labeling. Serial PET imaging and biodistribution studies were carried out to evaluate the tumor targeting efficacy and pharmacokinetics in the MDA-MB-231 TNBC model, which expresses high levels of TF on the tumor cells. Blocking studies, histological assessment, as well as RT-PCR were performed to confirm TF specificity of {sup 64}Cu-NOTA-ALT-836-Fab. ALT-836-Fab was produced with high purity, which exhibited superb TF binding affinity and specificity. Serial PET imaging revealed rapid and persistent tumor uptake of {sup 64}Cu-NOTA-ALT-836-Fab (5.1 ± 0.5 %ID/g at 24 h post-injection; n = 4) and high tumor/muscle ratio (7.0 ± 1.2 at 24 h post-injection; n = 4), several-fold higher than that of the blocking group and tumor models that do not express significant level of TF, which was confirmed by biodistribution studies. TF specificity of the tracer was also validated by histology and RT-PCR. {sup 64}Cu-NOTA-ALT-836-Fab exhibited prominent tissue factor targeting efficiency in MDA-MB-231 TNBC model. The use of a Fab fragment led to fast tumor uptake and good tissue/muscle ratio, which may be translated into same-day immunoPET imaging in the clinical setting to improve TNBC patient management. (orig.)

  15. Clave de las especies de Conoderus Grupo II (Coleoptera: Elateridae Key of the species of Conoderus Group II (Coleoptera: Elateridae

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    Marta E. Guzmán De Tomé


    Full Text Available RESUMEN: Se presenta una reseña histórica, diagnosis y clave de identificación de 33 especies exclusivamente neotropicales, del género Conoderus Eschscholtz 1829 Grupo II, (Coleoptera, Elateridae proporcionando, datos de su distribución e ilustraciones de cuatro especies representativas de la región.ABSTRACT. An identification of 33 species of Conoderus Group II, Eschscholtz 1829 (Coleoptera, Elateridae with full diagnosis, distribution, with representative illustrations of four species of the neotropical region.

  16. New species of Hemilophini (Coleoptera, Cerambycidae, Lamiinae) from Colombia and Ecuador. (United States)

    Monné, Marcela L; Monné, Miguel A


    Three new species of Hemilophini (Coleoptera, Cerambycidae, Lamiinae) are described: Chrysaperda mimica sp. nov. and Malacoscylus nearnsi sp. nov. from Ecuador, and Eulachnesia boteroi sp. nov. from Colombia.

  17. Repellency of Hydroethanolic Extracts of Ricinus communis (Euphorbiaceae) to Scyphophorus acupunctatus (Coleoptera: Curculionidae) in the Laboratory

    National Research Council Canada - National Science Library

    Cinthia Pacheco-Sánchez; Patricia Villa-Ayala; Roberto Montes-Belmont; Rodolfo Figueroa-Brito; Alfredo Jiménez-Pérez


    ABSTRACT The agave snout weevil Scyphophorus acupunctatus Gyllenhal (Coleoptera: Curculionidae) is an ubiquitous insect and the main pest of blue tequila agave, Agave tequilana Weber, and other agaves...

  18. β-Hydroxyacyl-acyl Carrier Protein Dehydratase (FabZ) from Francisella tularensis and Yersinia pestis : Structure Determination, Enzymatic Characterization, and Cross-Inhibition Studies

    Energy Technology Data Exchange (ETDEWEB)

    McGillick, Brian E.; Kumaran, Desigan; Vieni, Casey; Swaminathan, Subramanyam


    The bacterial system for fatty acid biosynthesis (FAS) contains several enzymes whose sequence and structure are highly conserved across a vast array of pathogens. This, coupled with their low homology and difference in organization compared to the equivalent system in humans, makes the FAS pathway an excellent target for antimicrobial drug development. To this end, we have cloned, expressed, and purified the β-hydroxyacyl-acyl carrier protein dehydratase (FabZ) from both Francisella tularensis (FtFabZ) and Yersinia pestis (YpFabZ). We also solved the crystal structures and performed an enzymatic characterization of both enzymes and several mutant forms of YpFabZ. Additionally, we have discovered two novel inhibitors of FabZ, mangostin and stictic acid, which show similar potencies against both YpFabZ and FtFabZ. Lastly, we selected several compounds from the literature that have been shown to be active against single homologues of FabZ and tested them against both YpFabZ and FtFabZ. These results have revealed clues as to which scaffolds are likely to lead to broad-spectrum antimicrobials targeted against FabZ as well as modifications to existing FabZ inhibitors that may improve potency.

  19. Structure of a human monoclonal antibody Fab fragment against gp41 of human immunodeficiency virus type (United States)

    He, X. M.; Ruker, F.; Casale, E.; Carter, D. C.


    The three-dimensional structure of a human monoclonal antibody (Fab), which binds specifically to a major epitope of the transmembrane protein gp41 of the human immunodeficiency virus type 1, has been determined by crystallographic methods to a resolution of 2.7 A. It has been previously determined that this antibody recognizes the epitope SGKLICTTAVPWNAS, belongs to the subclass IgG1 (kappa), and exhibits antibody-dependent cellular cytotoxicity. The quaternary structure of the Fab is in an extended conformation with an elbow bend angle between the constant and variable domains of 175 degrees. Structurally, four of the hypervariable loops can be classified according to previously recognized canonical structures. The third hypervariable loops of the heavy (H3) and light chain (L3) are structurally distinct. Hypervariable loop H3, residues 102H-109H, is unusually extended from the surface. The complementarity-determining region forms a hydrophobic binding pocket that is created primarily from hypervariable loops L3, H3, and H2.

  20. Nurturing Creativity and Innovation Through FabKids: A Case Study (United States)

    Beyers, Ronald Noel


    This paper will report on a case study that was conducted involving Grade 10 learners who were exposed to a high-tech rapid-prototyping environment of a Fabrication Laboratory as part of a FabKids experience. This project must be viewed in the context of a global shortage of key skills placing a higher priority on the initiation and development of a pipeline to attract youth into science, engineering and technology careers. Creativity and innovation feature high on the skills agenda but more importantly preliminary results indicate that learners from a broad range of schools were able to operate effectively in this post constructivist environment. Participants had to apply their knowledge, skill, attitudes and values in order to produce a solution to the challenges provided. The fundamentals of the design process of investigate, design, make, evaluate and communicate were emphasized where the FabKids had to draw on their own collective knowledge using a range of technologies available to them.

  1. Anti-neuropilin 1 antibody Fab' fragment conjugated liposomal docetaxel for active targeting of tumours. (United States)

    Manjappa, Arehalli S; Goel, Peeyush N; Gude, Rajiv P; Ramachandra Murthy, Rayasa S


    Neuropilin-1, a transmembrane receptor entailed in wide range of human tumour cell lines and diverse neoplasms, mediates the effects of VEGF and Semaphorins during the processes of cellular proliferation, survival and migration. In view of this, we had developed and evaluated in vitro and in vivo efficacy of anti-neuropilin-1 immunoliposomes against neuropilin-1 receptor expressing tumours. The PEGylated liposomes loaded with docetaxel were prepared using thin film hydration method. Functionalised PEGylated liposomes were prepared using post-insertion technique. Anti-neuropilin-1 immunoliposomes were prepared by covalently conjugating Fab' fragments of neuropilin-1 antibody to functionalised PEGylated liposomes via thioether linkage. In vivo evaluation of Taxotere and liposomal formulations was performed using intradermal tumour model to demonstrate anti-angiogenic and tumour regression ability. The modified Fab' fragments and immunoliposomes were found to be immunoreactive against A549 cells. Further, docetaxel loaded PEGylated liposomes and PEGylated immunoliposomes demonstrated higher in vitro cytotoxicity than Taxotere formulation at the same drug concentration and exposure time. The live imaging showed distinctive cellular uptake of functional immunoliposomes. Further, significant decrease in micro-blood vessel density and tumour volumes was observed using bio-engineered liposomes. The results clearly highlight the need to seek neuropilin-1 as one of the prime targets in developing an anti-angiogenic therapy.

  2. Feeding ω-3 PUFA enriched rotifers to Galaxias maculatus (Jenyns, 1842 larvae reared at different salinity conditions: effects on growth parameters, survival and fatty acids profile

    Directory of Open Access Journals (Sweden)

    Patricio Dantagnan


    Full Text Available Despite the well known importance of ω-3 polyunsaturated fatty acids (PUFA in marine and freshwater fish larvae, there are few studies on how essential fatty acid requirements and composition on whole body can be altered by changes in water salinity. The present study aimed to determine the effect of salinity on ω-3 PUFA requirements, larval growth survival and fatty acid composition of Galaxias maculatus larvae cultured at two different salinities (0 and 15 g L-1 for 20 days while fed rotifers containing two different levels of ω-3 PUFA (1.87 and 3.16%. The results denoted a marked difference in ω-3 PUFA requirements and in the pattern of fatty acid deposition in the whole body of larvae reared at different salinities, depending of ω-3 PUFA in diets. Thus, to improve growth and survival larvae of G. maculatus reared at 0 g L-1 require higher levels of ω-3 PUFA, principally 18:3 ω-3. Larvae reared at salinities of 15 g L-1 require low levels of ω-3 PUFA for optimal survival, especially 18:3 ω-3. Eicosapentaenoic acid and docosahexaenoic acid content in the whole body of larvae was also affected by water salinity.

  3. Vicilin-derived peptides are transferred from males to females as seminal nuptial gift in the seed-feeding beetle Callosobruchus maculatus. (United States)

    Alexandre, Daniel; Linhares, Ricardo T; Queiroz, Bruna; Fontoura, Luisa; Uchôa, Adriana F; Samuels, Richard I; Macedo, Maria Lígia R; Bezerra, Cezar S; Oliveira, Eliana M; Demartini, Diogo R; Carlini, Célia R; Silva, Carlos P


    The fate of vicilins ingested by Callosobruchus maculatus and the physiological importance of these proteins in larvae and adults have been recently investigated. Vicilins have been demonstrated to be absorbed through the midgut epithelium, circulate in their trimeric form in the haemolymph and are deposited in the fat body. In fat body cells of both sexes, vicilins are partially hydrolyzed and the fragments are eventually deposited in the eggs. Tracking the fate of FITC-labelled vicilins in adult males revealed that the labelled vicilin fragments were also detected in oöcytes and eggs, when the males copulated with non-labelled females. Based on the results presented here, we propose that following absorption, vicilins accumulate in the fat body, where they are partially degraded. These peptides are retained throughout the development of the males and are eventually sequestered by the gonads and passed to the female gonads during copulation. It is possible that accumulation in the eggs is a defensive strategy against pathogen attack, as these peptides are known to have antimicrobial activity. The contribution of vicilin-derived peptides from seminal fluids may be an investment that helps to increase the offspring survival. This study provides additional insights into the possible contributions of males to female fecundity following copulation in C. maculatus.

  4. Identification of Albizia lebbeck seed coat chitin-binding vicilins (7S globulins with high toxicity to the larvae of the bruchid Callosobruchus maculatus

    Directory of Open Access Journals (Sweden)

    A.J. Souza


    Full Text Available Seed coat is a specialized maternal tissue that interfaces the embryo and the external environment during embryogenesis, dormancy and germination. In addition, it is the first defensive barrier against penetration by pathogens and herbivores. Here we show that Albizia lebbeck seed coat dramatically compromises the oviposition, eclosion and development of the bruchid Callosobruchus maculatus. Dietary supplementation of bruchid larvae with A. lebbeck seed coat flour causes severe weight loss and reduces survival. By means of protein purification, mass spectrometry and bioinformatic analyses, we show that chitin-binding vicilins are the main source of A. lebbeck tegumental toxicity to C. maculatus. At concentrations as low as 0.1%, A. lebbeck vicilins reduce larval mass from 8.1 ± 1.7 (mass of control larvae to 1.8 ± 0.5 mg, which corresponds to a decrease of 78%. Seed coat toxicity constitutes an efficient defense mechanism, hindering insect predation and preventing embryo damage. We hypothesize that A. lebbeck vicilins are good candidates for the genetic transformation of crop legumes to enhance resistance to bruchid predation.

  5. Bioefficacy of a long-lasting insecticide impregnated net on blood feeding inhibition of Anopheles maculatus Theobald and Culex quinquefasciatus Say. (United States)

    Beng, Tan Swee; Vythilingam, Indra; Lim, Lee Han


    This study aimed to evaluate the bioefficacy and blood feeding inhibition of mosquitoes under laboratory conditions using the WHO tunnel test method on unwashed and washed long-lasting insecticide impregnated net with extrinsic heat treatment of 30 degrees C followed by 80 degrees C on the same net during washing. PermaNet exhibited fairy high durability to washing (5 washes) and had fairy long-lasting bioefficacy against Anopheles maculatus for blood feeding inhibition on both unwashed (39 months) and washed (26 months) nets. However, Perma-Net exhibited lower bioefficacy against Culex quinquefasciatus. This study also suggested that the application of extrinsic heat treatment of 30 degrees C followed by an increased heat at 80 degrees C on the same net exerted significant differences (p 0.05). An. maculatus exhibited significant differences in resting preference after a successful blood meal, as more blood-fed and live females preferred to rest and stay near the bait in the mouse cage, and more dead and unfed females were found in the outer cage of both the unwashed and washed nets (p < 0.05). Conversely, fully blood-fed and live Cx. quinquefasciatus females did not show any resting preference between the mouse cage and outer cage, but there were more dead and unfed females in the mouse cage of both the unwashed and washed nets (p < 0.05).

  6. Activity of the thyroid and the pituitary gland in the viviparous Cyprinodonts Lebistes reticulatus (Peters), Xiphophorus helleri Heckel and Xiphophorus maculatus Günther during the development of the gonopodium

    NARCIS (Netherlands)

    Stolk, A.


    A description is given of the activity of the thyroid and of the pituitary gland during the development of the gonopodium in the viviparous Cyprinodonts Lebistes reticulatus (Peters), Xiphophorus helleri (Heckel) and Xiphophorus maculatus (Günther). During this process the thyroid gland and the glan

  7. Noteworthy records of Hispines from Belize (Coleoptera: Chrysomelidae)


    Robert F C Naczi; Staines, Charles L.


    Cephaloleia consanguinea Baly, Cephaloleia fulvolimbata Baly, Cephaloleia ruficollis Baly, Chalepus amabilis Baly, Chalepus brevicornis (Baly), Chalepus pici Descarpentries and Villiers, Microrhopala erebus (Newman), Octhispa bimaculata Uhmann, Octotoma championi Baly, Pseudispa tuberculata Staines, Sceloenopla erudita (Baly), Stenispa guatemalensis Uhmann, Sumitrosis gestroi (Weise), and Sumitrosis terminatus (Baly) (Coleoptera: Chrysomelidae: Cassidinae) are new country records of hispine c...

  8. Elmidae Curtis, 1830 (Coleoptera, Polyphaga, Byrrhoidea) of the Neotropical region. (United States)

    Segura, Melissa Ottoboni; Dos Passos, Maria Ines Da Silva; Fonseca-Gessner, Alaíde Aparecida; Froehlich, Claudio Gilberto


    A bibliography of significant taxonomic papers on Elmidae (Coleoptera, Polyphaga, Byrrhoidea), and a checklist of valid species and subspecies and their geographic distributions are provided. Forty-seven genera and 430 valid species are included. Maps show the geographic distribution of the genera by country.

  9. Diet based fitness variability of Coccinella novemnotata (Coleoptera: Coccinellidae) (United States)

    Coccinella novemnotata (Herbst) is a species of North American native lady beetle (Coleoptera: Coccinellidae) that has come under great ecological duress over the past 30 years and is experiencing a significant decline throughout its native range. This species once was widely distributed across mos...

  10. Temperature-dependent development of Xyleborus glabratus (Coleoptera: Curculionidae: Scolytinae) (United States)

    Redbay ambrosia beetle, Xyleborus glabratus Eichhoff (Coleoptera: Curculionidae: Scolytinae) is a nonnative pest that vectors the pathogenic fungus Raffaelea lauricola, which causes laurel wilt in trees of the family Lauraceae. Laurel wilt is present in the commercial growing areas of avocado (Perse...

  11. Morphometric analysis of instar variation in Tenebrio molitor (Coleoptera: Tenebrionidae) (United States)

    Measurements of head capsule, mandible, metanotum, and body weight were done on larvae of Tenebrio molitor L. (Coleoptera: Tenebrionide) from the second to the last instar. Instar number varied from 14 to 18, but 15 or 16 instars were the most common. The value of dimensional measurements was evalua...

  12. A coffee berry borer (Coleoptera: Curculionidae: Scolytinae) bibliography (United States)

    One hundred years ago, one of the most significant biological invasions of an agricultural insect pest in the Americas was initiated. Endemic to Africa, the coffee berry borer (Hypothenemus hampei; Coleoptera: Curculionidae) was accidentally introduced to Brazil in 1913 and years later invaded coffe...

  13. Complexes of neutralizing and non-neutralizing affinity matured Fabs with a mimetic of the internal trimeric coiled-coil of HIV-1 gp41.

    Directory of Open Access Journals (Sweden)

    Elena Gustchina

    Full Text Available A series of mini-antibodies (monovalent and bivalent Fabs targeting the conserved internal trimeric coiled-coil of the N-heptad repeat (N-HR of HIV-1 gp41 has been previously constructed and reported. Crystal structures of two closely related monovalent Fabs, one (Fab 8066 broadly neutralizing across a wide panel of HIV-1 subtype B and C viruses, and the other (Fab 8062 non-neutralizing, representing the extremes of this series, were previously solved as complexes with 5-Helix, a gp41 pre-hairpin intermediate mimetic. Binding of these Fabs to covalently stabilized chimeric trimers of N-peptides of HIV-1 gp41 (named (CCIZN363 or 3-H has now been investigated using X-ray crystallography, cryo-electron microscopy, and a variety of biophysical methods. Crystal structures of the complexes between 3-H and Fab 8066 and Fab 8062 were determined at 2.8 and 3.0 Å resolution, respectively. Although the structures of the complexes with the neutralizing Fab 8066 and its non-neutralizing counterpart Fab 8062 were generally similar, small differences between them could be correlated with the biological properties of these antibodies. The conformations of the corresponding CDRs of each antibody in the complexes with 3-H and 5-Helix are very similar. The adaptation to a different target upon complex formation is predominantly achieved by changes in the structure of the trimer of N-HR helices, as well as by adjustment of the orientation of the Fab molecule relative to the N-HR in the complex, via rigid-body movement. The structural data presented here indicate that binding of three Fabs 8062 with high affinity requires more significant changes in the structure of the N-HR trimer compared to binding of Fab 8066. A comparative analysis of the structures of Fabs complexed to different gp41 intermediate mimetics allows further evaluation of biological relevance for generation of neutralizing antibodies, as well as provides novel structural insights into immunogen

  14. Binding of Shewanella FadR to the fabA fatty acid biosynthetic gene: implications for contraction of the fad regulon. (United States)

    Zhang, Huimin; Zheng, Beiwen; Gao, Rongsui; Feng, Youjun


    The Escherichia coli fadR protein product, a paradigm/prototypical FadR regulator, positively regulates fabA and fabB, the two critical genes for unsaturated fatty acid (UFA) biosynthesis. However the scenario in the other Ɣ-proteobacteria, such as Shewanella with the marine origin, is unusual in that Rodionov and coworkers predicted that only fabA (not fabB) has a binding site for FadR protein. It raised the possibility of fad regulon contraction. Here we report that this is the case. Sequence alignment of the FadR homologs revealed that the N-terminal DNA-binding domain exhibited remarkable similarity, whereas the ligand-accepting motif at C-terminus is relatively-less conserved. The FadR homologue of S. oneidensis (referred to FadR_she) was over-expressed and purified to homogeneity. Integrative evidence obtained by FPLC (fast protein liquid chromatography) and chemical cross-linking analyses elucidated that FadR_she protein can dimerize in solution, whose identity was determined by MALDI-TOF-MS. In vitro data from electrophoretic mobility shift assays suggested that FadR_she is almost functionally-exchangeable/equivalent to E. coli FadR (FadR_ec) in the ability of binding the E. coli fabA (and fabB) promoters. In an agreement with that of E. coli fabA, S. oneidensis fabA promoter bound both FadR_she and FadR_ec, and was disassociated specifically with the FadR regulatory protein upon the addition of long-chain acyl-CoA thioesters. To monitor in vivo effect exerted by FadR on Shewanella fabA expression, the native promoter of S. oneidensis fabA was fused to a LacZ reporter gene to engineer a chromosome fabA-lacZ transcriptional fusion in E. coli. As anticipated, the removal of fadR gene gave about 2-fold decrement of Shewanella fabA expression by β-gal activity, which is almost identical to the inhibitory level by the addition of oleate. Therefore, we concluded that fabA is contracted to be the only one member of fad regulon in the context of fatty acid

  15. Construction of a human recombinant polyclonal Fab fragment antibody library using peripheral blood lymphocytes of snake bitten victims

    Directory of Open Access Journals (Sweden)

    Motedayen, M.H.


    Full Text Available Human snake bitten poisoning is a serious threat in many tropical and subtropical countries such as Iran. The best acceptable treatment of envenomated humans is antivenoms; however they have a series of economic and medical problems and need more improvements. In this study a combinatorial human immunoglobulin gene library against some of Iranian snakes venoms was constructed. Total RNA prepared from peripheral blood lymphocytes of two recovered snake victims. RT-PCR was used for cDNA synthesis and amplification of the heavy (Fd segment and kappa light chains of IgG antibody. After digestion of the heavy chain with SpeI and XhoI and light chain with XbaI and SacI enzymes, inserted successively into the cloning vector pComb3x, and then recombinant vector transformed to TG1 bacteria to construct the Fab library. For determination insertion rate of Fab segment into cloning vector, plasmids of 12 clones of library were extracted and digested with SfiI. Length of amplified Fd and κ chains, were 650 - 750 bp. Primary library size was determined to contain 4.9×105 members out of which half of them contained the same size of Fab fragment. This result is comparable to some researchers and shows that this method could be appropriate tool for the production of human polyclonal Fab fragment antibodies for management of poisonous snake bitted victims.

  16. [Molecular dynamics of immune complex of photoadduct-containing DNA with Fab-Anti-DNA antibody fragment]. (United States)

    Akberova, N I; Zhmurov, A A; Nevzorova, T A; Litvinov, R I


    Antibodies to DNA play an important role in the pathogenesis of autoimmune diseases. The elucidation of structural mechanisms of both the antigen recognition and the interaction of anti-DNA antibodies with DNA will help to understand the role of DNA-containing immune complexes in various pathologies and can provide a basis for new treatment modalities. Moreover, the DNA-antibody complex is an analog of specific intracellular DNA-protein interactions. In this work, we used in silico molecular dynamic simulations of bimolecular complexes of the dsDNA segment containing the Fab fragment of an anti-DNA antibody to obtain the detailed thermodynamic and structural characteristics of dynamic intermolecular interactions. Using computationally modified crystal structure of the Fab-DNA complex (PDB ID: 3VW3), we studied the equilibrium molecular dynamics of the 64M-5 antibody Fab fragment associated with the dsDNA fragment containing the thymine dimer, the product of DNA photodamage. Amino acid residues that constitute paratopes and the complementary nucleotide epitopes for the Fab-DNA construct were identified. Stacking and electrostatic interactions were found to play the main role in mediating the most specific antibody-dsDNA contacts, while hydrogen bonds were less significant. These findings may shed light on the formation and properties of pathogenic anti-DNA antibodies in autoimmune diseases, such as systemic lupus erythematosus associated with skin photosensitivity and DNA photodamage.

  17. Generation of a haptoglobin-hemoglobin complex-specific Fab antibody blocking the binding of the complex to CD163

    DEFF Research Database (Denmark)

    Horn, Ivo R; Nielsen, Marianne Jensby; Madsen, Mette


    During intravascular hemolysis hemoglobin (Hb) binds to haptoglobin (Hp) leading to endocytosis of the complex by the macrophage receptor, CD163. In the present study, we used a phage-display Fab antibody strategy to explore if the complex formation between Hp and Hb leads to exposure of antigenic...

  18. Predation by Flat Bark Beetles (Coleoptera: Silvanidae and Laemophloeidae) on Coffee Berry Borer (Coleoptera: Curculionidae) in Hawaii coffee (United States)

    Coffee berry borer(CBB), Hypothenemus hampei, is a serious pest of coffee worldwide and a new invasive pest in Hawaii. Adult flat bark beetles, mainly Leptophloeus sp.(75%) and Cathartus quadricollis(21%) (Coleoptera: Laemophloeidae and Silvanidae, respectively), were found feeding in CBB-infested c...

  19. FabSim: Facilitating computational research through automation on large-scale and distributed e-infrastructures (United States)

    Groen, Derek; Bhati, Agastya P.; Suter, James; Hetherington, James; Zasada, Stefan J.; Coveney, Peter V.


    We present FabSim, a toolkit developed to simplify a range of computational tasks for researchers in diverse disciplines. FabSim is flexible, adaptable, and allows users to perform a wide range of tasks with ease. It also provides a systematic way to automate the use of resources, including HPC and distributed machines, and to make tasks easier to repeat by recording contextual information. To demonstrate this, we present three use cases where FabSim has enhanced our research productivity. These include simulating cerebrovascular bloodflow, modelling clay-polymer nanocomposites across multiple scales, and calculating ligand-protein binding affinities. Catalogue identifier: AFAO_v1_0 Program summary URL: Program obtainable from: CPC Programme Library, Queen's University, Belfast, N. Ireland Licensing provisions: BSD 3-Clause No. of lines in distributed program, including test data, etc.: 268282 No. of bytes in distributed program, including test data, etc.: 2791792 Distribution format: tar.gz Programming language: Python. Computer: PC or Mac. Operating system: Unix, OSX. RAM: 1 Gbytes Classification: 3, 4, 6.5. External routines: NumPy, SciPy, Fabric (1.5 or newer), PyYaml Nature of problem: Running advanced computations using remote resources is an activity that requires considerable time and human attention. These activities, such as organizing data, configuring software and setting up individual runs often vary slightly each time they are performed. To lighten this burden, we required an approach that introduced little burden of its own to set up and adapt, beyond which very substantial productivity ensues. Solution method: We present a toolkit which helps to simplify and automate the activities which surround computational science research. FabSim is aimed squarely at the experienced computational scientist, who can use the command line interface and a system of modifiable content to quickly automate sets of

  20. The change of the scFv into the Fab format improves the stability and in vivo toxin neutralization capacity of recombinant antibodies. (United States)

    Quintero-Hernández, Veronica; Juárez-González, Victor R; Ortíz-León, Mauricio; Sánchez, Rosalba; Possani, Lourival D; Becerril, Baltazar


    The antigen-binding fragment (Fab) has been considered a more functionally stable version of recombinant antibodies than single chain antibody fragments (scFvs), however this intuitive consideration has not been sufficiently proven in vivo. This communication shows that three out of four specific scFvs against a scorpion toxin, with different affinities and stabilities, become neutralizing in vivo when expressed as Fabs, despite the fact that they are not neutralizing in the scFv format. A scFv fragment previously obtained from a neutralizing mouse antibody (BCF2) was used to produce three derived scFvs by directed evolution. Only one of them was neutralizing, however when expressed as Fab, all of them became neutralizing fragments in vivo. The initial scFvBCF2 (earlier used for directed evolution) was not neutralizing in the scFv format. After expressing it as Fab did not become a neutralizing fragment, but did reduce the intoxication symptoms of experimental mice. The stability of the four Fabs derived from their respective scFvs was improved when tested in the presence of guanidinium chloride. The in vitro stability of the Fab format has been shown earlier, but the physiological consequences of this stability are shown in this communication. The present results indicate that improved functional stability conferred by the Fab format can replace additional maturation steps, when the affinity and stability are close to the minimum necessary to be neutralizing.

  1. Application of {sup 99m}Tc-labeled chimeric Fab fragments of monoclonal antibody A7 for radioimmunoscintigraphy of pancreatic cancer

    Energy Technology Data Exchange (ETDEWEB)

    Matsumura, Hiroomi [Kyoto Prefectural Univ. of Medicine (Japan)


    Pancreatic cancer is one of the most lethal diseases and its prognosis is still poor. To improve the survival rate, it is essential to develop new technologies for early and definitive diagnosis. In this study, chimeric Fab fragments of monoclonal antibody A7 were successfully radio-labeled with {sup 99m}Tc, preventing depression of the antigen-binding activity. {sup 99m}Tc-labeled monoclonal antibody A7, {sup 99m}Tc-labeled chimeric Fab fragments of monoclonal antibody A7, {sup 99m}Tc-labeled normal mouse IgG and {sup 99m}Tc-labeled Fab fragments of normal mouse IgG were injected intravenously into nude mice bearing human pancreatic cancer xenografts and the radioactivity was subsequently measured. The tumor accumulation was significantly higher with labeled monoclonal antibody A7 than with normal mouse IgG, and higher with chimeric Fab fragments of monoclonal antibody A7 than with Fab fragments of normal mouse IgG. The tumor/blood ratio of radioactivity increased rapidly over time with chimeric Fab fragments of monoclonal antibody A7. These results suggest that chimeric Fab fragments of monoclonal antibody A7 may be useful for diagnosing pancreatic cancer by means of radioimmunoscintigraphy. (author)

  2. Fab'-bearing siRNA TNFα-loaded nanoparticles targeted to colonic macrophages offer an effective therapy for experimental colitis. (United States)

    Laroui, Hamed; Viennois, Emilie; Xiao, Bo; Canup, Brandon S B; Geem, Duke; Denning, Timothy L; Merlin, Didier


    Patients suffering from inflammatory bowel disease (IBD) are currently treated by systemic drugs that can have significant side effects. Thus, it would be highly desirable to target TNFα siRNA (a therapeutic molecule) to the inflamed tissue. Here, we demonstrate that TNFα siRNA can be efficiently loaded into nanoparticles (NPs) made of poly (lactic acid) poly (ethylene glycol) block copolymer (PLA-PEG), and that grafting of the Fab' portion of the F4/80 Ab (Fab'-bearing) onto the NP surface via maleimide/thiol group-mediated covalent bonding improves the macrophage (MP)-targeting kinetics of the NPs to RAW264.7 cells in vitro. Direct binding was shown between MPs and the Fab'-bearing NPs. Next, we orally administered hydrogel (chitosan/alginate)-encapsulated Fab'-bearing TNFα-siRNA-loaded NPs to 3% dextran sodium sulfate (DSS)-treated mice and investigated the therapeutic effect on colitis. In vivo, the release of TNFα-siRNA-loaded NPs into the mouse colon attenuated colitis more efficiently when the NPs were covered with Fab'-bearing, compared to uncovered NPs. All DSS-induced parameters of colonic inflammation (e.g., weight loss, myeloperoxidase activity, and Iκbα accumulation) were more attenuated Fab'-bearing NPs loaded with TNFα siRNA than without the Fab'-bearing. Grafting the Fab'-bearing onto the NPs improved the kinetics of endocytosis as well as the MP-targeting ability, as indicated by flow cytometry. Collectively, our results show that Fab'-bearing PLA-PEG NPs are powerful and efficient nanosized tools for delivering siRNAs into colonic macrophages.

  3. Building blocks X-FAB SOI 0.18 μm (United States)

    Cizel, J.-B.; Ahmad, S.; Callier, S.; Cornat, R.; Dulucq, F.; Fleury, J.; Martin-Chassard, G.; Raux, L.; de La Taille, C.; Thienpont, D.


    This work has been done in order to study a new technology provided by X-FAB named xt018. It is an SOI (Silicon On Insulator) technology with a minimal gate length of 180 nm. Building blocks have been done to test the advantages and drawbacks of this technology compared to the one currently used (AMS SiGe 0.35 μm). These building blocks have been designed to fit in an existing experience housed by the CALICE collaboration: the read-out chip for the Electromagnetic CALorimeter (ECAL) of the foreseen International Linear Collider (ILC). Performances will be compared to those of the SKIROC2 chip designed by the OMEGA laboratory, trying to fit the same requirements. The chip is being manufactured and will be back for measurements in December, the displayed results are only simulation results and thus the conclusions concerning the performances of these building blocks are subject to change.

  4. Photomask film degradation effects in the wafer fab: how to detect and monitor over time (United States)

    Whittey, John; Hess, Carl; Garcia, Edgardo; Wagner, Mark; Duffy, Brian


    As a result of repeated cleanings and exposure effects such as chrome migration or MoSi oxidation some photomasks in the semiconductor fabs exhibit changes in critical dimension uniformity (CDU) over time. Detecting these effects in a timely manner allows for better risk management and process control in manufacturing. By monitoring changes in film reflectance intensity due to the various degradation mechanisms it is possible to predict when they may begin to influence across chip line width variations (ACLV). By accurately predicting the magnitude of these changes it is possible for semiconductor manufacturers to replace the photomasks before they have an impact on yields. This paper looks at possible causes of CDU variations on reticles during use and how this information might be used to improve or monitor reticle CDU changes over time.


    Institute of Scientific and Technical Information of China (English)

    刘宁; 罗德元; 等


    An antigastric cancer monoclonal antibody,3H11,and its Fab fragment,were labeled using p-(211At)-astatobenzoic acid(pAtBA) intermediate,in the yields of more than 30%.The results of in vitro experiments show that 211 At-3H11 and 211At-3H11 Fab are stable,and have specific immunoreactivities and cytotoxic effects to human gastric cancer cell M85,The cytotoxic effects are dependent on the concentration of 211At-3H11 or 211 At-3H11 Fab and obviously stronger than that of Na211At.

  6. Efecto de la temperatura, humedad y dieta sobre el desarrollo de Gnathocerus cornutus Fab. (Coleopera: Tenebrionidae

    Directory of Open Access Journals (Sweden)

    Núñez Fernando


    Full Text Available The influence of temperature, relative humídíty and diet composítíon in the duration and velocity of the development cicle of Gnathoccnis comutus Fab., was studied. Each of two diets (Aand Bl was tried in presence of 5 different combinations of temperature and humídíty. Optimal conditions fOI development were found to be: 28°C and 85-90% R.H.. in which the complete cycle lasts 54 days. the larval stage is the critical one in the development in which a htgh variation among individuals in both, velocity and span of development, are notorious. The cycle duration is virtually the same for both diets, but in the diet A. a better adaptability, expressed as a proportion of descendents, was observed. Life tables and surviving curves were made.
    Se estudió la influencia de la temperatura. la humedad relativa y la composición de la dieta sobre la duración del ciclo de desarrollo y la velocidad del mismo en Gnathocerus comutus Fab, Para el efecto. cada una de las dos dietas (A y B] se ensayó en presencia de 5 combinaciones distintas de temperatura y humedad. Se encontró que las condiciones óptimas para el desarrollo. son: 28°C y 85-90% H.R.. en las cuales. el ciclo completo se desenvuelve en 54 días. El estado crítico en cuanto al desarrollo. es el larval en el cual es notoria una alta variación entre los individuos en velocidad y tiempo de desarrollo. La duración del ciclo es virtualmente la misma en ambas dietas. pero en la A se observa una mejor adaptabilidad expresada en proporctón de descendientes. Se construyeron tablas de vida y curvas de supervivencia.

  7. The Intrinsic Dynamics and Unfolding Process of an Antibody Fab Fragment Revealed by Elastic Network Model

    Directory of Open Access Journals (Sweden)

    Ji-Guo Su


    Full Text Available Antibodies have been increasingly used as pharmaceuticals in clinical treatment. Thermal stability and unfolding process are important properties that must be considered in antibody design. In this paper, the structure-encoded dynamical properties and the unfolding process of the Fab fragment of the phosphocholine-binding antibody McPC603 are investigated by use of the normal mode analysis of Gaussian network model (GNM. Firstly, the temperature factors for the residues of the protein were calculated with GNM and then compared with the experimental measurements. A good result was obtained, which provides the validity for the use of GNM to study the dynamical properties of the protein. Then, with this approach, the mean-square fluctuation (MSF of the residues, as well as the MSF in the internal distance (MSFID between all pairwise residues, was calculated to investigate the mobility and flexibility of the protein, respectively. It is found that the mobility and flexibility of the constant regions are higher than those of the variable regions, and the six complementarity-determining regions (CDRs in the variable regions also exhibit relative large mobility and flexibility. The large amplitude motions of the CDRs are considered to be associated with the immune function of the antibody. In addition, the unfolding process of the protein was simulated by iterative use of the GNM. In our method, only the topology of protein native structure is taken into account, and the protein unfolding process is simulated through breaking the native contacts one by one according to the MSFID values between the residues. It is found that the flexible regions tend to unfold earlier. The sequence of the unfolding events obtained by our method is consistent with the hydrogen-deuterium exchange experimental results. Our studies imply that the unfolding behavior of the Fab fragment of antibody McPc603 is largely determined by the intrinsic dynamics of the protein.

  8. Generation, characterization, and docking studies of DNA-hydrolyzing recombinant F(ab) antibodies. (United States)

    Zein, Haggag S; El-Sehemy, Ahmed A; Fares, Mohamed O; ElHefnawi, Mahmoud; da Silva, Jaime A Teixeira; Miyatake, Kazutaka


    Previously we established a series of catalytic antibodies (catAbs) capable of hydrolyzing DNA prepared by hybridoma technology. A group of these catAbs exhibited high reactivity and substrate specificity. To determine the molecular basis for these catAbs, we cloned, sequenced, and expressed the variable regions of this group of antibodies as functional F(ab) fragments. The nucleotide and deduced amino acid sequences of the expressed light chain (Vκ) germline gene assignments confidently belonged to germline family Vκ1A, gene bb1.1 and GenBank accession number EF672207 while heavy chain variable region V(H) genes belonged to V(H) 1/V(H) J558, gene V130.3 and GenBank accession number EF672221. A well-established expression system based on the pARA7 vector was examined for its ability to produce catalytically active antibodies. Recombinant F(ab) (rF(ab) ) fragments were purified and their hydrolyzing activity was analyzed against supercoiled pUC19 plasmid DNA (scDNA). The study of rF(ab) provides important information about the potential catalytic activities of antibodies whose structure allows us to understand their basis of catalysis. Molecular surface analysis and docking studies were performed on the molecular interactions between the antibodies and poly(dA9), poly(dG9), poly(dT9), and poly(dC9) oligomers. Surface analysis identified the important sequence motifs at the binding sites, and different effects exerted by arginine and tyrosine residues at different positions in the light and heavy chains. This study demonstrates the potential usefulness of the protein DNA surrogate in the investigation of the origin of anti-DNA antibodies. These studies may define important features of DNA catAbs.

  9. Ergasilus chelangulatus sp. nov. (Copepoda: Ergasilidae) a branchial parasite of the freshwater catfish, Pimelodus maculatus from the upper São Francisco River, Brazil Ergasilus chelangulatus sp. nov. (Copepoda: Ergasilidae) parasito das brânquias do "mandi", Pimelodus maculatus do alto rio São Francisco, Brasil


    Vernon E. Thatcher; Brasil-Sato,Marilia C.


    Ergasilus chelangulatus sp. nov. is described from the freshwater fish, Pimelodus maculatus Lacépède, 1803 (Siluriformes: Pimelodidae) of the upper São Francisco River, Minas Gerais State, Brazil. The new species is based on female specimens and the male is unknown. This species has a serrate seta on exopod one and a two-segmented first endopod, as do most Amazonian species of this genus. The body is elongate and produced anteriorly. The antennae are elongate and have prominent sensilla on se...

  10. Two new species of Scymnini (Coleoptera: Coccinellidae) from Karnataka, India. (United States)

    Poorani, J


    The Scymnini (Coleoptera: Coccinellidae) of the Indian region is rich and highly speciose, with nearly 90 described species and scores of undescribed species (Poorani 2002). There is a dire need to systematically revise the genera and species of this tribe from the Indian region. Due to paucity of representative collections covering the entire region and lack of access to types, it is difficult to identify most of the Scymnini of the Indian region to species. As a result, many economically important species remain poorly characterized, or worse, unnamed. Two economically important and unique species of Scymnini (Coccinellidae) belonging to Horniolus Weise (1900) and Scymnus (Pullus) Mulsant (1846) from the Southern Indian state of Karnataka that have remained unnamed for long are treated in this paper. These species are externally similar to other known species and often misidentified. Horniolussororius sp. n. and Scymnus (Pullus) rajeshwariae sp. n. (Coleoptera: Coccinellidae) are described here and illustrated with notes on their biology and related species.

  11. Structure-based design,synthesis of novel inhibitors of Mycobacterium tuberculosis FabH as potential anti-tuberculosis agents

    Institute of Scientific and Technical Information of China (English)

    Xue Hui Zhang; Hong Yu; Wu Zhong; Li Li Wang; Song Li


    Mycobacterium tuberculosis FabH,an essential enzyme in mycolic acids biosynthetic pathway,is an attractive target for novel anti-tuberculosis agents.Structure-based design,synthesis of novel inhibitors of mrFabH was reported in this paper.A novel scaffold structure was designed,and 12 candidate compounds that displayed favorable binding with the active site were identified and synthesized.

  12. Allergen-specific regulation of allergic rhinitis in mice by intranasal exposure to IgG1 monoclonal antibody Fab fragments against pathogenic allergen. (United States)

    Matsuoka, Daiko; Mizutani, Nobuaki; Sae-Wong, Chutha; Yoshino, Shin


    Fab fragments (Fabs) have the ability to bind to specific antigens but lack the Fc portion for binding to receptors on immune and inflammatory cells that play a critical role in allergic diseases. In the present study, we investigated whether Fabs of an allergen-specific IgG1 monoclonal antibody (mAb) inhibited allergic rhinitis in mice. BALB/c mice sensitized by intraperitoneal injections of ovalbumin (OVA) plus alum on days 0 and 14 were intranasally challenged with OVA on days 28-30, and 35. Fabs prepared by the digestion of an anti-OVA IgG1 mAb (O1-10) with papain were also intranasally administered 15min before each OVA challenge. The results showed that treatment with O1-10 Fabs significantly suppressed the sneezing frequency, associated with decrease of OVA-specific IgE in the serum and infiltration by mast cells in the nasal mucosa seen following the fourth antigenic challenge; additionally, the level of mouse mast cell protease-1, a marker of mast cell activation, in serum was decreased. Furthermore, infiltration of eosinophils and goblet cell hyperplasia in the nasal mucosa at the fourth challenge were inhibited by treatment with O1-10 Fabs. In conclusion, these results suggest that intranasal exposure to Fabs of a pathogenic antigen-specific IgG1 mAb may be effective in regulating allergic rhinitis through allergen capture by Fabs in the nasal mucosa before the interaction of the intact antibody and allergen.

  13. Immunoglobulin G (IgG) Fab glycosylation analysis using a new mass spectrometric high-throughput profiling method reveals pregnancy-associated changes. (United States)

    Bondt, Albert; Rombouts, Yoann; Selman, Maurice H J; Hensbergen, Paul J; Reiding, Karli R; Hazes, Johanna M W; Dolhain, Radboud J E M; Wuhrer, Manfred


    The N-linked glycosylation of the constant fragment (Fc) of immunoglobulin G has been shown to change during pathological and physiological events and to strongly influence antibody inflammatory properties. In contrast, little is known about Fab-linked N-glycosylation, carried by ∼ 20% of IgG. Here we present a high-throughput workflow to analyze Fab and Fc glycosylation of polyclonal IgG purified from 5 μl of serum. We were able to detect and quantify 37 different N-glycans by means of MALDI-TOF-MS analysis in reflectron positive mode using a novel linkage-specific derivatization of sialic acid. This method was applied to 174 samples of a pregnancy cohort to reveal Fab glycosylation features and their change with pregnancy. Data analysis revealed marked differences between Fab and Fc glycosylation, especially in the levels of galactosylation and sialylation, incidence of bisecting GlcNAc, and presence of high mannose structures, which were all higher in the Fab portion than the Fc, whereas Fc showed higher levels of fucosylation. Additionally, we observed several changes during pregnancy and after delivery. Fab N-glycan sialylation was increased and bisection was decreased relative to postpartum time points, and nearly complete galactosylation of Fab glycans was observed throughout. Fc glycosylation changes were similar to results described before, with increased galactosylation and sialylation and decreased bisection during pregnancy. We expect that the parallel analysis of IgG Fab and Fc, as set up in this paper, will be important for unraveling roles of these glycans in (auto)immunity, which may be mediated via recognition by human lectins or modulation of antigen binding.

  14. Active site modification of the β-ketoacyl-ACP synthase FabF3 of Streptomyces coelicolor affects the fatty acid chain length of the CDA lipopeptides. (United States)

    Lewis, Richard A; Nunns, Laura; Thirlway, Jenny; Carroll, Kathleen; Smith, Colin P; Micklefield, Jason


    Using site directed mutagenesis we altered an active site residue (Phe107) of the enzyme encoded by fabF3 (SCO3248) in the Streptomyces coelicolor gene cluster required for biosynthesis of the calcium dependent antibiotics (CDAs), successfully generating two novel CDA derivatives comprising truncated (C4) lipid side chains and confirming that fabF3 encodes a KAS-II homologue that is involved in determining CDA fatty acid chain length.

  15. A checklist of stag beetles (Coleoptera: Scarabaeoidea: Lucanidae) from Iran. (United States)

    Bartolozzi, Luca; Ghahari, Hassan; Sprecher-Uebersax, Eva; Zilioli, Michele


    An updated checklist of the Lucanidae (Coleoptera) from Iran is given. New locality records are listed and some dubious distributional records are discussed. Dorcus vavrai Nonfried, 1905 is placed in synonymy with Dorcus peyronis Reiche and Saulcy, 1856 (new synonymy) The female of Lucanus xerxes Král, 2004 is described. A key for the identification of the Iranian stag beetle species is also provided and all the species are figured.

  16. Invasions by ladybugs, ladybirds, and other predatory Coleoptera


    Evans, Edward W.; Soares, António O.; Yasuda, Hironori


    Copyright © International Organization for Biological Control (IOBC) 2011. Species of predatory Coleoptera have become abundant in new geographic regions recently, raising concerns for invaded ecosystems. We address this topic by focusing on invasive alien ladybird beetles (Coccinellidae; known also as ladybugs). Humans appear directly or indirectly responsible for all or most ladybird invasions. Factors hypothesized to have promoted ladybird invasions include genetic diversity (e.g., for ...

  17. Studies of nontarget-mediated distribution of human full-length IgG1 antibody and its FAb fragment in cardiovascular and metabolic-related tissues. (United States)

    Davidsson, Pia; Söderling, Ann-Sofi; Svensson, Lena; Ahnmark, Andrea; Flodin, Christine; Wanag, Ewa; Screpanti-Sundqvist, Valentina; Gennemark, Peter


    Tissue distribution and pharmacokinetics (PK) of full-length nontargeted antibody and its antigen-binding fragment (FAb) were evaluated for a range of tissues primarily of interest for cardiovascular and metabolic diseases. Mice were intravenously injected with a dose of 10 mg/kg of either human IgG1or its FAb fragment; perfused tissues were collected at a range of time points over 3 weeks for the human IgG1 antibody and 1 week for the human FAb antibody. Tissues were homogenized and antibody concentrations were measured by specific immunoassays on the Gyros system. Exposure in terms of maximum concentration (Cmax ) and area under the curve was assessed for all nine tissues. Tissue exposure of full-length antibody relative to plasma exposure was found to be between 1% and 10%, except for brain (0.2%). Relative concentrations of FAb antibody were the same, except for kidney tissue, where the antibody concentration was found to be ten times higher than in plasma. However, the absolute tissue uptake of full-length IgG was significantly higher than the absolute tissue uptake of the FAb antibody. This study provides a reference PK state for full-length whole and FAb antibodies in tissues related to cardiovascular and metabolic diseases that do not include antigen or antibody binding. © 2015 Wiley Periodicals, Inc. and the American Pharmacists Association.

  18. Transcription of the Escherichia coli fatty acid synthesis operon fabHDG is directly activated by FadR and inhibited by ppGpp. (United States)

    My, Laetitia; Rekoske, Brian; Lemke, Justin J; Viala, Julie P; Gourse, Richard L; Bouveret, Emmanuelle


    In Escherichia coli, FadR and FabR are transcriptional regulators that control the expression of fatty acid degradation and unsaturated fatty acid synthesis genes, depending on the availability of fatty acids. In this report, we focus on the dual transcriptional regulator FadR. In the absence of fatty acids, FadR represses the transcription of fad genes required for fatty acid degradation. However, FadR is also an activator, stimulating transcription of the products of the fabA and fabB genes responsible for unsaturated fatty acid synthesis. In this study, we show that FadR directly activates another fatty acid synthesis promoter, PfabH, which transcribes the fabHDG operon, indicating that FadR is a global regulator of both fatty acid degradation and fatty acid synthesis. We also demonstrate that ppGpp and its cofactor DksA, known primarily for their role in regulation of the synthesis of the translational machinery, directly inhibit transcription from the fabH promoter. ppGpp also inhibits the fadR promoter, thereby reducing transcription activation of fabH by FadR indirectly. Our study shows that both ppGpp and FadR have direct roles in the control of fatty acid promoters, linking expression in response to both translation activity and fatty acid availability.

  19. Nocturnal Migration of Coleoptera: Carabidae in North China

    Institute of Scientific and Technical Information of China (English)


    Vertical-looking radar (VLR) has allowed long-term automatic monitoring of the altitudinal and temporal dynamics of high-flying insect populations. To investigate whether ground beetle, insect of Coleoptera, was capable of migrating and its migration pattern by taking advantages of capability of the VLR for long-term real-time automatic monitoring, the migration of Coleoptera ground beetle was investigated by setting up radar observation points, making long-term observation using the VLR and related supplementary equipment, and analyzing low altitude air current and large area circulating current in combination with the meteorological data. Information obtained in 2005 and 2006 showed that the seasonal activities of ground beetles traps of trap lamps were mainly from late June to late August, peak period was mainly in August, seasonal traps of high-altitude lamps and ground lamps were featured by sudden increase and sudden decrease; in peak period, the height of radar echo point could be as high as 600 m, while it was mainly below the height of 450 m; night activities mainly occurred from 20:00 to 22:00, in very few nights, radar echo could last until about 04:00, changes in numbers of ground beetles within the searchlights were consistent with radar echo intensity; ground beetle images were successfully trapped in the sweep nets carried by captive balloons at the height of 200 m. Some species of Carabidaes had some degrees of migration, thus providing the foundation for investigating the migration of Coleoptera insects.

  20. Resistance Mechanisms and the Future of Bacterial Enoyl-Acyl Carrier Protein Reductase (FabI) Antibiotics. (United States)

    Yao, Jiangwei; Rock, Charles O


    Missense mutations leading to clinical antibiotic resistance are a liability of single-target inhibitors. The enoyl-acyl carrier protein reductase (FabI) inhibitors have one intracellular protein target and drug resistance is increased by the acquisition of single-base-pair mutations that alter drug binding. The spectrum of resistance mechanisms to FabI inhibitors suggests criteria that should be considered during the development of single-target antibiotics that would minimize the impact of missense mutations on their clinical usefulness. These criteria include high-affinity, fast on/off kinetics, few drug contacts with residue side chains, and no toxicity. These stringent criteria are achievable by structure-guided design, but this approach will only yield pathogen-specific drugs. Single-step acquisition of resistance may limit the clinical application of broad-spectrum, single-target antibiotics, but appropriately designed pathogen-specific antibiotics have the potential to overcome this liability.

  1. Construction of human Fab library and screening of a single-domain antibody of amyloid-beta 42 oligomers

    Institute of Scientific and Technical Information of China (English)

    Zuanning Yuan; Minge Du; Yiwen Chen; Fei Dou


    Screening humanized antibodies from a human Fab phage display library is an effective and quick method to obtain beta-amyloid oligomers. Thus, the present study prepared amyloid-beta 42 oli-gomers and constructed a naïve human Fab phage display library based on blood samples from six healthy people. After three rounds of biopanning in vitro, a human single-domain antibody that spe-cifical y recognized amyloid-beta 42 oligomers was identified. Western blot and enzyme-linked immunosorbent assay demonstrated this antibody bound specifical y to human amyloid-beta 42 te-tramer and nonamer, but not the monomer or high molecular weight oligomers. This study suc-cessful y constructed a human phage display library and screened a single-domain antibody that specifical y recognized amyloid-beta 42 oligomers.

  2. Structure of Mycobacterium tuberculosis mtFabD, a malonyl-CoA:acyl carrier protein transacylase (MCAT). (United States)

    Ghadbane, Hemza; Brown, Alistair K; Kremer, Laurent; Besra, Gurdyal S; Fütterer, Klaus


    Mycobacteria display a unique and unusual cell-wall architecture, central to which is the membrane-proximal mycolyl-arabinogalactan-peptidoglycan core (mAGP). The biosynthesis of mycolic acids, which form the outermost layer of the mAGP core, involves malonyl-CoA:acyl carrier protein transacylase (MCAT). This essential enzyme catalyses the transfer of malonyl from coenzyme A to acyl carrier protein AcpM, thus feeding these two-carbon units into the chain-elongation cycle of the type II fatty-acid synthase. The crystal structure of M. tuberculosis mtFabD, the mycobacterial MCAT, has been determined to 3.0 A resolution by multi-wavelength anomalous dispersion. Phasing was facilitated by Ni2+ ions bound to the 20-residue N-terminal affinity tag, which packed between the two independent copies of mtFabD.

  3. Sex- and Size-Related Patterns of Carrion Visitation in Necrodes littoralis (Coleoptera: Silphidae) and Creophilus maxillosus (Coleoptera: Staphylinidae). (United States)

    Mądra-Bielewicz, Anna; Frątczak-Łagiewska, Katarzyna; Matuszewski, Szymon


    The estimation of postmortem interval (PMI) based on successional patterns of adult insects is largely limited, due to the lack of potential PMI markers. Sex and size of adult insects could be easily used for such estimation. In this study, sex- and size-related patterns of carrion attendance by adult insects were analyzed in Necrodes littoralis (Coleoptera: Silphidae) and Creophilus maxillosus (Coleoptera: Staphylinidae). For both species, abundance of males and females changed similarly during decomposition. A slightly female-biased sex ratio was recorded in N. littoralis. Females of N. littoralis started visiting carcasses, on average, one day earlier than males. There was a rise in size of males of N. littoralis at the end of decomposition, whereas for females of both species and males of C. maxillosus, no size-related patterns of carrion visitation were found. Current results demonstrate that size and sex of adult carrion beetles are poor indicators of PMI.

  4. Estados inmaduros de Ancognatha ustulata (Coleoptera: Melolonthidae: Dynastinae: Cyclocephalini Immature stages of Ancognatha ustulata (Coleoptera: Melolonthidae: Dynastinae: Cyclocephalini

    Directory of Open Access Journals (Sweden)

    Jhon César Neita-Moreno


    Full Text Available Se describen y se ilustran por primera vez la larva de tercer estadio y la pupa de Ancognatha ustulata Burmeister, 1847 (Coleoptera: Melolonthidae: Dynastinae: Cyclocephalini. Se aportan datos sobre la biología de la especie y su distribución en Colombia. Se proporciona una clave para la identificación de las larvas de tercer estadio conocidas de las especies del género Ancognatha Erichson.The third instar larva and pupa of Ancognatha ustulata Burmeister, 1847 (Coleoptera: Melolonthidae: Dynastinae: Cyclocephalini are described and illustrated for the first time. New data on larval biology and distribution of the species in Colombia are included. A key to the known third stage larvae of Ancognatha Erichson is provided.

  5. Enterococcus faecalis endocarditis severity in rabbits is reduced by IgG Fabs interfering with aggregation substance.

    Directory of Open Access Journals (Sweden)

    Patrick M Schlievert

    Full Text Available BACKGROUND: Enterococcus faecalis is a significant cause of infective endocarditis, an infection of the heart endothelium leading to vegetation formation (microbes, fibrin, platelets, and host cells attached to underlying endothelial tissue. Our previous research determined that enterococcal aggregation substance (AS is an important virulence factor in causation of endocarditis, although endocarditis may occur in the absence of AS production. Production of AS by E. faecalis causes the organism to form aggregates through AS binding to enterococcal binding substance. In this study, we assessed the ability of IgGs and IgG Fabs against AS to provide protection against AS+ E. faecalis endocarditis. METHODOLOGY/PRINCIPAL FINDINGS: When challenged with AS+ E. faecalis, 10 rabbits actively immunized against AS+ E. faecalis developed more significant vegetations than 9 animals immunized against AS⁻E. faecalis, and 9/10 succumbed compared to 2/9 (p<0.005, suggesting enhanced aggregation by IgG contributes significantly to disease. IgG antibodies against AS also enhanced enterococcal aggregation as tested in vitro. In contrast, Fab fragments of IgG from rabbits immunized against purified AS, when passively administered to rabbits (6/group immediately before challenge with AS+E. faecalis, reduced total vegetation (endocarditis lesion microbial counts (7.9 x 10⁶ versus 2.0 x 10⁵, p = 0.02 and size (40 mg versus 10, p = 0.05. In vitro, the Fabs prevented enterococcal aggregation. CONCLUSIONS/SIGNIFICANCE: The data confirm the role of AS in infective endocarditis formation and suggest that use of Fabs against AS will provide partial protection from AS+E. faecalis illness.

  6. Construction of a large synthetic human Fab antibody library on yeast cell surface by optimized yeast mating. (United States)

    Baek, Du-San; Kim, Yong-Sung


    Yeast surface-displayed antibody libraries provide an efficient and quantitative screening resource for given antigens, but suffer from typically modest library sizes owing to low yeast transformation efficiency. Yeast mating is an attractive method for overcoming the limit of yeast transformation to construct a large, combinatorial antibody library, but the optimal conditions have not been reported. Here, we report a large synthetic human Fab (antigen binding fragment) yeast surface-displayed library generated by stepwise optimization of yeast mating conditions. We first constructed HC (heavy chain) and LC (light chain) libraries, where all of the six CDRs (complementarity-determining regions) of the variable domains were diversified mimicking the human germline antibody repertoires by degenerate codons, onto single frameworks of VH3-23 and Vkappa1-16 germline sequences, in two haploid cells of opposite mating types. Yeast mating conditions were optimized in the order of cell density, media pH, and cell growth phase, yielding a mating efficiency of ~58% between the two haploid cells carrying HC and LC libraries. We constructed two combinatorial Fab libraries with CDR-H3 of 9 or 11 residues in length with colony diversities of more than 10(9) by one round of yeast mating between the two haploid HC and LC libraries, with modest diversity sizes of ~10(7). The synthetic human Fab yeast-displayed libraries exhibited relative amino acid compositions in each position of the six CDRs that were very similar to those of the designed repertoires, suggesting that they are a promising source for human Fab antibody screening.

  7. Use of the gills of Pimelodus maculatus (Lacèpéde, 1803 (Siluriformes, Pimelodidae as a biomarker of pollution in the reservoir of UHE Marechal Mascarenhas de Moraes, Minas Gerais, Brazil

    Directory of Open Access Journals (Sweden)

    Diego José Nogueira


    Full Text Available The gills of fish are biomarkers of environmental toxicity which are often used to assess the impacts of xenobiotics. This study aims to evaluate the contamination of the reservoir Marechal Mascarenhas de Moraes, through the estimation ofthe rate of morphological changes in the gills of Pimelodus maculatus caused by toxic agents. The specimens of P. maculatus were collected from three points of the reservoir of UHE Marechal Mascarenhas de Moraes. In the local of collection, immediately, the gills of the collected individuals were removed and fixed in Bouin. They were stained with Hematoxylin-Eosin for light microscopic analysis. The quantification of changes was carried out through stereology and the indices of change were estimated, classifying the lesions from moderate to severe in the three points under study.

  8. The Response Regulator YycF Inhibits Expression of the Fatty Acid Biosynthesis Repressor FabT in Streptococcus pneumoniae. (United States)

    Mohedano, Maria L; Amblar, Mónica; de la Fuente, Alicia; Wells, Jerry M; López, Paloma


    The YycFG (also known as WalRK, VicRK, MicAB, or TCS02) two-component system (TCS) is highly conserved among Gram-positive bacteria with a low G+C content. In Streptococcus pneumoniae the YycF response regulator has been reported to be essential due to its control of pcsB gene expression. Previously we showed that overexpression of yycF in S. pneumoniae TIGR4 altered the transcription of genes involved in cell wall metabolism and fatty acid biosynthesis, giving rise to anomalous cell division and increased chain length of membrane fatty acids. Here, we have overexpressed the yycFG system in TIGR4 wild-type strain and yycF in a TIGR4 mutant depleted of YycG, and analyzed their effects on expression of proteins involved in fatty acid biosynthesis during activation of the TCS. We demonstrate that transcription of the fab genes and levels of their products were only altered in the YycF overexpressing strain, indicating that the unphosphorylated form of YycF is involved in the regulation of fatty acid biosynthesis. In addition, DNA-binding assays and in vitro transcription experiments with purified YycF and the promoter region of the FabTH-acp operon support a direct inhibition of transcription of the FabT repressor by YycF, thus confirming the role of the unphosphorylated form in transcriptional regulation.

  9. Characterization and crystallization of a recombinant IgE Fab fragment in complex with the bovine β-lactoglobulin allergen

    Energy Technology Data Exchange (ETDEWEB)

    Niemi, Merja, E-mail:; Jänis, Janne [Department of Chemistry, University of Joensuu, PO Box 111, FIN-80101 Joensuu (Finland); Jylhä, Sirpa [VTT Technical Research Centre of Finland, PO Box 1000, FIN-02044 VTT (Finland); Kallio, Johanna M.; Hakulinen, Nina [Department of Chemistry, University of Joensuu, PO Box 111, FIN-80101 Joensuu (Finland); Laukkanen, Marja-Leena; Takkinen, Kristiina [VTT Technical Research Centre of Finland, PO Box 1000, FIN-02044 VTT (Finland); Rouvinen, Juha [Department of Chemistry, University of Joensuu, PO Box 111, FIN-80101 Joensuu (Finland)


    The high-resolution mass-spectrometric characterization, crystallization and X-ray diffraction studies of a recombinant IgE Fab fragment in complex with bovine β-lactoglobulin are reported. A D1 Fab fragment containing the allergen-binding variable domains of the IgE antibody was characterized by ESI FT–ICR mass spectrometry and crystallized with bovine β-lactoglobulin (BLG) using the hanging-drop vapour-diffusion method at 293 K. X-ray data suitable for structure determination were collected to 2.8 Å resolution using synchrotron radiation. The crystal belonged to the orthorhombic space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 67.0, b = 100.6, c = 168.1 Å. The three-dimensional structure of the D1 Fab fragment–BLG complex will provide the first insight into IgE antibody–allergen interactions at the molecular level.

  10. Crystallization and preliminary X-ray analysis of enoyl-acyl carrier protein reductase (FabK) from Streptococcus pneumoniae

    Energy Technology Data Exchange (ETDEWEB)

    Saito, Jun, E-mail:; Yamada, Mototsugu; Watanabe, Takashi; Kitagawa, Hideo; Takeuchi, Yasuo [Pharmaceutical Research Center, Meiji Seika Kaisha Ltd, 760 Morooka-cho, Kohoku-ku, Yokohama 222-8567 (Japan)


    Enoyl-acyl carrier protein (ACP) reductases are responsible for bacterial type II fatty-acid biosynthesis and are attractive targets for developing novel antibiotics. The S. pneumoniae enoyl-ACP reductase (FabK) was crystallized and selenomethionine MAD data were collected to 2 Å resolution. The enoyl-acyl carrier protein (ACP) reductase from Streptococcus pneumoniae (FabK; EC is responsible for catalyzing the final step in each elongation cycle of fatty-acid biosynthesis. Selenomethionine-substituted FabK was purified and crystallized by the hanging-drop vapour-diffusion method at 277 K. The crystal belongs to space group P2{sub 1}, with unit-cell parameters a = 50.26, b = 126.70, c = 53.63 Å, β = 112.46°. Diffraction data were collected to 2.00 Å resolution using synchrotron beamline BL32B2 at SPring-8. Two molecules were estimated to be present in the asymmetric unit, with a solvent content of 45.1%.

  11. Molecular modeling and simulation of FabG, an enzyme involved in the fatty acid pathway of Streptococcus pyogenes. (United States)

    Shafreen, Rajamohmed Beema; Pandian, Shunmugiah Karutha


    Streptococcus pyogenes (SP) is the major cause of pharyngitis accompanied by strep throat infections in humans. 3-keto acyl reductase (FabG), an important enzyme involved in the elongation cycle of the fatty acid pathway of S. pyogenes, is essential for synthesis of the cell-membrane, virulence factors and quorum sensing-related mechanisms. Targeting SPFabG may provide an important aid for the development of drugs against S. pyogenes. However, the absence of a crystal structure for FabG of S. pyogenes limits the development of structure-based drug designs. Hence, in the present study, a homology model of FabG was generated using the X-ray crystallographic structure of Aquifex aeolicus (PDB ID: 2PNF). The modeled structure was refined using energy minimization. Furthermore, active sites were predicted, and a large dataset of compounds was screened against SPFabG. The ligands were docked using the LigandFit module that is available from Discovery Studio version 2.5. From this list, 13 best hit ligands were chosen based on the docking score and binding energy. All of the 13 ligands were screened for Absorption, Distribution, Metabolism, Excretion and Toxicity (ADMET) properties. From this, the two best descriptors, along with one descriptor that lay outside the ADMET plot, were selected for molecular dynamic (MD) simulation. In vitro testing of the ligands using biological assays further substantiated the efficacy of the ligands that were screened based on the in silico methods.

  12. The Response Regulator YycF Inhibits Expression of the Fatty Acid Biosynthesis Repressor FabT in Streptococcus pneumoniae (United States)

    Mohedano, Maria L.; Amblar, Mónica; de la Fuente, Alicia; Wells, Jerry M.; López, Paloma


    The YycFG (also known as WalRK, VicRK, MicAB, or TCS02) two-component system (TCS) is highly conserved among Gram-positive bacteria with a low G+C content. In Streptococcus pneumoniae the YycF response regulator has been reported to be essential due to its control of pcsB gene expression. Previously we showed that overexpression of yycF in S. pneumoniae TIGR4 altered the transcription of genes involved in cell wall metabolism and fatty acid biosynthesis, giving rise to anomalous cell division and increased chain length of membrane fatty acids. Here, we have overexpressed the yycFG system in TIGR4 wild-type strain and yycF in a TIGR4 mutant depleted of YycG, and analyzed their effects on expression of proteins involved in fatty acid biosynthesis during activation of the TCS. We demonstrate that transcription of the fab genes and levels of their products were only altered in the YycF overexpressing strain, indicating that the unphosphorylated form of YycF is involved in the regulation of fatty acid biosynthesis. In addition, DNA-binding assays and in vitro transcription experiments with purified YycF and the promoter region of the FabTH-acp operon support a direct inhibition of transcription of the FabT repressor by YycF, thus confirming the role of the unphosphorylated form in transcriptional regulation. PMID:27610104

  13. A fully synthetic human Fab antibody library based on fixed VH/VL framework pairings with favorable biophysical properties. (United States)

    Tiller, Thomas; Schuster, Ingrid; Deppe, Dorothée; Siegers, Katja; Strohner, Ralf; Herrmann, Tanja; Berenguer, Marion; Poujol, Dominique; Stehle, Jennifer; Stark, Yvonne; Heßling, Martin; Daubert, Daniela; Felderer, Karin; Kaden, Stefan; Kölln, Johanna; Enzelberger, Markus; Urlinger, Stefanie


    This report describes the design, generation and testing of Ylanthia, a fully synthetic human Fab antibody library with 1.3E+11 clones. Ylanthia comprises 36 fixed immunoglobulin (Ig) variable heavy (VH)/variable light (VL) chain pairs, which cover a broad range of canonical complementarity-determining region (CDR) structures. The variable Ig heavy and Ig light (VH/VL) chain pairs were selected for biophysical characteristics favorable to manufacturing and development. The selection process included multiple parameters, e.g., assessment of protein expression yield, thermal stability and aggregation propensity in fragment antigen binding (Fab) and IgG1 formats, and relative Fab display rate on phage. The framework regions are fixed and the diversified CDRs were designed based on a systematic analysis of a large set of rearranged human antibody sequences. Care was taken to minimize the occurrence of potential posttranslational modification sites within the CDRs. Phage selection was performed against various antigens and unique antibodies with excellent biophysical properties were isolated. Our results confirm that quality can be built into an antibody library by prudent selection of unmodified, fully human VH/VL pairs as scaffolds.

  14. Enterococcus faecalis Endocarditis Severity in Rabbits Is Reduced by IgG Fabs Interfering with Aggregation Substance (United States)

    Schlievert, Patrick M.; Chuang-Smith, Olivia N.; Peterson, Marnie L.; Cook, Laura C. C.; Dunny, Gary M.


    Background Enterococcus faecalis is a significant cause of infective endocarditis, an infection of the heart endothelium leading to vegetation formation (microbes, fibrin, platelets, and host cells attached to underlying endothelial tissue). Our previous research determined that enterococcal aggregation substance (AS) is an important virulence factor in causation of endocarditis, although endocarditis may occur in the absence of AS production. Production of AS by E. faecalis causes the organism to form aggregates through AS binding to enterococcal binding substance. In this study, we assessed the ability of IgGs and IgG Fabs against AS to provide protection against AS+ E. faecalis endocarditis. Methodology/Principal Findings When challenged with AS+ E. faecalis, 10 rabbits actively immunized against AS+ E. faecalis developed more significant vegetations than 9 animals immunized against AS− E. faecalis, and 9/10 succumbed compared to 2/9 (pendocarditis lesion) microbial counts (7.9×106 versus 2.0×105, p = 0.02) and size (40 mg versus 10, p = 0.05). In vitro, the Fabs prevented enterococcal aggregation. Conclusions/Significance The data confirm the role of AS in infective endocarditis formation and suggest that use of Fabs against AS will provide partial protection from AS+ E. faecalis illness. PMID:20957231

  15. Antigen binding of human IgG Fabs mediate ERK-associated proliferation of human breast cancer cells. (United States)

    Wen, Yue-Jin; Mancino, Anne; Pashov, Anastas; Whitehead, Tracy; Stanley, Joseph; Kieber-Emmons, Thomas


    Serum-circulating antibody can be linked to poor outcomes in some cancer patients. To investigate the role of human antibodies in regulating tumor cell growth, we constructed a recombinant cDNA expression library of human IgG Fab from a patient with breast cancer. Clones were screened from the library with breast tumor cell lysate. Sequence analysis of the clones showed somatic hypermutations when compared to their closest VH/VL germ-line genes. Initial characterizations focused on five clones. All tested clones displayed stronger binding to antigen derived from primary breast cancers and established breast cancer cell lines than to normal breast tissues. In vitro functional studies showed that four out of five tested clones could stimulate the growth of MDA-MB-231 breast cancer cell lines, and one out of five was able to promote MCF-7 cell growth as well. Involvement of ERK2 pathway was observed. By 1H-NMR spectra and Western blot analysis, it was evident that two tested antibody Fabs are capable of interacting with sialic acid. Our study suggests a possible role for human antibody in promoting tumor cell growth by direct binding of IgG Fab to breast tumor antigen. Such studies prompt speculation regarding the role of serum antibodies in mediating tumor growth as well as their contribution to disease progression.

  16. Independent evolution of Fc- and Fab-mediated HIV-1-specific antiviral antibody activity following acute infection (United States)

    Dugast, Anne-Sophie; Stamatatos, Leonidas; Tonelli, Andrew; Suscovich, Todd J.; Licht, Anna F.; Mikell, Iliyana; Ackerman, Margaret E.; Streeck, Hendrik; Klasse, P.J.; Moore, John P.; Alter, Galit


    Fc-related antibody activities, such as antibody-dependent cellular cytotoxicity (ADCC), or more broadly, antibody-mediated cellular viral inhibition (ADCVI), play a role in curbing early SIV viral replication, are enriched in human long-term infected non-progressors, and could potentially contribute to protection from infection. However, little is known about the mechanism by which such humoral immune responses are naturally induced following infection. Here we focused on the early evolution of the functional antibody response, largely driven by the Fc portion of the antibody, in the context of the evolving binding and neutralizing antibody response, which is driven mainly by the antibody binding fragment (Fab). We show that ADCVI/ADCC-inducing responses in humans are rapidly generated following acute HIV-1 infection, peak at approximately 6 months post-infection, but decay rapidly in the setting of persistent immune activation, as Fab-related activities persistently increase. Moreover, the loss of Fc activity occurred in synchrony with a loss of HIV-specific IgG3 responses. Our data strongly suggest that Fc- and Fab-related antibody functions are modulated in a distinct manner following acute HIV infection. Vaccination strategies intended to optimally induce both sets of antiviral antibody activities may, therefore, require a fine-tuning of the inflammatory response. PMID:25043633

  17. Slow-Onset Inhibition of the FabI Enoyl Reductase from Francisella tularensis: Residence Time and in Vivo Activity

    Energy Technology Data Exchange (ETDEWEB)

    Lu, H.; England, K; Ende, C; Truglio, J; Luckner, S; Reddy, B; Marlenee, N; Knudson, S; Knudson, D; et. al.


    Francisella tularensis is a highly virulent and contagious Gram-negative intracellular bacterium that causes the disease tularemia in mammals. The high infectivity and the ability of the bacterium to survive for weeks in a cool, moist environment have raised the possibility that this organism could be exploited deliberately as a potential biological weapon. Fatty acid biosynthesis (FAS-II) is essential for bacterial viability and has been validated as a target for the discovery of novel antibacterials. The FAS-II enoyl reductase ftuFabI has been cloned and expressed, and a series of diphenyl ethers have been identified that are subnanomolar inhibitors of the enzyme with MIC90 values as low as 0.00018 ?g mL-1. The existence of a linear correlation between the Ki and MIC values strongly suggests that the antibacterial activity of the diphenyl ethers results from direct inhibition of ftuFabI within the cell. The compounds are slow-onset inhibitors of ftuFabI, and the residence time of the inhibitors on the enzyme correlates with their in vivo activity in a mouse model of tularemia infection. Significantly, the rate of breakdown of the enzyme-inhibitor complex is a better predictor of in vivo activity than the overall thermodynamic stability of the complex, a concept that has important implications for the discovery of novel chemotherapeutics that normally rely on equilibrium measurements of potency.

  18. Identification of highly reactive cysteine residues at less exposed positions in the Fab constant region for site-specific conjugation. (United States)

    Shiraishi, Yasuhisa; Muramoto, Takashige; Nagatomo, Kazutaka; Shinmi, Daisuke; Honma, Emiko; Masuda, Kazuhiro; Yamasaki, Motoo


    Engineered cysteine residues are currently used for the site-specific conjugation of antibody-drug conjugates (ADC). In general, positions on the protein surface have been selected for substituting a cysteine as a conjugation site; however, less exposed positions (with less than 20% of accessible surface area [ASA]) have not yet been evaluated. In this study, we engineered original cysteine positional variants of a Fab fragment, with less than 20% of ASA, and evaluated their thiol reactivities through conjugation with various kinds of payloads. As a result, we have identified three original cysteine positional variants (heavy chain: Hc-A140C, light chain: Lc-Q124C and Lc-L201C), which exhibited similar monomer content, thermal stability, and antigen binding affinity in comparison to the wild-type Fab. In addition, the presence of cysteine in these positions made it possible for the Fab variants to react with variable-sized molecules with high efficiency. The favorable physical properties of the cysteine positional variants selected in our study suggest that less exposed positions, with less than 20% of ASA, provide an alternative for creating conjugation sites.

  19. Dual Constant Domain-Fab: A novel strategy to improve half-life and potency of a Met therapeutic antibody. (United States)

    Cignetto, Simona; Modica, Chiara; Chiriaco, Cristina; Fontani, Lara; Milla, Paola; Michieli, Paolo; Comoglio, Paolo M; Vigna, Elisa


    The kinase receptor encoded by the Met oncogene is a sensible target for cancer therapy. The chimeric monovalent Fab fragment of the DN30 monoclonal antibody (MvDN30) has an odd mechanism of action, based on cell surface removal of Met via activation of specific plasma membrane proteases. However, the short half-life of the Fab, due to its low molecular weight, is a severe limitation for the deployment in therapy. This issue was addressed by increasing the Fab molecular weight above the glomerular filtration threshold through the duplication of the constant domains, in tandem (DCD-1) or reciprocally swapped (DCD-2). The two newly engineered molecules showed biochemical properties comparable to the original MvDN30 in vitro, acting as full Met antagonists, impairing Met phosphorylation and activation of downstream signaling pathways. As a consequence, Met-mediated biological responses were inhibited, including anchorage-dependent and -independent cell growth. In vivo DCD-1 and DCD-2 showed a pharmacokinetic profile significantly improved over the original MvDN30, doubling the circulating half-life and reducing the clearance. In pre-clinical models of cancer, generated by injection of tumor cells or implant of patient-derived samples, systemic administration of the engineered molecules inhibited the growth of Met-addicted tumors.

  20. 抗轮状病毒IgY和Fab的研制%Preparation of IgY and Fab against HRV

    Institute of Scientific and Technical Information of China (English)

    孙淑清; 孟岩; 段春燕; 胡彦涛



  1. Site-Specific Photolabeling of the IgG Fab Fragment Using a Small Protein G Derived Domain. (United States)

    Kanje, Sara; von Witting, Emma; Chiang, Samuel C C; Bryceson, Yenan T; Hober, Sophia


    Antibodies are widely used reagents for recognition in both clinic and research laboratories all over the world. For many applications, antibodies are labeled through conjugation to different reporter molecules or therapeutic agents. Traditionally, antibodies are covalently conjugated to reporter molecules via primary amines on lysines or thiols on cysteines. While efficient, such labeling is variable and nonstoichiometric and may affect an antibody's binding to its target. Moreover, an emerging field for therapeutics is antibody-drug conjugates, where a toxin or drug is conjugated to an antibody in order to increase or incorporate a therapeutic effect. It has been shown that homogeneity and controlled conjugation are crucial in these therapeutic applications. Here we present two novel protein domains developed from an IgG-binding domain of Streptococcal Protein G. These domains show obligate Fab binding and can be used for site-specific and covalent attachment exclusively to the constant part of the Fab fragment of an antibody. The two different domains can covalently label IgG of mouse and human descent. The labeled antibodies were shown to be functional in both an ELISA and in an NK-cell antibody-dependent cellular cytotoxicity assay. These engineered protein domains provide novel tools for controlled labeling of Fab fragments and full-length IgG.

  2. Sexing murine embryos with an indirect immunofluorescence assay using phage antibody B9-Fab against SDM antigen. (United States)

    Wang, Naidong; Yuan, Anwen; Ma, Jun; Deng, Zhibang; Xue, Liqun


    The use of serologically detectable male (SDM; also called H-Y) antigens to identify male embryos may be limited by the source of anti-SDM antibody. In the present study, novel anti-SDM B9-Fab recombinant clones (obtained by chain shuffling of an A8 original clone) were used to detect SDM antigens on murine embryos. Murine morulae and blastocysts (n=138) were flushed from the oviducts of Kunming mice and incubated with anti-SDM B9-Fab for 30 min at 37°C. With an indirect immunofluorescence assay, the membrane and inner cell mass had bright green fluorescence (presumptive males). Overall, 43.5% (60/138) were classified as presumptive males and 56.5% (78/138) as presumptive females, with 85.0 and 88.5% of these, respectively, confirmed as correct predictions (based on PCR analysis of a male-specific [Sry] sequence). We concluded that the anti-SDM B9-Fab molecule had potential for non-invasive, technically simple immunological sexing of mammalian embryos.

  3. Crystal structure of human TWEAK in complex with the Fab fragment of a neutralizing antibody reveals insights into receptor binding.

    Directory of Open Access Journals (Sweden)

    Alfred Lammens

    Full Text Available The tumor necrosis factor-like weak inducer of apoptosis (TWEAK is a multifunctional cytokine playing a key role in tissue regeneration and remodeling. Dysregulation of TWEAK signaling is involved in various pathological processes like autoimmune diseases and cancer. The unique interaction with its cognate receptor Fn14 makes both ligand and receptor promising targets for novel therapeutics. To gain insights into this important signaling pathway, we determined the structure of soluble human TWEAK in complex with the Fab fragment of an antibody selected for inhibition of receptor binding. In the crystallized complex TWEAK is bound by three Fab fragments of the neutralizing antibody. Homology modeling shows that Fab binding overlaps with the putative Fn14 binding site of TWEAK. Docking of the Fn14 cysteine rich domain (CRD to that site generates a highly complementary interface with perfectly opposing charged and hydrophobic residues. Taken together the presented structure provides new insights into the biology of TWEAK and the TWEAK/Fn14 pathway, which will help to optimize the therapeutic strategy for treatment of related cancer types and autoimmune diseases.

  4. The response regulator YycF inhibits expression of the fatty acid biosynthesis repressor FabT in Streptococcus pneumoniae

    Directory of Open Access Journals (Sweden)

    Maria Luz Mohedano


    Full Text Available The YycFG (also known as WalRK, VicRK, MicAB or TCS02 two-component system (TCS is highly conserved among Gram-positive bacteria with a low G+C content. In Streptococcus pneumoniae the YycF response regulator has been reported to be essential due to its control of pcsB gene expression. Previously we showed that overexpression of yycF in S. pneumoniae TIGR4 altered the transcription of genes involved in cell wall metabolism and fatty acid biosynthesis, giving rise to anomalous cell division and increased chain length of membrane fatty acids. Here, we have overexpressed the yycFG system in TIGR4 wild-type strain and yycF in a TIGR4 mutant depleted of YycG, and analyzed their effects on expression of proteins involved in fatty acid biosynthesis during activation of the TCS. We demonstrate that transcription of the fab genes and levels of their products were only altered in the YycF overexpressing strain, indicating that the unphosphorylated form of YycF is involved in the regulation of fatty acid biosynthesis. In addition, DNA-binding assays and in vitro transcription experiments with purified YycF and the promoter region of the FabTH-acp operon support a direct inhibition of transcription of the FabT repressor by YycF, thus confirming the role of the unphosphorylated form in transcriptional regulation.

  5. Structural and functional insights into the anti-BACE1 Fab fragment that recognizes the BACE1 exosite. (United States)

    Gutiérrez, Lucas Joel; Andujar, Sebastián Antonio; Enriz, Ricardo Daniel; Baldoni, Héctor Armando


    A molecular modeling study giving structural, functional, and mutagenesis insights into the anti-BACE1 Fab fragment that recognizes the BACE1 exosite is reported. Our results allow extending experimental data resulting from X-ray diffraction experiments in order to examine unknown aspects for the Fab-BACE1 recognition and its binding mode. Thus, the study performed here allows extending the inherently static nature of crystallographic structures in order to gain a deeper understanding of the structural and dynamical basis at the atomic level. The characteristics and strength of the interatomic interactions involved in the immune complex formation are exhaustively analyzed. The results might explain how the anti-BACE1 Fab fragment and other BACE1 exosite binders are capable to produce an allosteric modulation of the BACE1 activity. Our site-directed mutagenesis study indicated that the functional anti-BACE1 paratope, residues Tyr32 (H1), Trp50 (H2), Arg98 (H3), Phe101 (H3), Trp104 (H3) and Tyr94 (L3), strongly dominates the binding energetics with the BACE1 exosite. The mutational studies described in this work might accelerate the development of new BACE1 exosite binders with interesting pharmacological activity.

  6. Generation and characterization of a human neutralizing bivalent antibody Fab094-DDD against rabies virus%人源特异性抗狂犬病毒二价Fab094-DDD抗体的制备及鉴定

    Institute of Scientific and Technical Information of China (English)


    目的:利用蛋白激酶A(protein kinase A,PKA)调节亚基(R亚基)的二聚体化功能结构域(DDD)的二聚化功能,制备具有中和活性的人源特异性抗狂犬病毒二价抗体.方法:优化合成linker-C-DDD序列,设计引物扩增抗狂犬病毒抗体Fab094的Fd段和轻链可变区序列(Vκ),通过overlap PCR将Fab094的Fd段和linker-C-DDD基因重组为Fd-DDD,将其和Vκ分别克隆到真核表达载体中,共转染293Free style细胞,表达纯化该抗体.应用SDS-PAGE 、Western blot、ELISA、免疫共沉淀、亲和力测定及免疫荧光试验检测该抗体的免疫学特性,用荧光抗体病毒中和试验检测其中和活性.结果:成功构建了全人源抗狂犬病毒二价抗体Fab094-DDD的真核表达载体,获得的二价Fab094-DDD抗体与抗原保持着较好的亲和力,能特异性结合狂犬病毒,中和效价为213.2 U/mg.结论:成功制备了抗狂犬病毒二价Fab094-DDD抗体,具有较高的中和活性,为进一步研发狂犬病治疗性抗体药物奠定了基础,对于其他疾病双表位人源特异性抗体的制备也具有借鉴作用.

  7. Medically important beetles (insecta: coleoptera of Iran

    Directory of Open Access Journals (Sweden)

    MR Nikbakhtzadeh


    Full Text Available This study focused on coleopteran species that are responsible for the emergence of recent cases of dermatological manifestations in Iran. To the best of our knowledge, five species of the family Meloidae and nine species of the genus Paederus are by far the only beetles recognized as medically important in Iran. The staphylinids consists of Paederus ilsae, P. iliensis, P. fuscipes, P. kalalovae, P. balcanicus, P. lenkoranus, P. littoralis, P. carpathicus, P. nigricornis, while the meloids are Mylabris impressa, M. guerini, Muzimes iranicus, Alosimus smyrnensis and Epicauta sharpi. Most cases of linear dermatitis in this country occur in areas bordering the Caspian Sea. This problem is caused by beetles of the genus Paederus which are present as adults from mid-April to October with particularly high incidences from May to August. Fars (in southern Iran ranks second in number of cases of insect-induced dermatitis. The third major region in which this type of dermatitis has been recorded is Hamedan Province, in the west of the country. Meloid dermatitis showed its highest severity in 2001, when a considerable number of patients sought medical help in Toyserkan and Nahavand counties. New cases of skin blistering were reported along the Persian Gulf coast and the agent was identified as Epicauta sharpi (Coleoptera: Meloidae. In all these regions, it was observed that recorded cases of lesions coincided precisely with the yearly peaks of the beetles. Paederus fuscipes and P. kalalovae are the predominant species along the Caspian Sea shore. It appears that P. fuscipes is homogeneously distributed throughout the Caspian Sea region while the distribution of the other species is more irregular. Paederus fuscipes is probably the major agent that causes linear dermatitis in northern Iran. Whereas this disease is a rural difficulty in the south, mainly in villages or small towns, it is an urban problem in northern provinces along the Caspian Sea shore

  8. Route of infection and hematological effect of Metarhizium anisopliae (Metsch.) Sorokin on Dysdercus cingulatus (Fab.) adult. (United States)

    Sahayaraj, Kitherian; Borgio, Jesu Francis; Lucini, Luigi


    The primary objective of this work was to identify, under laboratory conditions, the route of infection and hemogram of Dysdercus cingulatus (Fab.) adults by Metarhizium anisopliae. The infection process in D. cingulatus by M. anisopliae involved the conidia adherence to the host cuticle and germination after 24 h post-infection, accompanied by falling of bristles. The subsequent step, within 24-48 h post-infection, comprised penetration of fungus through spiracles, root of bristles, hemolymph, and the three dorsal sacs. Subsequently, within 72-96 h post-infection, the fungus penetrated into trachea and sacs, then emerged on cuticular surface and was found to be maximum in hemolymph. A great decrease in hemocytes count was observed within 96 h from infection. The hemosomic index (HSI) decreased gradually as the incubation period increased. As far as we know, this is the first study to know the mechanism of action of M. anisopliae to D. cingulatus.

  9. Characterization of silicon photomultipliers at National Nano-Fab Center for PET-MR. (United States)

    Kim, Hyoungtaek; Sul, Woo Suk; Cho, Gyuseong


    The silicon photomultipliers (SiPMs) were fabricated for magnetic resonance compatible positron emission tomography (PET) applications using customized CMOS processes at National NanoFab Center. Each micro-cell consists of a shallow n+/p well junction on a p-type epitaxial wafer and passive quenching circuit was applied. The size of the SiPM is 3 × 3 mm(2) and the pitch of each micro-cell is 65 μm. In this work, several thousands of SiPMs were packaged and tested to build a PET ring detector which has a 60 mm axial and 390 mm radial field of view. I-V characteristics of the SiPMs are shown good uniformity and breakdown voltage is around 20 V. The photon detection efficiency was measured via photon counting method and the maximum value was recorded as 16% at 470 nm. The gamma ray spectrum of a Ge-68 isotope showed nearly 10% energy resolution at 511 keV with a 3 × 3 × 20 mm(3) LYSO crystal.

  10. Imaging of cardiac allograft rejection in dogs using indium-111 monoclonal antimyosin Fab

    Energy Technology Data Exchange (ETDEWEB)

    Addonizio, L.J.; Michler, R.E.; Marboe, C.; Esser, P.E.; Johnson, L.L.; Seldin, D.W.; Gersony, W.M.; Alderson, P.O.; Rose, E.A.; Cannon, P.J.


    The acute rejection of cardiac allografts is currently diagnosed by the presence of myocyte necrosis on endomyocardial biopsy. We evaluated the efficacy of noninvasive scintigraphic imaging with indium-111-labeled anticardiac myosin Fab fragments (indium-111 antimyosin) to detect and quantify cardiac allograft rejection. Six dogs that had intrathoracic heterotopic cardiac allograft transplantation were injected with indium-111 antimyosin and planar and single photon emission computed tomographic (SPECT) images were obtained in various stages of acute and subacute rejection. Four dogs had an allograft older than 8 months and had been on long-term immunosuppressive therapy; two dogs had an allograft less than 2 weeks old and were not on immunosuppressive therapy. Count ratios comparing heterotopic with native hearts were calculated from both SPECT images and in vitro scans of excised and sectioned hearts and were compared with the degree of rejection scored by an independent histopathologic review. Indium-111 antimyosin uptake was not visible in planar or SPECT images of native hearts. Faint diffuse uptake was apparent in cardiac allografts during long-term immunosuppression and intense radioactivity was present in hearts with electrocardiographic evidence of rejection. The heterotopic to native heart count ratios in SPECT images correlated significantly with the count ratios in the excised hearts (r = 0.93) and with the histopathologic rejection score (r = 0.97). The distribution of indium-111 antimyosin activity in right and left ventricles corresponded to areas of histopathologic abnormalities.

  11. Characterization of silicon photomultipliers at National Nano-Fab Center for PET-MR (United States)

    Kim, Hyoungtaek; Sul, Woo Suk; Cho, Gyuseong


    The silicon photomultipliers (SiPMs) were fabricated for magnetic resonance compatible positron emission tomography (PET) applications using customized CMOS processes at National NanoFab Center. Each micro-cell consists of a shallow n+/p well junction on a p-type epitaxial wafer and passive quenching circuit was applied. The size of the SiPM is 3 × 3 mm2 and the pitch of each micro-cell is 65 μm. In this work, several thousands of SiPMs were packaged and tested to build a PET ring detector which has a 60 mm axial and 390 mm radial field of view. I-V characteristics of the SiPMs are shown good uniformity and breakdown voltage is around 20 V. The photon detection efficiency was measured via photon counting method and the maximum value was recorded as 16% at 470 nm. The gamma ray spectrum of a Ge-68 isotope showed nearly 10% energy resolution at 511 keV with a 3 × 3 × 20 mm3 LYSO crystal.

  12. An efficient process of generating bispecific antibodies via controlled Fab-arm exchange using culture supernatants. (United States)

    Paul, Suparna; Connor, Judy; Nesspor, Tom; Haytko, Peter; Boakye, Ken; Chiu, Mark L; Jiang, Haiyan


    Bispecific antibody generation is actively pursued for therapeutic and research antibody development. Although there are multiple strategies for generating bispecific antibodies (bsAbs); the common challenge is to develop a scalable method to prepare bsAbs with high purity and yield. The controlled Fab-arm exchange (cFAE) method combines two parental monoclonal antibodies (mAbs), each with a matched point mutation, F405L and K409R in the respective CH3 domains. The conventional process employs two steps: the purification of two parental mAbs from culture supernatants followed by cFAE. Following a reduction/oxidation reaction, the bispecific mAb is formed with greater than 95% heterodimerization efficiency. In this study, cFAE was initiated in culture supernatants expressing the two parental mAbs, thereby eliminating the need to first purify the parental mAbs. The bsAbs formed in culture supernatant was then purified using a Protein A affinity chromatography. The BsAbs generated in this manner had efficiency comparable to the conventional method using purified parental mAbs. BsAbs prepared by two different routes showed indistinguishable characteristics by SDS capillary electrophoresis, analytical size exclusion, and cation exchange chromatography. This alternative method significantly shortened timelines and reduced resources required for bsAb generation, providing an improved process with potential benefits in large-scale bsAb preparation, as well as for HTP small-scale bsAb matrix selection.

  13. Purification and characterization of Fab fragments with rapid reaction kinetics against myoglobin. (United States)

    Song, Hyung-Nam; Kim, Dong-Hyung; Park, Sung-Goo; Lee, Myung Kyu; Paek, Se-Hwan; Woo, Eui-Jeon


    Myoglobin is an early biomarker for acute myocardial infarction. Recently, we isolated the antibody IgG-Myo2-7ds, which exhibits unique rapid reaction kinetics toward human myoglobin antigen. Antibodies with rapid dissociation kinetics are thought to be premature IgG forms that are produced during the early stage of in vivo immunization. In the present study, we identified the epitope region of the IgG-Myo2-7ds antibody to be the C-terminal region of myoglobin, which corresponds to 144-154 aa. The Fab fragment was directly purified by papain cleavage and protein G affinity chromatography and demonstrated kinetics of an association constant of 4.02 × 10(4) M(-1) s(-1) and a dissociation constant of 2.28 × 10(-2) s(-1), which retained the unique reaction kinetics of intact IgG-Myo2-7ds antibodies. Because a rapid dissociation antibody can be utilized for antibody recycling, the results from this study would provide a platform for the development of antibody engineering in potential diagnostic areas such as a continuous monitoring system for heart disease.

  14. Integration of the APC framework with AMD's Fab25 factory system (United States)

    Bushman, Scott; Campbell, William J.; Miller, Michael L.


    This paper discusses the integration and development of advanced process control technologies with AMD's Fab25 factory systems using the Advance Process Control Framework. The Framework is an open software architecture that allows the integration of existing factory systems, such as the manufacturing execution systems, configurable equipment interfaces, recipe management systems, metrology tools, process tools, and add-on sensors, into a system which provides advanced process control specific functionality. The Advanced Process Control Framework project was formulated to enable effective integration of Advanced Process Control applications into a semiconductor facility to improve manufacturing productivity and product yields. The main communication link between the factory system and the Framework is the Configurable Equipment Interface. It interfaces through a specialized component in the framework, the Machine Interface, which converts the factory system communication protocol, ISIS, to the Framework protocol, CORBA. The Framework is a distributed architecture that uses CORBA as a communication protocol between specialized components. A generalized example of how the Framework is integrated into the semiconductor facility is provided, as well as a description of the overall architecture used for process control strategy development. The main development language, Tcl/Tk, provides for increased development and deployment over traditional coding methods.

  15. Family-Group Names In Coleoptera (Insecta

    Directory of Open Access Journals (Sweden)

    Patrice Bouchard


    Full Text Available We synthesize data on all known extant and fossil Coleoptera family-group names for the first time. A catalogue of 4887 family-group names (124 fossil, 4763 extant based on 4707 distinct genera in Coleoptera is given. A total of 4492 names are available, 183 of which are permanently invalid because they are based on a preoccupied or a suppressed type genus. Names are listed in a classification framework. We recognize as valid 24 superfamilies, 211 families, 541 subfamilies, 1663 tribes and 740 subtribes. For each name, the original spelling, author, year of publication, page number, correct stem and type genus are included. The original spelling and availability of each name were checked from primary literature. A list of necessary changes due to Priority and Homonymy problems, and actions taken, is given. Current usage of names was conserved, whenever possible, to promote stability of the classification. New synonymies (family-group names followed by genus-group names: Agronomina Gistel, 1848 syn. n. of Amarina Zimmermann, 1832 (Carabidae, Hylepnigalioini Gistel, 1856 syn. n. of Melandryini Leach, 1815 (Melandryidae, Polycystophoridae Gistel, 1856 syn. n. of Malachiinae Fleming, 1821 (Melyridae, Sclerasteinae Gistel, 1856 syn. n. of Ptilininae Shuckard, 1839 (Ptinidae, Phloeonomini Ádám, 2001 syn. n. of Omaliini MacLeay, 1825 (Staphylinidae, Sepedophilini Ádám, 2001 syn. n. of Tachyporini MacLeay, 1825 (Staphylinidae, Phibalini Gistel, 1856 syn. n. of Cteniopodini Solier, 1835 (Tenebrionidae; Agronoma Gistel 1848 (type species Carabus familiaris Duftschmid, 1812, designated herein syn. n. of Amara Bonelli, 1810 (Carabidae, Hylepnigalio Gistel, 1856 (type species Chrysomela caraboides Linnaeus, 1760, by monotypy syn. n. of Melandrya Fabricius, 1801 (Melandryidae, Polycystophorus Gistel, 1856 (type species Cantharis aeneus Linnaeus, 1758, designated herein syn. n. of Malachius Fabricius, 1775 (Melyridae, Sclerastes Gistel, 1856 (type species

  16. A kit to prepare (111)In-DTPA-trastuzumab (Herceptin) Fab fragments injection under GMP conditions for imaging or radioimmunoguided surgery of HER2-positive breast cancer. (United States)

    Scollard, Deborah A; Chan, Conrad; Holloway, Claire M B; Reilly, Raymond M


    The human epidermal growth factor receptor-2 (HER2) gene is amplified in 25% of invasive breast cancers, and receptor overexpression has been noted in up to 60% of early stages of the disease [ductal carcinoma in situ (DCIS)]. Preclinical studies have revealed high tumor/blood ratios (>27:1) for (111)In-labeled Fab fragments of the HER2 monoclonal antibody, trastuzumab (Herceptin) ((111)In-DTPA-trastuzumab Fab) at 72 h pi in athymic mice bearing subcutaneous human breast cancer xenografts. Our aim in this study was to formulate a kit for preparation of (111)In-DTPA-trastuzumab Fab injection under good manufacturing practice (GMP) conditions suitable for human administration in a Phase I clinical trial of imaging and radioimmunoguided surgery (RIGS) of HER2-positive breast cancer. Fab fragments were produced by digestion of trastuzumab IgG (Herceptin) with immobilized papain for 20 h at 37°C. Fab fragments were purified by ultrafiltration, then reacted with a 10-fold molar excess of diethylenetriaminepentaacetic acid (DTPA) dianhydride. DTPA-Fab fragments were purified, then sterilized by filtration into unit dose glass vials (kits). Kits were tested against specifications for volume (0.9-1.1 ml), protein concentration (0.45-0.55 mg/ml), pH (5.5-6.5), DTPA substitution (0.5-4.0 mol DTPA/mol Fab), appearance (clear, colorless and particle free), labeling efficiency (≥ 85%), and sterility and apyrogenicity (USP XXXII). Immunoreactivity of (111)In-DTPA-trastuzumab Fab towards HER2 was measured by saturation radioligand binding assays using SKBR-3 human breast cancer cells (specifications: K(a) = 0.6-9.6 × 10(7) L/mol; B(max) = 0.6-10.4 × 10(6) sites/cell). (111)In-DTPA-trastuzumab Fab injection was prepared by adding 80-100 MBq of (111)InCl(3) to a single kit vial and incubating for 30 min at room temperature. (111)In-DTPA-trastuzumab Fab was assayed for the amount of radioactivity and tested for pH, radiochemical purity (RCP), appearance and sterility. Pure and

  17. A kit to prepare {sup 111}In-DTPA-trastuzumab (Herceptin) Fab fragments injection under GMP conditions for imaging or radioimmunoguided surgery of HER2-positive breast cancer

    Energy Technology Data Exchange (ETDEWEB)

    Scollard, Deborah A.; Chan, Conrad [Department of Pharmaceutical Sciences, University of Toronto, Toronto, ON, M5S 3M2 (Canada); Holloway, Claire M.B. [Department of Surgery, Sunnybrook Health Sciences Centre, Toronto, ON, M4N 1H1 (Canada); Reilly, Raymond M., E-mail: raymond.reilly@utoronto.c [Department of Pharmaceutical Sciences, University of Toronto, Toronto, ON, M5S 3M2 (Canada); Department of Medical Imaging, University of Toronto, Toronto, ON, M5S 3E2 (Canada); Toronto General Research Institute, University Health Network, Toronto, ON, M5G 2M9 (Canada)


    Introduction: The human epidermal growth factor receptor-2 (HER2) gene is amplified in 25% of invasive breast cancers, and receptor overexpression has been noted in up to 60% of early stages of the disease [ductal carcinoma in situ (DCIS)]. Preclinical studies have revealed high tumor/blood ratios (>27:1) for {sup 111}In-labeled Fab fragments of the HER2 monoclonal antibody, trastuzumab (Herceptin) ({sup 111}In-DTPA-trastuzumab Fab) at 72 h pi in athymic mice bearing subcutaneous human breast cancer xenografts. Our aim in this study was to formulate a kit for preparation of {sup 111}In-DTPA-trastuzumab Fab injection under good manufacturing practice (GMP) conditions suitable for human administration in a Phase I clinical trial of imaging and radioimmunoguided surgery (RIGS) of HER2-positive breast cancer. Methods: Fab fragments were produced by digestion of trastuzumab IgG (Herceptin) with immobilized papain for 20 h at 37{sup o}C. Fab fragments were purified by ultrafiltration, then reacted with a 10-fold molar excess of diethylenetriaminepentaacetic acid (DTPA) dianhydride. DTPA-Fab fragments were purified, then sterilized by filtration into unit dose glass vials (kits). Kits were tested against specifications for volume (0.9-1.1 ml), protein concentration (0.45-0.55 mg/ml), pH (5.5-6.5), DTPA substitution (0.5-4.0 mol DTPA/mol Fab), appearance (clear, colorless and particle free), labeling efficiency ({>=}85%), and sterility and apyrogenicity (USP XXXII). Immunoreactivity of {sup 111}In-DTPA-trastuzumab Fab towards HER2 was measured by saturation radioligand binding assays using SKBR-3 human breast cancer cells (specifications: K{sub a}=0.6-9.6x10{sup 7} L/mol; B{sub max}=0.6-10.4x10{sup 6} sites/cell). {sup 111}In-DTPA-trastuzumab Fab injection was prepared by adding 80-100 MBq of {sup 111}InCl{sub 3} to a single kit vial and incubating for 30 min at room temperature. {sup 111}In-DTPA-trastuzumab Fab was assayed for the amount of radioactivity and tested for p

  18. Targeting Mast Cells and Basophils with Anti-FcεRIα Fab-Conjugated Celastrol-Loaded Micelles Suppresses Allergic Inflammation. (United States)

    Peng, Xia; Wang, Juan; Li, Xianyang; Lin, Lihui; Xie, Guogang; Cui, Zelin; Li, Jia; Wang, Yuping; Li, Li


    Mast cells and basophils are effector cells in the pathophysiology of allergic diseases. Targeted elimination of these cells may be a promising strategy for the treatment of allergic disorders. Our present study aims at targeted delivery of anti-FcεRIα Fab-conjugated celastrol-loaded micelles toward FcεRIα receptors expressed on mast cells and basophils to have enhanced anti-allergic effect. To achieve this aim, we prepared celastrol-loaded (PEO-block-PPO-block-PEO, Pluronic) polymeric nanomicelles using thin-film hydration method. The anti-FcεRIα Fab Fragment was then conjugated to carboxyl groups on drug-loaded micelles via EDC amidation reaction. The anti-FcεRIα Fab-conjugated celastrol-loaded micelles revealed uniform particle size (93.43 ± 12.93 nm) with high loading percentage (21.2 ± 1.5% w/w). The image of micelles showed oval and rod like. The anti-FcεRIα Fab-conjugated micelles demonstrated enhanced cellular uptake and cytotoxity toward target KU812 cells than non-conjugated micelles in vitro. Furthermore, diffusion of the drug into the cells allowed an efficient induction of cell apoptosis. In mouse model of allergic asthma, treatment with anti-FcεRIα Fab-conjugated micelles increased lung accumulation of micelles, and significantly reduced OVA-sIgE, histamine and Th2 cytokines (IL-4, IL-5, TNF-α) levels, eosinophils infiltration and mucus production. In addition, in mouse model of passive cutaneous anaphylaxis, anti-FcεRIα Fab-conjugated celastrol-loaded micelles treatment significantly decreased extravasated evan's in the ear. These results indicate that anti-FcεRIα Fab-conjugated celastrol-loaded micelles can target and selectively kill mast cells and basophils which express FcεRIα, and may be efficient reagents for the treatment of allergic disorders and mast cell related diseases.

  19. Conjugation of 10 kDa Linear PEG onto Trastuzumab Fab' Is Sufficient to Significantly Enhance Lymphatic Exposure while Preserving in Vitro Biological Activity. (United States)

    Chan, Linda J; Ascher, David B; Yadav, Rajbharan; Bulitta, Jürgen B; Williams, Charlotte C; Porter, Christopher J H; Landersdorfer, Cornelia B; Kaminskas, Lisa M


    The lymphatic system is a major conduit by which many diseases spread and proliferate. There is therefore increasing interest in promoting better lymphatic drug targeting. Further, antibody fragments such as Fabs have several advantages over full length monoclonal antibodies but are subject to rapid plasma clearance, which can limit the lymphatic exposure and activity of Fabs against lymph-resident diseases. This study therefore explored ideal PEGylation strategies to maximize biological activity and lymphatic exposure using trastuzumab Fab' as a model. Specifically, the Fab' was conjugated with single linear 10 or 40 kDa PEG chains at the hinge region. PEGylation led to a 3-4-fold reduction in binding affinity to HER2, but antiproliferative activity against HER2-expressing BT474 cells was preserved. Lymphatic pharmacokinetics were then examined in thoracic lymph duct cannulated rats after intravenous and subcutaneous dosing at 2 mg/kg, and the data were evaluated via population pharmacokinetic modeling. The Fab' displayed limited lymphatic exposure, but conjugation of 10 kDa PEG improved exposure by approximately 11- and 5-fold after intravenous (15% dose collected in thoracic lymph over 30 h) and subcutaneous (9%) administration, respectively. Increasing the molecular weight of the PEG to 40 kDa, however, had no significant impact on lymphatic exposure after intravenous (14%) administration and only doubled lymphatic exposure after subcutaneous administration (18%) when compared to 10 kDa PEG-Fab'. The data therefore suggests that minimal PEGylation has the potential to enhance the exposure and activity of Fab's against lymph-resident diseases, while no significant benefit is achieved with very large PEGs.

  20. A Human Anti-Toll Like Receptor 4 Fab Fragment Inhibits Lipopolysaccharide-Induced Pro-Inflammatory Cytokines Production in Macrophages. (United States)

    Wang, Maorong; Zheng, Wenkai; Zhu, Xuhui; Xu, Jing; Cai, Binggang; Zhang, Yiqing; Zheng, Feng; Zhou, Linfu; Yang, Zhiguo; Zhang, Xin; Wang, Changjun; Nie, Shinan; Zhu, Jin


    The results of clinical and experimental studies suggest that endotoxin/toll-like receptor 4 (TLR4)-mediated proinflammatory and profibrotic signaling activation is critical in the development of hepatic fibrosis. However, studies examining the role of specific TLR4 inhibitor are still lacking. The present study was aimed to prepare a human anti-TLR4 Fab fragment, named hTLR4-Fab01, and to explore its immune activity. We screened the positive clone of anti-human TLR4 phagemid from a human phage-display antibody library using recombinant TLR4 protein, which was used as template cDNA for the amplification of variable regions of the heavy (VH) chain and light chain (VL), then coupled with highly conserved regions of the heavy chain domain 1 (CH1) and the light chain (CL), respectively. Thus, the prokaryotic expression vector pETDuet-1 of hTLR4-Fab01 was constructed and transformed into Escherichia coli (E. coli) BL21. The characteristic of hTLR4-Fab01 was examined by SDS-PAGE, Western blotting, ELISA, affinity and kinetics assay. Further, our data demonstrate that hTLR4-Fab01 could specifically bind to TLR4, and its treatment obviously attenuated the proinflammatory effect, characterized by less LPS-induced TNF-α, IL-1, IL-6 and IL-8 production in human macrophages. In conclusion, we have successfully prepared the hTLR4-Fab01 with efficient activity for blocking LPS-induced proinflammatory cytokines production, suggesting that the hTLR4-Fab01 may be a potential candidate for the treatment of hepatic fibrosis.

  1. Improved renal clearance and tumor targeting of {sup 99m}Tc-labeled anti-Tac monoclonal antibody Fab by chemical modifications

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Meyoung-kon; Jeong, Hyeh-Jean; Kao, Chih-Hao K.; Yao, Zhengsheng; Paik, David S.; Pie, Jae Eun; Kobayashi, Hisataka; Waldmann, Thomas A.; Carrasquillo, Jorge A.; Paik, Chang H. E-mail:


    This study was undertaken to improve the renal clearance and tumor targeting properties of {sup 99m}Tc-labeled humanized anti-Tac (HuTac) monoclonal antibody Fab fragments using two chemical approaches: 1) labeling with a renal secretion agent {sup 99m}Tc-mercaptoacetyltriglycine (MAG3) and 2) lowering its isoelectric point (pI) by acylation. HuTac Fab (3.3 mg/mL) was reacted with a trifluorophenyl ester (TFP) of {sup 99m}Tc-MAG3 alone or was additionally reacted with TFP-glycolate to reduce the pI. In Balb/c mice, {sup 99m}Tc-MAG3-Fab (pI>9.3) rapidly accumulated in the kidneys (177% injected dose [ID]/g at 15 min) and then gradually cleared out of the kidneys. In contrast, the glycolation (pI 4.6{approx}6.6) drastically reduced the renal uptake (31% ID/g) and also the whole-body retention (82% ID vs 101% for the nonglycolated) at 15 min, indicating that the glycolated {sup 99m}Tc-MAG3-Fab (pI 4.6{approx}6.6) was rapidly excreted. The glycolated remained in the blood longer than the nonglycolated (1.2% vs 0.3% ID/g at 360 min), but this effect was less drastic than the effect shown on the renal uptake. In nude mice bearing receptor-positive (ATAC4) tumors, the glycolated {sup 99m}Tc-MAG3-Fab increased the peak tumor uptake to 14.8% ID/g from 8.3% ID/g for {sup 99m}Tc-MAG3-Fab, whereas the glycolation resulted in a drastic reduction of the renal uptake at 15 min. We demonstrated that the renal clearance and the tumor targeting of Fab could be optimized by chemical modifications.

  2. 具斑芒毛苣苔抑制α-葡萄糖苷酶活性成分研究%α-Glucosidase inhibitory activity of Aeschynanthus maculatus

    Institute of Scientific and Technical Information of China (English)

    田璞玉; 康文艺


    Objective: To study the inhibitory activity of Aeschynanthus maculatus on α-glucosidase. Method; The inhibilitory model of in vitro α-glucosidase was established. Active extracts of A. maculatus were isolated and identified bymultiple chromatographic methods, and their molecular structures were identifiied by spectral techniques. Result; Seven coumpounts were isolated from A. maculatus and isolated as lupeol(l) , stigmasterol(2) , ursolic acid(3) , stigmast-5 ,22( E)-diene-3β-ol(4) , β-daucosterol(5) , 3-hydro xy-12-taraxasten-28-oic-acid(6) and oleanic acid(7). Compounds 1 (IC50 25.41 mg · L-1 ) ,3( IC50 4. 42 mg · L-1),4(IC50 11.50 mg · L-1 ) ,6( IC50 14. 17 mg · L-1 ) and 7(IC5() 2. 88 mg · L-1 ) had higher inhibitory activities than that of acarbose( IC501 103.01 mg · L-1 ) as the positive control drug. Conclusion; Compound 1-7 were isolated from this plant for the first time. Compound 6 was isolated from Gesneriaceae family for the first time. Compound 7 was isolated from Aeschynanthus genus for the first time.%目的:研究具斑芒毛苣苔抑制α-葡萄糖苷酶的活性成分.方法:用体外α-葡萄糖苷酶抑制模型筛选;采用各种色谱法对高活性部位分离,运用多种波谱技术鉴定结构.结果:从具斑芒毛苣苔分离得到7个化合物,分别为羽扇豆醇(1),豆甾醇(2),熊果酸(3),豆甾-5,22(E)-二烯-3β-醇(4)和β-胡萝卜苷(5),3-hydroxy-12-taraxasten-28-oic-acid(6),齐墩果酸(7).化合物1(IC50 25.41 mg·L-1),3(IC50 4.42 mg·L-1),4(IC50 11.50 mg·L-1),6(IC50 14.17 mg·L-1)和7(IC50 2.88mg·L-1)的体外抑制α-葡萄糖苷酶活性高于阳性对照药物acarbose(IC50 1 103.01 mg· L-1).结论:化合物1~7为首次从该植物中分离得到,6为首次从该科中分离得到,7为首次从该属植物中分离得到.

  3. Parasitylenchus sp. (Tylenchomorpha: Allantonematidae) parasitizing field populations of Harmonia axyridis (Coleoptera: Coccinellidae)

    DEFF Research Database (Denmark)

    Harding, Susanne; Poinar, George O. Jr.; Dimitrova, Desislava V.;


    Adults of the invasive harlequin ladybird, Harmonia axyridis (Coleoptera: Coccinellidae), were found to be parasitized by nematodes (Tylenchomorpha: Allantonematidae) in Denmark. The nematodes were identified as Parasitylenchus sp. Major morphological characters of the nematodes did not differ...

  4. Illustrated guide to the emerald ash borer, Agrilus planipennis Fairmaire and related species (Coleoptera, Buprestidae) (United States)

    The 33 species of Agrilus (Coleoptera: Buprestidae) hypothesized to be most closely related to Agrilus planipennis Fairmaire (the emerald ash borer), are described and illustrated. Morphology (adults and immatures), biology, distribution, detailed taxonomic history and systematics are presented fo...

  5. Apostasimerini (Coleoptera: Curculionidae: Baridinae). Rectification of authorship, year of publication, rank, and taxa included (United States)

    The following nomenclatural changes are proposed in the Coleoptera, Curculionidae: the author of Apostasimerini is Schoenherr (1844), not Lacordaire (1866); Madopterini Lacordaire, 1866 is demoted to subtribe of Apostasimerini; Erirhinus mourei Bondar, 1943 is a new synonym of Apostasimerus serriros...

  6. An unusual new species of Micraspis Chevrolat (Coleoptera: Coccinellidae) from northeastern India. (United States)

    Poorani, J


    Micraspispusillus sp. n. (Coleoptera: Coccinellidae) is described and illustrated from the northeastern region of India. It is unusual in possessing very large eye canthus and is the smallest species of the genus known from India so far.

  7. First record of Molorchus minor minor (Linnaeus (Coleoptera, Cerambycidae in Brazil

    Directory of Open Access Journals (Sweden)

    Ubirajara R. Martins


    Full Text Available Molorchus minor minor (Linnaeus (Coleoptera, Cerambycidae is recorded for the first time in Brazil (Bahia. It was originally described from Europe and is currently widely distributed in that continent and Asia.

  8. A new species of Scymnobius Casey (Coleoptera, Coccinellidae, Scymnini from Pernambuco, Brazil

    Directory of Open Access Journals (Sweden)

    José Adriano Giorgi


    Full Text Available A new species of Scymnobius Casey (Coleoptera, Coccinellidae, Scymnini from Pernambuco, Brazil. Scymnobius pernambucensis sp. nov. from Pernambuco, Brazil, is described and illustrated. This is the third species of this genus recorded from Brazil.

  9. A new species of the genus Falsoibidion Pic (Coleoptera, Cerambycidae) from Korea. (United States)

    Lee, Seunghyun; Lee, Seunghwan


    A new species of the genus Falsoibidion Pic, 1922 (Coleoptera, Cerambycidae, Cerambycinae, Callidiopini) from Korea is described. Habitus and genitalia of male and female of the new species are illustrated.

  10. Methods for assessing infestations of sunflower stem weevil (Coleoptera: Curculionidae) in sunflower stems (United States)

    The sunflower stem weevil, Cylindrocopturus adspersus LeConte (Coleoptera: Curculionidae), reduces sunflower, Helianthus annuus L. (Asteraceae), yields by spreading pathogens, damaging vascular tissues, and promoting lodging of sunflower plants. To assess weevil populations for host plant resistanc...

  11. Intracellular routing in breast cancer cells of streptavidin-conjugated trastuzumab Fab fragments linked to biotinylated doxorubicin-functionalized metal chelating polymers. (United States)

    Liu, Peng; Cai, Zhongli; Kang, Jae W; Boyle, Amanda J; Adams, Jarret; Lu, Yijie; Ngo Ndjock Mbong, Ghislaine; Sidhu, Sachdev; Reilly, Raymond M; Winnik, Mitchell A


    We describe the synthesis of a heterotelechelic metal-chelating polymer (Bi-MCP-Dox), a polyacrylamide with a number average degree of polymerization DPn = 50 (PDI = 1.2), with biotin (Bi) and doxorubicin (Dox) as functional chain ends and diethylenetriaminepentaacetic acid (DTPA) pendant groups as the binding sites for metal ions. We compared its behavior in cell-uptake experiments with a similar polymer (Bi-MCP) without Dox. These MCPs were complexed with trastuzumab Fab (tmFab) fragments covalently linked to streptavidin (SAv) to form tmFab-SAv-Bi-MCP-Dox and tmFab-SAv-Bi-MCP via the strong affinity between Bi and SAv. tmFab targets human epidermal growth factor receptor-2 (HER2), which is overexpressed on certain human breast cancer cells. Surface plasmon resonance (SPR) experiments with the extracellular domain (ECD) of HER2 showed that incorporation of the MCPs in these complexes had no significant effect on the association or dissociation rate with the HER2 ECD and the dissociation constants. The tmFab-complexed MCPs were subsequently labeled with (111)In (an Auger electron emitting radionuclide). Auger electrons can cause lethal DNA double strand breaks (DSBs) but only if they are emitted intracellularly and especially, in close proximity to the nucleus. To evaluate the cellular and nuclear uptake of tmFab-SAv-Bi-MCP-Dox, we incubated HER2+ SK-BR-3 human breast cancer cells with the complexes saturated with stable In(3+) and visualized their distribution by confocal fluorescence microscopy, monitoring the fluorescence of Dox. In parallel, we carried out cell fractionation studies on tmFab-SAv-Bi-MCP-Dox and on tmFab-SAv-Bi-MCP labeled with (111)In. Both radiolabeled complexes showed cell internalization and nuclear localization. We conclude that metal-chelating polymers with this composition appear to encourage internalization, nuclear uptake, and chromatin (DNA) binding of trastuzumab fragments modified with streptavidin in human breast cancer cells

  12. Specific Conjugation of the Hinge Region for Homogeneous Preparation of Antibody Fragment-Drug Conjugate: A Case Study for Doxorubicin-PEG-anti-CD20 Fab' Synthesis. (United States)

    Zhou, Zhan; Zhang, Jing; Zhang, Yan; Ma, Guanghui; Su, Zhiguo


    Conventional preparation strategies for antibody-drug conjugates (ADCs) result in heterogeneous products with various molecular sizes and species. In this study, we developed a homogeneous preparation strategy by site-specific conjugation of the anticancer drug with an antibody fragment. The model drug doxorubicin (DOX) was coupled to the Fab' fragment of anti-CD20 IgG at its permissive sites through a heterotelechelic PEG linker, generating an antibody fragment-drug conjugate (AFDC). Anti-CD20 IgG was digested and reduced specifically with β-mercaptoethylamine to generate the Fab' fragment with two free mercapto groups in its hinge region. Meanwhile, DOX was conjugated with α-succinimidylsuccinate ω-maleimide polyethylene glycol (NHS-PEG-MAL) to form MAL-PEG-DOX, which was subsequently linked to the free mercapto containing Fab' fragment to form a Fab'-PEG-DOX conjugate. The dual site-specific bioconjugation was achieved through the combination of highly selective reduction of IgG and introduction of heterotelechelic PEG linker. The resulting AFDC provides an utterly homogeneous product, with a definite ratio of one fragment to two drugs. Laser confocal microscopy and cell ELISA revealed that the AFDC could accumulate in the antigen-positive Daudi tumor cell. In addition, the Fab'-PEG-DOX retained appreciable targeting ability and improved antitumor activity, demonstrating an excellent therapeutic effect on the lymphoma mice model for better cure rate and significantly reduced side effects.

  13. Structure of the Francisella tularensis enoyl-acyl carrier protein reductase (FabI) in complex with NAD[superscript +] and triclosan

    Energy Technology Data Exchange (ETDEWEB)

    Mehboob, Shahila; Truong, Kent; Santarsiero, Bernard D.; Johnson, Michael E. (UIC)


    Enoyl-acyl carrier protein reductase (FabI) catalyzes the last rate-limiting step in the elongation cycle of the fatty-acid biosynthesis pathway and has been validated as a potential antimicrobial drug target in Francisella tularensis. The development of new antibiotic therapies is important both to combat potential drug-resistant bioweapons and to address the broader societal problem of increasing antibiotic resistance among many pathogenic bacteria. The crystal structure of FabI from F. tularensis (FtuFabI) in complex with the inhibitor triclosan and the cofactor NAD{sup +} has been solved to a resolution of 2.1 {angstrom}. Triclosan is known to effectively inhibit FabI from different organisms. Precise characterization of the mode of triclosan binding is required to develop highly specific inhibitors. Comparison of our structure with the previously determined FtuFabI structure (PDB code 2jjy) which is bound to only NAD{sup +} reveals the conformation of the substrate-binding loop, electron density for which was missing in the earlier structure, and demonstrates a shift in the conformation of the NAD{sup +} cofactor. This shift in the position of the phosphate groups allows more room in the active site for substrate or inhibitor to bind and be better accommodated. This information will be crucial for virtual screening studies to identify novel scaffolds for development into new active inhibitors.

  14. Imaging of HER2/neu-positive BT-474 human breast cancer xenografts in athymic mice using {sup 111}In-trastuzumab (Herceptin) Fab fragments

    Energy Technology Data Exchange (ETDEWEB)

    Tang Ying [Division of Nuclear Medicine, University Health Network, Toronto, Ontario, M5G 2C4 (Canada); Department of Pharmaceutical Sciences, University of Toronto, Toronto, Ontario, M5S 2S2 (Canada); Wang, Judy [Division of Nuclear Medicine, University Health Network, Toronto, Ontario, M5G 2C4 (Canada); Scollard, Deborah A. [Division of Nuclear Medicine, University Health Network, Toronto, Ontario, M5G 2C4 (Canada); Mondal, Hridya [Division of Nuclear Medicine, University Health Network, Toronto, Ontario, M5G 2C4 (Canada); Holloway, Claire [Sunnybrook and Women' s College Health Sciences Center, Toronto, Ontario, M4N 3M5 (Canada); Kahn, Harriette J. [Sunnybrook and Women' s College Health Sciences Center, Toronto, Ontario, M4N 3M5 (Canada); Reilly, Raymond M. [Division of Nuclear Medicine, University Health Network, Toronto, Ontario, M5G 2C4 (Canada) and Department of Pharmaceutical Sciences, University of Toronto, Toronto, Ontario, M5S 2S2 (Canada) and Department of Medical Imaging, University of Toronto, Toronto, Ontario, M5S 3E2 (Canada)]. E-mail:


    Trastuzumab (Herceptin) Fab were prepared by digestion of intact IgG with immobilized papain, derivatized with diethylenetriaminepentaacetic acid (DTPA) and radiolabeled with {sup 111}In. The dissociation constant (K{sub d}) for binding of Fab to HER2/neu-positive SK-BR-3 human breast cancer cells was two- to threefold higher than for intact IgG (14-36 vs. 8-14 nM). The binding affinity was not significantly decreased after DTPA derivatization (K{sub d}=47 nM). {sup 111}In-trastuzumab Fab localized specifically in HER2/neu-positive BT-474 human breast cancer xenografts in athymic mice with tumor uptake of 7.8{+-}0.7% injected dose (ID)/g and tumor/blood ratio of 25.2{+-}1.6 at 72 h postinjection compared with 2.7{+-}0.7% ID/g and 7.0{+-}0.9 for {sup 111}In-HuM195 anti-CD33 Fab (significantly different, P<.001). Small (3-5 mm in diameter) BT-474 tumors were imaged with {sup 111}In-trastuzumab Fab as early as 24 h postinjection.

  15. Deletion of the β-acetoacetyl synthase FabY in Pseudomonas aeruginosa induces hypoacylation of lipopolysaccharide and increases antimicrobial susceptibility. (United States)

    Six, David A; Yuan, Yanqiu; Leeds, Jennifer A; Meredith, Timothy C


    The β-acetoacetyl-acyl carrier protein synthase FabY is a key enzyme in the initiation of fatty acid biosynthesis in Pseudomonas aeruginosa. Deletion of fabY results in an increased susceptibility of P. aeruginosa in vitro to a number of antibiotics, including vancomycin and cephalosporins. Because antibiotic susceptibility can be influenced by changes in membrane lipid composition, we determined the total fatty acid profile of the ΔfabY mutant, which suggested alterations in the lipid A region of the lipopolysaccharide. The majority of lipid A species in the ΔfabY mutant lacked a single secondary lauroyl group, resulting in hypoacylated lipid A. Adding exogenous fatty acids to the growth media restored the wild-type antibiotic susceptibility profile and the wild-type lipid A fatty acid profile. We suggest that incorporation of hypoacylated lipid A species into the outer membrane contributes to the shift in the antibiotic susceptibility profile of the ΔfabY mutant.

  16. Structural and biophysical characterization of an epitope-specific engineered Fab fragment and complexation with membrane proteins: implications for co-crystallization. (United States)

    Johnson, Jennifer L; Entzminger, Kevin C; Hyun, Jeongmin; Kalyoncu, Sibel; Heaner, David P; Morales, Ivan A; Sheppard, Aly; Gumbart, James C; Maynard, Jennifer A; Lieberman, Raquel L


    Crystallization chaperones are attracting increasing interest as a route to crystal growth and structure elucidation of difficult targets such as membrane proteins. While strategies to date have typically employed protein-specific chaperones, a peptide-specific chaperone to crystallize multiple cognate peptide epitope-containing client proteins is envisioned. This would eliminate the target-specific chaperone-production step and streamline the co-crystallization process. Previously, protein engineering and directed evolution were used to generate a single-chain variable (scFv) antibody fragment with affinity for the peptide sequence EYMPME (scFv/EE). This report details the conversion of scFv/EE to an anti-EE Fab format (Fab/EE) followed by its biophysical characterization. The addition of constant chains increased the overall stability and had a negligible impact on the antigen affinity. The 2.0 Å resolution crystal structure of Fab/EE reveals contacts with larger surface areas than those of scFv/EE. Surface plasmon resonance, an enzyme-linked immunosorbent assay, and size-exclusion chromatography were used to assess Fab/EE binding to EE-tagged soluble and membrane test proteins: namely, the β-barrel outer membrane protein intimin and α-helical A2a G protein-coupled receptor (A2aR). Molecular-dynamics simulation of the intimin constructs with and without Fab/EE provides insight into the energetic complexities of the co-crystallization approach.

  17. Label-free Fab and Fc affinity/avidity profiling of the antibody complex half-life for polyclonal and monoclonal efficacy screening. (United States)

    Read, Thomas; Olkhov, Rouslan V; Williamson, E Diane; Shaw, Andrew M


    A unified approach to affinity screening for Fab and Fc interactions of an antibody for its antigen and FcγR receptor has been developed. An antigen array is used for the Fab affinity and cross-reactivity screening and protein A/G proxy is the FcγR receptor. The affinities are derived using a simple 1:1 binding model with a consistent error analysis. The association and dissociation kinetics are measured over optimised times for accurate determination. The Fab/Fc affinities are derived for ten antibodies: mAb-actin (mouse), pAb-BSA (sheep), pAb-collagen V (rabbit), pAb-CRP (goat), mAb-F1 (mouse), mAbs (mouse) 7.3, 12.3, 29.3, 36.3 and 46.3 raised against LcrV in Yersinia pestis. The rate of the dissociation of antigen-antibody complexes relates directly to their immunological function as does the Fc-FcγR complex and a new half-life plot has been defined with a Fab/Fc half-life range of 17-470 min. The upper half-life value points to surface avidity. Two antibodies that are protective as an immunotherapy define a Fab half-life >250 min and an Fc half-life >50 min as characteristics of ideal interactions which can form the basis of an antibody screen for immunotherapy.

  18. Construction and diversification of yeast cell surface displayed libraries by yeast mating: application to the affinity maturation of Fab antibody fragments. (United States)

    Blaise, Lydia; Wehnert, Anita; Steukers, Mieke P G; van den Beucken, Twan; Hoogenboom, Hennie R; Hufton, Simon E


    Yeast display is a powerful technology for the affinity maturation of human antibody fragments. However, the technology thus far has been limited by the size of antibody libraries that can be generated, as using current transformation protocols libraries of only between 10(6) and 10(7) are typically possible. We have recently shown that Fab antibodies can be displayed on the cell surface of Saccharomyces cerevisiae [van den Beucken, T., Pieters, H., Steukers, M., van der Vaart, M., Ladner, R.C., Hoogenboom, H.R., Hufton, S.E., 2003. Affinity maturation of Fab antibody fragments by fluorescent-activated cell sorting of yeast-displayed libraries. FEBS Lett. 546, 288-294]. This discovery and the knowledge that Fab antibodies are heterodimeric suggest that independent repertoires of heavy chain (HC) and light chain (LC) can be constructed in haploid yeast strains of opposite mating type. These separate repertoires can then be combined by highly efficient yeast mating. Using this approach, we have rapidly generated a naive human Fab yeast display library of over 10(9) clones. In addition, utilizing error-prone polymerase chain reaction, we have diversified Fab sequences and generated combinatorial and hierarchical chain shuffled libraries with complexities of up to 5 x 10(9) clones. These libraries have been selected for higher affinity using a repeating process of mating-driven chain shuffling and flow cytometric sorting.

  19. Best-practice evaluation-methods for wafer-fab photomask-requalification inspection tools (United States)

    Cho, Chan Seob; Mungmode, Ashish; Taylor, Ron; Cho, David; Koh, Hui Peng


    Requalifying semiconductor photomasks remains critically important and is increasingly challenging for 20nm and 14nm node logic reticles. Patterns are becoming more complex on the photomask, and defect sensitivity requirements are more stringent than ever before. Reticle inspection tools are equally important for effective process development and the successful ramp and sustained yield for high volume manufacturing. The inspection stages considered were: incoming inspection to match with Mask Shop Outgoing result and to detect defects generated during transport; requalification by routine cycle inspection to detect Haze and any other defects; and inspection by in-house or Mask shop at the post cleaning. There are many critical capability and capacity factors for the decision for best inspection tool and strategy for high volume manufacturing, especially objective Lens NA, wavelength, power, pixel size, throughput, full-automation inspection linked with Overhead Transport, algorithm application, engineering application function, and inspection of PSM and OMOG . These tools are expensive but deliver differentiated value in terms of performance and throughput as well as extendibility. Performing a thorough evaluation and making a technically sound choice which explores these many factors is critical for success of a fab. This paper examines the methodology for evaluating two different photomask inspection tools. The focus is on ensuring production worthiness on real and advanced product photomasks requiring accurate evaluation of sensitivity, throughput, data analysis function and engineering work function on those product photomasks. Photomasks used for data collection are production reticles, PDM(Program defect Mask), SiN spray defect Reticle which is described that evaluates how the tools would perform on a contaminated plate.

  20. Distribuição e alimentação de duas espécies simpátricas de piranhas Serrasalmus maculatus e Pygocentrus nattereri (Characidae, Serrasalminae do rio Ibicuí, Rio Grande do Sul, Brasil Distribution and feeding of two sympatric species of piranhas Serrasalmus maculatus and Pygocentrus nattereri (Characidae, Serrasalminae of the Ibicuí river, State of Rio Grande do Sul, Brazil

    Directory of Open Access Journals (Sweden)

    Everton R. Behr


    Full Text Available Duas espécies de piranhas ocorrem no rio Ibicuí, um dos principais afluentes do rio Uruguai. Com o objetivo de analisar a distribuição e a alimentação destas espécies, foram realizadas coletas de dezembro de 1999 a janeiro de 2002, utilizando-se diferentes artes de pesca em três pontos do rio, sendo que cada ponto foi amostrado em dois ambientes (lêntico e lótico. Foram capturados 203 indivíduos de Serrasalmus maculatus Kner, 1858 e 86 de Pygocentrus nattereri Kner, 1858, sendo que a maior captura de ambas as espécies ocorreu no ambiente lêntico e próximo ao rio Uruguai, onde P. nattereri parece estar mais concentrada. O número de exemplares capturados por horário não apresentou variações significativas em P. nattereri enquanto que S. maculatus apresentou a maior captura na revisão da meia-noite. Juvenis de S. maculatus (2-4 cm de comprimento padrão consumiram preferencialmente nadadeiras e insetos. Nas demais classes de tamanho de ambas as espécies, restos de peixes foi o alimento predominante, havendo sobreposição alimentar intra e interespecífica nas maiores classes (8-16 cm e >16 cm de comprimento padrão. Não foram constatadas diferenças no índice de repleção entre os horários e os ambientes para as duas espécies.Two species of piranhas occur in the Ibicuí River, one of the main tributaries of the Uruguay River in southern Brazil, Serrasalmus maculatus Kner, 1858 and Pygocentrus nattereri Kner, 1858. In order to analyze the distribution and feeding habits of these species we conducted collecting expeditions from December 1999 to January 2002, using a variety of fishing gears in three sites along the river, both in lentic and lotic habitats. A total of 203 individuals of S. maculatus and 86 of P. nattereri were caught, mostly in lentic habitats and closer to the Uruguay River, were P. nattereri seems to be more concentrated. The number of individuals captured at different times did not show variation in P

  1. Ergasilus chelangulatus sp. nov. (Copepoda: Ergasilidae a branchial parasite of the freshwater catfish, Pimelodus maculatus from the upper São Francisco River, Brazil Ergasilus chelangulatus sp. nov. (Copepoda: Ergasilidae parasito das brânquias do "mandi", Pimelodus maculatus do alto rio São Francisco, Brasil

    Directory of Open Access Journals (Sweden)

    Vernon E. Thatcher


    Full Text Available Ergasilus chelangulatus sp. nov. is described from the freshwater fish, Pimelodus maculatus Lacépède, 1803 (Siluriformes: Pimelodidae of the upper São Francisco River, Minas Gerais State, Brazil. The new species is based on female specimens and the male is unknown. This species has a serrate seta on exopod one and a two-segmented first endopod, as do most Amazonian species of this genus. The body is elongate and produced anteriorly. The antennae are elongate and have prominent sensilla on segments two and three. Ergasilus chelangulatus sp. nov. can be distinguished from all known species of Ergasilus by the fourth antennal segment (claw which is bent at nearly a right angle.Ergasilus chelangulatus sp. nov. é descrita do peixe de água doce, Pimelodus maculatus Lacépède, 1803 (Siluriformes: Pimelodidae do alto rio São Francisco, Minas Gerais, Brasil. A nova espécie está baseada em espécimes fêmeas e o macho é desconhecido. Esta espécie tem uma seta serrilhada no primeiro exopodito e tem o primeiro endopodito bi-segmentado, como a maioria das espécies amazônicas deste gênero. O corpo é alongado e projetado anteriormente. As antenas são alongadas com sensila proeminente no segundo e no terceiro segmentos. Ergasilus chelangulatus sp. nov. é distinta de todas as outras espécies conhecidas de Ergasilus por ter o quarto segmento antenal (garra dobrado em ângulo quase reto.

  2. Passalidae (Coleoptera: Scarabaeoidea) of the Greater and Lesser Antilles. (United States)

    Jiménez-Ferbans, Larry; Reyes-Castillo, Pedro; Schuster, Jack C


    We present a synthesis of the state of knowledge concerning the species of Passalidae (Coleoptera) of the West Indies and we present a key to the species. The recently described genus Antillanax Boucher renders the subgenus Passalus (Pertinax) Kaup paraphyletic, therefore we place Antillanax in synonymy with Passalus (Pertinax) and we propose a new combination for Passalus (Pertinax) doesburgi (Boucher). The island richest in species is Hispaniola, with five species, three of them endemic. Excluding Trinidad and Tobago, the passalid fauna of the West Indies comprises 13 species; this is low richness, but with high endemism (50%), especially for the Greater Antilles.

  3. Revision of the genus Altitatiayus Weinreich(Coleoptera, Lucanidae, Lucaninae

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    Paschoal C. Grossi


    Full Text Available Revision of the genus Altitatiayus Weinreich (Coleoptera, Lucanidae, Lucaninae. The South American genus Altitatiayus Weinreich is revised and now includes six species, A. rotundatus (Boileau, A. ruficollis (Lüderwaldt, A. godinhorum (Bomans & Arnaud, A. dulceae (Bomans & Arnaud, A. trifurcatus (Grossi & Racca-Filho and A. koikei sp. nov. (Minas Gerais, Brazil. All species are described and illustrated. For the first time male and female genitalia are illustrated for five species and observations on the behavior of two species are included.

  4. The tribe Phanaeini (Coleoptera: Scarabaeidae: Scarabaeinae in Peru

    Directory of Open Access Journals (Sweden)

    Luis Figueroa


    Full Text Available We examine the occurrence of the tribe Phanaeini (Coleoptera: Scarabaeidae: Scarabaeinae in Peru based on the collection at Museo de Historia Natural of the Universidad Nacional Mayor de San Marcos and on data provided in literature. Each species is presented with citations of its diagnosis, distribution and related comments. Peruvian Phanaeini includes 30 species in nine genera: Coprophanaeus, Dendropaemon, Gromphas, Oruscatus, Oxysternon, Phanaeus, Sulcophanaeus, Tetramereia and Megatharsis. Oruscatus davus is the only species distributed in the high Andes; Phanaeus lunaris and P. achilles occur in the northern arid zone shared by Peru and Ecuador; the remaining species are Amazonian.

  5. Revision of the genus Endochilus Weise (Coleoptera: Coccinellidae: Chilocorini). (United States)

    Łączyński, Piotr; Tomaszewska, Wioletta


    The members of the endemic African genus Endochilus Weise, 1898 (Coleoptera: Coccinellidae: Chilocorini) are redescribed, diagnosed, and illustrated. Lectotypes are designated for Endochilus compater Weise, Endochilus minor Weise, Endochilus plagiatus Sicard, Endochilus rubicundus Weise, and Endochilus styx Sicard. One new species is described: Endochilus abdominalis sp nov. Notes on the genus and nomenclatural history for each species are provided. A key for identification of all species is presented. Adult characters concerning similarities of Endochilus to other genera of African Chilocorini are discussed. This is an open access paper. We use the Creative Commons Attribution 3.0 license that permits unrestricted use, provided that the paper is properly attributed.

  6. Review of the tribe Hyperaspidini Mulsant (Coleoptera: Coccinellidae) from Iran. (United States)

    Biranvand, Amir; Tomaszewska, Wioletta; Nedvěd, Oldřich; Khormizi, Mehdi Zare; Nicolas, Vincent; Canepari, Claudio; Shakarami, Jahanshir; Fekrat, Lida; Fürsch, Helmut


    The Iranian species of the tribe Hyperaspidini Mulsant, 1846 (Coleoptera: Coccinellidae) are reviewed. The current list includes 12 species, all placed in a single genus Hyperaspis Chevrolat, 1836. Hyperapsis asiatica Lewis, 1896 and H. pumila Mulsant, 1850 are excluded from the Iranian list of Coccinellidae. Diagnoses of the tribe Hyperaspidini and the genus Hyperaspis are given. Images of adult beetles and diagnostic characters of the male genitalia of all species distributed in Iran are shown. A key to identification of the species is presented. Distribution records are provided for each species along with information on host plants and prey species when available.

  7. An annotated catalogue of the Buprestidae of Iran (Coleoptera: Buprestoidea). (United States)

    Ghahari, Hassan; Volkovitsh, Mark G; Bellamy, Charles L


    An annotated taxonomic catalogue of the jewel beetles (Coleoptera: Buprestidae) of Iran is given. Original descriptions and recent revisionary or catalogue data are included along with the distribution, both within and outside of Iran, ecological data and host plant associations, junior synonyms, and comments. A complete bibliography completes the catalogue. In total 428 species and 52 subspecies of jewel beetles belonging to 6 subfamilies (Julodinae, Polycestinae, Galbellinae, Chrysochroinae, Buprestinae, and Agrilinae), 20 tribes, and 38 genera are known from Iran including doubtful records and 4 nomina nuda. It is likely that the number of jewel beetle species from Iran will be between 460-480 and possibly even more species.

  8. Fossil history of Mesozoic weevils (Coleoptera:Curculionoidea)

    Institute of Scientific and Technical Information of China (English)

    Andrei A.Legalov


    The first synopsis of Mesozoic weevils (Curculionoidea: Coleoptera) is presented.Changes of family,genera and species abundance during the Mesozoic revealed three distributional patterns.The Jurassic (Karatau) fauna was dominated by the Nemonychidae.During the Early Cretaceous (beginning at the Jurassic/Cretaceous border),the Ithyceridae was the prevalent group with a significant role played by the Nemonychidae.In the Late Cretaceous (Cenomanian and Turonian),the major groups were the Curculionidae and Brentidae.Obviously,the change of weevil fauna during this period was due to the expansion of the angiosperms,which provided multiple niches in their vegetative and reproductive organs for weevil development.

  9. Electrophysiological responses of chafer beetle, Holotrichia serrata (F. (Coleoptera: Scarabaeidae

    Directory of Open Access Journals (Sweden)

    Kesavan Subaharan


    Full Text Available The chafer beetle, Holotrichia serrata F. (Coleoptera: Scarabaeidae in its larval stage is a serious pest on sugarcane, vegetables, groundnut and coconut in many parts of India. The antennal response of adults to host volatiles and pheromone gland extracts was assessed by electroantennography. Among the preferred host of H. serrata, the volatiles from neem, Azadirachta indica A. Juss leaf extract elicited higher antennal response than gulmohar Delonix regia L. flowers and Ailanthus excelsa (Roxb leaf extracts. The order of response was the same irrespective of the sex. In general the antennal response to pheromone gland and host extracts was higher in males than in females.

  10. Determination of Coleoptera fauna on carcasses in Ankara province, Turkey. (United States)

    Ozdemir, Senem; Sert, Osman


    In this study, 40 species from Staphylinidae, Histeridae, Dermestidae, Silphidae, Nitidulidae and Cleridae families of Coleoptera which were found in 12 pig (Sus scrofa L.) carcasses were identified and recorded during a one-year period at the Hacettepe University Beytepe Campus located in Ankara, Turkey. According to the duration of their presence on the carcasses, 22 of these species were accepted to be important in decomposition. Their distribution over the months and the duration of their presence in the various decomposition stages over the seasons were determined.





    Através do uso de armadilhas "pitfall" iscadas com massa fecal fresca de bovinos, realizou-se a coleta de besouros coprófagos (Coleoptera; Scarabaeidae), durante o período de 15 de abril de 1995 a 17 de fevereiro de 1996, em área de pastagem ao lado de confinamento de bovinos, em Piracicaba, SP. Coletou-se um total de 11 espécies distribuidas nos gêneros Aphodius, Ataenius, Trichillum, Eurysternus, Dichotomius e Canthon. Os besouros coprófagos de comportamento endocoprídeo representaram 72,73...

  12. Screening of Human Antibody Fab Fragment against HBsAg and the Construction of its dsFv Form

    Directory of Open Access Journals (Sweden)

    Leili Jia, Jiyun Yu, Hongbin Song, Xuelin Liu, Weina Ma, Yuanyong Xu, Chuanfu Zhang, Shicun Dong, Qiao Li


    Full Text Available The objective of this study was to pursue the techniques involving the screening of the human antibody Fab fragment against hepatitis B virus surface antigen (HBsAg and the construction of its disulfide-stabilized Fv fragment (dsFv. The phage antibody Fab fragments against HBsAg were screened from the human combinatorial immunoglobulin library. Sequence analysis revealed that its heavy chain gene was complete, but the light chain gene was lost. To improve the affinity of the antibody by chain shuffling, a human antibody light chain gene repertoire was generated by reverse transcriptase-polymerase chain reaction (RT-PCR from the human peripheral blood lymphocytes. A phage antibody sub-library was then constructed by inserting the light chain gene repertoire into the phagmid that contained the Fd gene. Five clones with appreciably higher absorbance than that of the original clone were obtained, which indicated that the affinity of the light chain-shuffled phage antibodies was improved. Then, the mutated genes of dsFv against HBsAg were constructed by using PCR-based point mutagenesis method. Purified VH and VL proteins were folded into a 25-kDa protein, designated as anti-HBsAg dsFv. ELISA and competition ELISA revealed that the dsFv maintained the specificity of the Fab by binding to HBsAg, even through with a lower binding activity. These results have facilitated the undertaking of further functional analyses of the constructed dsFv, and may therefore provide an improved technique for the production and application of dsFvs against HBsAg.

  13. Structure of anti-FLAG M2 Fab domain and its use in the stabilization of engineered membrane proteins

    Energy Technology Data Exchange (ETDEWEB)

    Roosild, Tarmo P.; Castronovo, Samantha; Choe, Senyon, E-mail: [Structural Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, California 92037 (United States)


    The X-ray crystallographic analysis of anti-FLAG M2 Fab is reported and the implications of the structure on FLAG epitope binding are described as a first step in the development of a tool for the structural and biophysical study of membrane proteins. The inherent difficulties of stabilizing detergent-solubilized integral membrane proteins for biophysical or structural analysis demand the development of new methodologies to improve success rates. One proven strategy is the use of antibody fragments to increase the ‘soluble’ portion of any membrane protein, but this approach is limited by the difficulties and expense associated with producing monoclonal antibodies to an appropriate exposed epitope on the target protein. Here, the stabilization of a detergent-solubilized K{sup +} channel protein, KvPae, by engineering a FLAG-binding epitope into a known loop region of the protein and creating a complex with Fab fragments from commercially available anti-FLAG M2 monoclonal antibodies is reported. Although well diffracting crystals of the complex have not yet been obtained, during the course of crystallization trials the structure of the anti-FLAG M2 Fab domain was solved to 1.86 Å resolution. This structure, which should aid future structure-determination efforts using this approach by facilitating molecular-replacement phasing, reveals that the binding pocket appears to be specific only for the first four amino acids of the traditional FLAG epitope, namely DYKD. Thus, the use of antibody fragments for improving the stability of target proteins can be rapidly applied to the study of membrane-protein structure by placing the short DKYD motif within a predicted peripheral loop of that protein and utilizing commercially available anti-FLAG M2 antibody fragments.

  14. Efficient production of human bivalent and trivalent anti-MUC1 Fab-scFv antibodies in Pichia pastoris

    Directory of Open Access Journals (Sweden)

    Haustraete Jurgen


    Full Text Available Abstract Background Tumour associated antigens on the surface of tumour cells, such as MUC1, are being used as specific antibody targets for immunotherapy of human malignancies. In order to address the poor penetration of full sized monoclonal antibodies in tumours, intermediate sized antibodies are being developed. The cost-effective and efficient production of these molecules is however crucial for their further success as anti-cancer therapeutics. The methylotropic P. pastoris yeast grows in cheap mineral media and is known for its short process times and the efficient production of recombinant antibody fragments like scFvs, bivalent scFvs and Fabs. Results Based on the anti-MUC1 PH1 Fab, we have developed bivalent PH1 bibodies and trivalent PH1 tribodies of intermediate molecular mass by adding PH1 scFvs to the C-terminus of the Fab chains using flexible peptide linkers. These recombinant antibody derivatives were efficiently expressed in both mammalian and P. pastoris cells. Stable production in NS0 cells produced 130.5 mg pure bibody and 27 mg pure tribody per litre. This high yield is achieved as a result of the high overall purification efficiency of 77%. Expression and purification of PH1 bibodies and tribodies from Pichia supernatant yielded predominantly correctly heterodimerised products, free of light chain homodimers. The yeast-produced bi- and tribodies retained the same specific activity as their mammalian-produced counterparts. Additionally, the yields of 36.8 mg pure bibody and 12 mg pure tribody per litre supernatant make the production of these molecules in Pichia more efficient than most other previously described trispecific or trivalent molecules produced in E. coli. Conclusion Bi- and tribody molecules are efficiently produced in P. pastoris. Furthermore, the yeast produced molecules retain the same specific affinity for their antigen. These results establish the value of P. pastoris as an efficient alternative expression

  15. Two new species of Myliocotyle (Monogenea: Monocotylidae) from the gills of Aetomylaeus maculatus and A. nichofii (Elasmobranchii: Myliobatidae) from Sarawak, Borneo, Malaysia. (United States)

    Chisholm, Leslie A; Whittington, Ian D


    Myliocotyle borneoensis sp. n. and M. multicrista sp. n. (Monocotylidae: Heterocotylinae) are described from the gills of the mottled eagle ray, Aetomylaeus maculatus (Gray), and the banded eagle ray A. nichofii (Bloch et Schneider) (Myliobatidae), respectively, collected from the northern coast of Malaysian Borneo. These are the first monogeneans to be described on elasmobranchs from Borneo. The formerly monotypic Myliocotyle (for M. pteromylaei) was distinguished from other monocotylids by the distribution and morphology of the eight sclerotised dorsal haptoral accessory structures and the morphology of the male copulatory organ. However, we have determined that M. pteromylaei has ten structures on the dorsal surface of the haptor. Myliocotyle borneoensis is distinguished from M. pteromylaei by the morphology of the male copulatory organ and its accessory piece. Myliocotyle multicrista has 12 sclerotised dorsal haptoral accessory structures and a male copulatory organ with two accessory pieces. Additional sclerotised ridges across the ventral surfaces of each loculus (except the posterior-most pair) are also present in M. multicrista. The diagnosis for Myliocotyle is revised given our discovery of additional dorsal haptoral accessory structures in the type species and to accommodate other new characters of the two new species. Anterior secretions of Myliocotyle are discussed.

  16. Helmintos parasitos das espécies Scomberomorus cavalla (Cuvier e Scomberomorus maculatus (Mitchill do litoral cearense: Contracaecum fortalezae sp. n. (Nematoda, Ascaridoidea

    Directory of Open Access Journals (Sweden)

    Vera Lucia M. Klein


    Full Text Available De nematódeos encontrados parasitando estômago, intestinos delgado e grosso de Scomberomorus cavalla (Cuv. e Scomberomorus maculatus (Mitch. o autor propões para o gênero Contracaecum Railliet et Henry, 1912 uma nova espécie, C. fortalezae sp. n., que mais se aproxima de C. clavatum (Rud., 1809 Baylis, 1920 dela se diferenciando por possuir espículos desiguais, um curto ovejetor, útero opistodelfo, ovos menores e por apresentar em todos exemplares estudados ceco intestinal curto e ceco esofagiano longo.In this paper the author proposes a new species of the genus Contracaecum Railliet et Henry, 1912. Of the species under Contracaecum, Contracaecum clavatum (Rudolphi, 1809 Baylis, 1920 is the nearest to Contracaecum fortalezae sp. n. differing from the latter by the following aspects: The males have unequal spicules. The females have a short ovijector and parallel uteri directed backward and the eggs are smaller. In both sexes the intestinal cecum is always short and the ventricular posterior appendix is always long, while Contracaecum clavatum presents these structures in an inverse way, considering the lenght of them. Besides, the thorn-like formations at the posterior end of the body are different.

  17. Biomarkers and bioindicators of the environmental condition using a fish species (Pimelodus maculatus Lacepède, 1803) in a tropical reservoir in Southeastern Brazil. (United States)

    Araújo, F G; Morado, C N; Parente, T T E; Paumgartten, F J R; Gomes, I D


    The Funil Reservoir receives a large amount of xenobiotics from the Paraíba do Sul River (PSR) from large number of industries and municipalities in the watershed. This study aimed to assess environmental quality along the longitudinal profile of the Paraíba do Sul River-Funil Reservoir system, by using biomarkers and bioindicators in a selected fish species. The raised hypothesis is that Funil Reservoir acts as a filter for the xenobiotics of the PSR waters, improving river water quality downstream the dam. Two biomarkers, the ethoxyresorufin-O-deethylase activity (EROD), measured as fluorimetricly in S9 hepatic fraction, and the micronuclei frequency (MN), observed in erythrocytes of the cytoplasm, and three bioindicators, the hepatosomatic index (HSI), gonadosomatic index (GSI) and condition factor (CF) were used in Pimelodus maculatus, a fish species widely distributed in the system. Four zones were searched through a longitudinal gradient: 1, river upstream from the reservoir; 2, upper reservoir; 3, lower reservoir; 4, river downstream of the reservoir. EROD activity and HSI and GSI had significant differences among the zones (Penvironmental condition occur at the river downstream of the reservoir, where it seems to occur more influence of xenobiotics, which could be associated with hydroelectric plant operation. The hypothesis that Funil reservoir acts as a filter decanting pollution from the Paraíba do Sul River waters was rejected. These results are novel information on this subject for a native fish species and could be useful for future comparisons with other environments.

  18. A KAS2 cDNA complements the phenotypes of the Arabidopsis fab1 mutant that differs in a single residue bordering the substrate binding pocket

    DEFF Research Database (Denmark)

    Carlsson, A.S.; LaBrie, S.T.; Kinney, A.J.;


    The fab1 mutant of Arabidopsis is partially deficient in activity of ß-ketoacyl-[acyl carrier protein] synthase II (KAS II). This defect results in increased levels of 16 : 0 fatty acid and is associated with damage and death of the mutants at low temperature. Transformation of fab1 plants with a c...... chain to bend. For functional analysis the equivalent Leu207Phe mutation was introduced into the fabB gene encoding the E. coli KAS I enzyme. Compared to wild-type, the Leu207Phe protein showed a 10-fold decrease in binding affinity for the fatty acid substrate, exhibited a modified behavior during size...

  19. Data of rational process optimization for the production of a full IgG and its Fab fragment from hybridoma cells. (United States)

    Röhm, Martina; Handl, Alina; König, Maria; Mavoungou, Chrystelle; Handrick, René; Schindowski, Katharina


    This data article focuses on the production of monoclonal antibodies (mAb) and their fragments Fab and F(ab')2. Here, we present the data of an optimization protocol to improve the product yield of a hybridoma cell process using a Design of Experiment (DoE) strategy. Furthermore, the data of the evaluated conditions were used to test feeding strategies in shake flasks. They were verified in controlled 2 L fed-batch bioreactor processes. Supplementing the culture medium with human insulin-like growth factor-I (IGF-I) and Pluronic F-68, as well as a nutrient rich additive for fed-batch, resulted in improved cell growth correlating with a 7 day elongated process time and a 4.5 fold higher product titer. Finally, a rapid Fab generation protocol and the respective data are presented using different papain digestion and a camelid anti-kappa light chain VHH affinity ligand.

  20. Crystallization and X-ray diffraction analysis of the β-ketoacyl-acyl carrier protein reductase FabG from Aquifex aeolicus VF5

    Energy Technology Data Exchange (ETDEWEB)

    Mao, Qilong [Hauptman-Woodward Medical Research Institute, 700 Ellicott Street, Buffalo, NY 14203 (United States); Duax, William L.; Umland, Timothy C., E-mail: [Hauptman-Woodward Medical Research Institute, 700 Ellicott Street, Buffalo, NY 14203 (United States); Department of Structural Biology, School of Medicine and Biomedical Sciences, University at Buffalo, Buffalo, NY (United States)


    FabG from A. aeolicus, a putative component of fatty-acid synthase II, has been overexpressed, purified and crystallized. Diffraction data have been collected to 1.8 Å resolution. The gene product of fabG from Aquifex aeolicus has been heterologously expressed in Escherichia coli. Purification of the protein took place using anion-exchange and size-exclusion chromatography and the protein was then crystallized. Diffraction data were collected to a maximum resolution of 1.8 Å and the initial phases were determined by molecular replacement. The A. aeolicus FabG protein is a putative β-ketoacyl-acyl carrier protein reductase. Structure–function studies of this protein are being performed as part of a larger project investigating naturally occurring deviations from highly conserved residues within the short-chain oxidoreductase (SCOR) family.

  1. Absorbed Doses and Risk Estimates of (211)At-MX35 F(ab')2 in Intraperitoneal Therapy of Ovarian Cancer Patients

    DEFF Research Database (Denmark)

    Cederkrantz, Elin; Andersson, Håkan; Bernhardt, Peter


    100 MBq/L, organ equivalent doses were less than 10% of the estimated tolerance dose. CONCLUSION: Intraperitoneal (211)At-MX35 F(ab')2 treatment is potentially a well-tolerated therapy for locally confined microscopic ovarian cancer. Absorbed doses to normal organs are low, but because the effective......, intraperitoneal (i.p.) targeted α therapy has been proposed as an adjuvant treatment for minimal residual disease after successful primary treatment. In the present study, we calculated absorbed and relative biological effect (RBE)-weighted (equivalent) doses in relevant normal tissues and estimated the effective...... dose associated with i.p. administration of (211)At-MX35 F(ab')2. METHODS AND MATERIALS: Patients in clinical remission after salvage chemotherapy for peritoneal recurrence of ovarian cancer underwent i.p. infusion of (211)At-MX35 F(ab')2. Potassium perchlorate was given to block unwanted accumulation...

  2. Taphonomic marks on pig tissue due to cadaveric Coleoptera activity under controlled conditions. (United States)

    Zanetti, Noelia I; Visciarelli, Elena C; Centeno, Néstor D


    The aim of this work was to study taphonomic marks that cadaveric coleopteran can produce under controlled conditions. To evaluate this, pig trotters were initially exposed to adults of Dermestes maculatus De Geer at 21 ± 5°C and a 12:12-h day/night cycle. Observations were made and photographs taken every 4-5 days for 9 months. When feeding and reproducing, D. maculatus produced, in both adult and larvae stages, different types of marks such as holes, striations, scratches, and pits in several kinds of tissue such as integumental, connective, and muscular, in both their fresh and dried stages. Bite marks were also evident. The results in this study provide not only taphonomic but also biological and forensic information. This is the first time that this kind of experiment has been performed. © 2014 American Academy of Forensic Sciences.

  3. Escherichia coli enoyl-acyl carrier protein reductase (FabI) supports efficient operation of a functional reversal of β-oxidation cycle. (United States)

    Vick, Jacob E; Clomburg, James M; Blankschien, Matthew D; Chou, Alexander; Kim, Seohyoung; Gonzalez, Ramon


    We recently used a synthetic/bottom-up approach to establish the identity of the four enzymes composing an engineered functional reversal of the -oxidation cycle for fuel and chemical production in Escherichia coli (J. M. Clomburg, J. E. Vick, M. D. Blankschien, M. Rodriguez-Moya, and R. Gonzalez, ACS Synth Biol 1:541–554, 2012, native enzymes that catalyze the first three steps of the pathway were identified, the identity of the native enzyme(s) acting as the trans-enoyl coenzyme A (CoA) reductase(s) remained unknown, limiting the amount of product that could be synthesized (e.g., 0.34 g/liter butyrate) and requiring the overexpression of a foreign enzyme (the Euglena gracilis trans-enoyl-CoA reductase [EgTER]) to achieve high titers (e.g., 3.4 g/liter butyrate). Here, we examine several native E. coli enzymes hypothesized to catalyze the reduction of enoyl-CoAs to acyl-CoAs. Our results indicate that FabI, the native enoyl-acyl carrier protein (enoyl-ACP) reductase (ENR) from type II fatty acid biosynthesis, possesses sufficient NADH-dependent TER activity to support the efficient operation of a -oxidation reversal. Overexpression of FabI proved as effective as EgTER for the production of butyrate and longer-chain carboxylic acids. Given the essential nature of fabI, we investigated whether bacterial ENRs from other families were able to complement a fabI deletion without promiscuous reduction of crotonyl-CoA. These characteristics from Bacillus subtilis FabL enabled deltaffabI complementation experiments that conclusively established that FabI encodes a native enoyl-CoA reductase activity that supports the β-oxidation reversal in E. coli.

  4. Initial volume of a drug before it reaches the volume of distribution: pharmacokinetics of F(ab')2 antivenoms and other drugs. (United States)

    Sevcik, Carlos; Salazar, Victor; Díaz, Patricia; D'Suze, Gina


    Fast disappearance of F(ab')2 antivenoms from the plasma compartment [Sevcik et al., 2004. Modelling Tityus scorpion venom and antivenom pharmacokinetics. Evidence of active immunoglobulin G's F(ab')2 extrusion mechanism from blood to tissues. Toxicon 44, 731-734; Vazquez et al., 2005. Pharmacokinetics of a F(ab')2 scorpion antivenom in healthy human volunteers. Toxicon 46, 797-805] suggests a quick time course to reach its final distribution volume. An equation was developed to describe how the volume occupied by a drug in the body grows with time. As discussed in the paper this equation is free of some shortcomings of an equation developed for the same purpose by Niazi [1976. Volume of distribution as a function of time. J. Pharm. Sci. 65, 452-454]. Fluorescence microscopy showed that the rapid initial decay in plasmatic F(ab')2 concentration may be related to uptake of F(ab')2 by vascular endothelium which, in combination with accumulation in the vascular wall connective tissue, may produce an intermediate plateau in F(ab')2 V(sl)(t), which reached its final value after 10 h. The V(sl)(t) equation predicts that the plasma concentration half-time of decay has little use to estimate how a drug distributes through the body to exert its action, and predicts that, in some instances, intermediate plateaus in the time course of V(sl)(t) exist. Data from the literature showed that the kinetic considerations for V(sl)(t) also apply to clevidipine, digoxin, digitoxin, lidocaine and thiopentone.

  5. The analysis of VH and VL genes repertoires of Fab library built from peripheral B cells of human rabies virus vaccinated donors. (United States)

    Houimel, Mehdi


    A human combinatorial Fab antibody library was generated from immune repertoire based on peripheral B cells of ten rabies virus vaccinated donors. The analysis of random Fab fragments from the unselected library presented some bias of V gene usage towards IGHV-genes and IGLV-gen families. The screening of the Fab library on rabies virus allowed specific human Fab antibody fragments characterized for their gene encoding sequences, binding and specificities to RV. Genetic analysis of selected Fabs indicated that the IGHV and IGLV differ from the germ-line sequence. At the level of nucleotide sequences, the IGHV and IGLV domains were found to share 74-92% and 90-96% homology with sequences encoded by the corresponding human germ-line genes respectively. IGHV domains are characterized most frequently by IGHV3 genes, and large proportions of the anti-RV heavy chain IGHV domains are obtained following a VDJ recombination process that uses IGHD3, IGHD2, IGHD1 and IGHD6 genes. IGHJ3 and IGHJ4 genes are predominantly used in RV-Fab. The IGLV domains are dominated by IGKV1, IGLV1 and IGLV3 genes. Numerous somatic hypermutations in the RV-specific IGHV are detected, but only limited amino acid replacement in most of the RV-specific IGLV particularly in those encoded by J proximal IGLV or IGKV genes are found. Furthermore, IGHV3-IGKV1, IGHV3-IGVL1, and IGHV3-IGLV3 germ-line family pairings are preferentially enriched after the screening on rabies virus.

  6. Fatty acid biosynthesis in Pseudomonas aeruginosa is initiated by the FabY class of β-ketoacyl acyl carrier protein synthases. (United States)

    Yuan, Yanqiu; Sachdeva, Meena; Leeds, Jennifer A; Meredith, Timothy C


    The prototypical type II fatty acid synthesis (FAS) pathway in bacteria utilizes two distinct classes of β-ketoacyl synthase (KAS) domains to assemble long-chain fatty acids, the KASIII domain for initiation and the KASI/II domain for elongation. The central role of FAS in bacterial viability and virulence has stimulated significant effort toward developing KAS inhibitors, particularly against the KASIII domain of the β-acetoacetyl-acyl carrier protein (ACP) synthase FabH. Herein, we show that the opportunistic pathogen Pseudomonas aeruginosa does not utilize a FabH ortholog but rather a new class of divergent KAS I/II enzymes to initiate the FAS pathway. When a P. aeruginosa cosmid library was used to rescue growth in a fabH downregulated strain of Escherichia coli, a single unannotated open reading frame, PA5174, complemented fabH depletion. While deletion of all four KASIII domain-encoding genes in the same P. aeruginosa strain resulted in a wild-type growth phenotype, deletion of PA5174 alone specifically attenuated growth due to a defect in de novo FAS. Siderophore secretion and quorum-sensing signaling, particularly in the rhl and Pseudomonas quinolone signal (PQS) systems, was significantly muted in the absence of PA5174. The defect could be repaired by intergeneric complementation with E. coli fabH. Characterization of recombinant PA5174 confirmed a preference for short-chain acyl coenzyme A (acyl-CoA) substrates, supporting the identification of PA5174 as the predominant enzyme catalyzing the condensation of acetyl coenzyme A with malonyl-ACP in P. aeruginosa. The identification of the functional role for PA5174 in FAS defines the new FabY class of β-ketoacyl synthase KASI/II domain condensation enzymes.

  7. Pharmacological efficacy of anti-IL-1β scFv, Fab and full-length antibodies in treatment of rheumatoid arthritis. (United States)

    Qi, Jianying; Ye, Xianlong; Ren, Guiping; Kan, Fangming; Zhang, Yu; Guo, Mo; Zhang, Zhiyi; Li, Deshan


    Rheumatoid arthritis (RA) is a chronic autoimmune inflammatory disease that mainly causes the synovial joint inflammation and cartilage destruction. Interleukin-1β (IL-1β) is an important proinflammatory cytokine involved in the pathogenesis of RA. In this study, we constructed and expressed anti-IL-1β-full-length antibody in CHO-K1-SV, anti-IL-1β-Fab and anti-IL-1β-scFv in Rosetta. We compared the therapeutic efficacy of three anti-IL-1β antibodies for CIA mice. Mice with CIA were subcutaneously injected with humanized anti-IL-1β-scFv, anti-IL-1β-Fab or anti-IL-1β-full-length antibody. The effects of treatment were determined by arthritis severity score, autoreactive humoral, cellular immune responses, histological lesion and cytokines production. Compared with anti-IL-1β-scFv treatments, anti-IL-1β-Fab and anti-IL-1β-full-length antibody therapy resulted in more significant effect in alleviating the severity of arthritis by preventing bone damage and cartilage destruction, reducing humoral and cellular immune responses, and down-regulating the expression of IL-1β, IL-6, IL-2, IFN-γ, TNF-α and MMP-3 in inflammatory tissue. The therapeutic effects of anti-IL-1β-Fab and anti-IL-1β-full-length antibodies on CIA mice had no significant difference. However, production of anti-IL-1β-full-length antibody in eukaryotic system is, in general, time-consuming and more expensive than that of anti-IL-1β-Fab in prokaryotic systems. In conclusion, as a small molecule antibody, anti-IL-1β-Fab is an ideal candidate for RA therapy.

  8. Sortase-catalyzed in vitro functionalization of a HER2-specific recombinant Fab for tumor targeting of the plant cytotoxin gelonin. (United States)

    Kornberger, Petra; Skerra, Arne


    We report on the preparation of a new type of immunotoxin via in vitro ligation of the αHer2 antigen binding fragment (Fab) of the clinically-validated antibody trastuzumab to the plant toxin gelonin, employing catalysis by the bacterial enzyme sortase A (SrtA). The αHer2 Fab was fused with the extended SrtA recognition motif LPET↓GLEH 6 at the C-terminus of its heavy chain, thereby preventing interference with antigen binding, while the toxin was equipped with a Gly 2 sequence at its N-terminus, distant to the catalytically active site in the C-terminal region. Site-specific in vitro transpeptidation led to a novel antibody-toxin conjugate wherein gelonin had effectively replaced the Fc region of a conventional (monomerized) immunoglobulin. After optimization of reaction conditions and incubation time, the resulting Fab-Gelonin ligation product was purified to homogeneity in a two-step procedure by means of Strep-Tactin affinity chromatography--utilizing the Strep-tag II appended to gelonin--and size exclusion chromatography. Binding activity of the immunotoxin for the Her2 ectodomain was indistinguishable from the unligated Fab as measured by real-time surface plasmon resonance spectroscopy. Specific cytotoxic potency of Fab-Gelonin was demonstrated against two Her2-positive cell lines, resulting in EC 50 values of ~1 nM or lower, indicating a 1000-fold enhanced cell-killing activity compared with gelonin itself. Thus, our strategy provides a convenient route to the modular construction of functional immunotoxins from Fabs of established tumor-specific antibodies with gelonin or related proteotoxins, also avoiding the elevated biosafety levels that would be mandatory for the direct biotechnological preparation of corresponding fusion proteins.

  9. Functional humanization of an anti-CD16 Fab fragment: obstacles of switching from murine {lambda} to human {lambda} or {kappa} light chains. (United States)

    Schlapschy, Martin; Fogarasi, Marton; Gruber, Helga; Gresch, Oliver; Schäfer, Claudia; Aguib, Yasmine; Skerra, Arne


    An alphaCD30xalphaCD16 bispecific monoclonal antibody (MAb) was previously shown to induce remission of Hodgkin's disease refractory to chemo- and radiotherapy through specific activation of natural killer (NK) cells, but the appearance of a human anti-mouse antibody (HAMA) response prevented its use for prolonged therapy. Here, we describe an effort to humanize the Fab arm directed against FcgammaRIII (CD16), which-in context with the previously humanized CD30 Fab fragment-provides the necessary component for the design of a clinically useful bispecific antibody. Thus, the CDRs of the anti-CD16 mouse IgG1/lambda MAb A9 were grafted onto human Ig sequences. In a first attempt, the murine V(lambda) domain was converted to a humanized lambda chain, which led, however, to complete loss of antigen-binding activity and extremely poor folding efficiency upon periplasmic expression in Escherichia coli. Hence, its CDRs were transplanted onto a human kappa light chain in a second attempt, which resulted in a functional recombinant Fab fragment, yet with 100-fold decreased antigen affinity. In the next step, an in vitro affinity maturation was performed, wherein random mutations were introduced into the humanized V(H) and V(kappa) domains through error-prone PCR, followed by a filter sandwich colony screening assay for increased binding activity towards the bacterially produced extracellular CD16 fragment. Finally, an optimized Fab fragment was obtained, which carries nine additional amino acid exchanges and exhibits an affinity that is within a factor of 2 identical to that of the original murine A9 Fab fragment. The resulting humanized Fab fragment was fully functional with respect to binding of the recombinant CD16 antigen in enzyme-linked immunosorbent assay and in cytofluorimetry with CD16-positive granulocytes, thus providing a promising starting point for the preparation of a fully human bispecific antibody that permits the therapeutic recruitment of NK cells.

  10. Metabolic flux between unsaturated and saturated fatty acids is controlled by the FabA:FabB ratio in the fully reconstituted fatty acid biosynthetic pathway of Escherichia coli. (United States)

    Xiao, Xirui; Yu, Xingye; Khosla, Chaitan


    The entire fatty acid biosynthetic pathway of Escherichia coli, starting from the acetyl-CoA carboxylase, has been reconstituted in vitro from 14 purified protein components. Radiotracer analysis verified stoichiometric conversion of acetyl-CoA and NAD(P)H to the free fatty acid product, allowing implementation of a facile spectrophotometric assay for kinetic analysis of this multienzyme system. At steady state, a maximal turnover rate of 0.5 s(-1) was achieved. Under optimal turnover conditions, the predominant products were C16 and C18 saturated as well as monounsaturated fatty acids. The reconstituted system allowed us to quantitatively interrogate the factors that influence metabolic flux toward unsaturated versus saturated fatty acids. In particular, the concentrations of the dehydratase FabA and the β-ketoacyl synthase FabB were found to be crucial for controlling this property. Via changes in these variables, the percentage of unsaturated fatty acid produced could be adjusted between 10 and 50% without significantly affecting the maximal turnover rate of the pathway. Our reconstituted system provides a powerful tool for understanding and engineering rate-limiting and regulatory steps in this complex and practically significant metabolic pathway.

  11. Descriptions of three new species of Strattis Pascoe, 1883 (Coleoptera: Curculionidae: Cryptorhynchinae) from the Indian subcontinent. (United States)

    Devi, Salam Rita; Ray, D C; Ramamurthy, V V


    The cryptorhynchine genus Strattis Pascoe originally described from Sri Lanka is recorded from India with the descriptions of three new species: S. pascoei sp. nov., S. maculatus sp. nov., and S. srilankaiensis sp. nov. from the Indian subcontinent. Strattis biguttatus is designated as type species for the genus. A key to species is provided. The character of a presumable stridulatory structure between the meta-femur and the first abdominal ventrite is discussed.

  12. Possible origin of B chromosome in Dichotomius sericeus (Coleoptera). (United States)

    Amorim, Igor Costa; Milani, Diogo; Cabral-de-Mello, Diogo Cavalcanti; Rocha, Marília França; Moura, Rita Cássia


    B chromosomes have so far been described in about 80 species of Coleoptera, mainly using conventional staining analysis. In this study, 152 individuals of the dung beetle Dichotomius sericeus (Coleoptera), collected from three isolated geographical areas in the State of Pernambuco, Brazil, were analyzed to determine the frequency, prevalence, distribution, meiotic behavior, and possible B chromosome origin. The cytogenetic analysis consisted of conventional staining, C-banding, triple fluorochrome staining (CMA3/DA/DAPI), and fluorescent in situ hybridization using ribosomal DNAs (rDNAs) and H3 histone gene as probes, as well as microdissection and chromosome painting of the B chromosome. The B chromosomes were detected in all populations analyzed. Analysis revealed the heterochromatic nature and the presence of G+C-rich blocks and 18S rDNA on the B chromosome. FISH with DNA from microdissected B chromosome painted the entire extension of the B chromosome for all populations, besides the pericentromeric regions of all the autosomes, as well as the X chromosome. Finally, cross-hybridization in nine related species of Dichotomius using the microdissected B chromosome as probe did not reveal any hybridization signal. The results suggest an intraspecific and monophyletic origin for B chromosomes in D. sericeus, probably from the second or third autosomal pair.

  13. Rove beetles of medical importance in Brazil (Coleoptera, Staphylinidae, Paederinae

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    Juliana S. Vieira


    Full Text Available Rove beetles of medical importance in Brazil (Coleoptera, Staphylinidae, Paederinae. The rove beetles of the genus Paederus Fabricius, 1775 are the most important group within Coleoptera causing dermatitis around the world. The medical importance of Paederus depends on its toxic hemolymph released when these beetles are crushed on human skin. The effects are mainly dermatitis linearis and some sporadic cases of conjunctivitis. In Brazil seven species of Paederus are known to cause dermatitis: P. amazonicus Sharp, 1876, P. brasiliensis Erichson, 1840, P. columbinus Laporte, 1835, P. ferus Erichson, 1840, P. mutans Sharp, 1876, P. protensus Sharp, 1876 stat. rev., and Paederus rutilicornis Erichson, 1840. Paederus mutans and P. protensus are for the first time recorded as of medical importance, whereas the record of P. rutilicornis in Brazil is doubtful. All seven species are redescribed and a dichotomous key is provided. The geographic distributions of all species are documented. The results provided here include the most recent and relevant taxonomic revision of Paederus of the Neotropical region, the first identification key for Brazilian species and the increase of recorded species of medical importance in the world.

  14. Variations on a Theme: Antennal Lobe Architecture across Coleoptera. (United States)

    Kollmann, Martin; Schmidt, Rovenna; Heuer, Carsten M; Schachtner, Joachim


    Beetles comprise about 400,000 described species, nearly one third of all known animal species. The enormous success of the order Coleoptera is reflected by a rich diversity of lifestyles, behaviors, morphological, and physiological adaptions. All these evolutionary adaptions that have been driven by a variety of parameters over the last about 300 million years, make the Coleoptera an ideal field to study the evolution of the brain on the interface between the basic bauplan of the insect brain and the adaptions that occurred. In the current study we concentrated on the paired antennal lobes (AL), the part of the brain that is typically responsible for the first processing of olfactory information collected from olfactory sensilla on antenna and mouthparts. We analyzed 63 beetle species from 22 different families and thus provide an extensive comparison of principal neuroarchitecture of the AL. On the examined anatomical level, we found a broad diversity including AL containing a wide range of glomeruli numbers reaching from 50 to 150 glomeruli and several species with numerous small glomeruli, resembling the microglomerular design described in acridid grasshoppers and diving beetles, and substructures within the glomeruli that have to date only been described for the small hive beetle, Aethina tumida. A first comparison of the various anatomical features of the AL with available descriptions of lifestyle and behaviors did so far not reveal useful correlations. In summary, the current study provides a solid basis for further studies to unravel mechanisms that are basic to evolutionary adaptions of the insect olfactory system.

  15. Site specific discrete PEGylation of (124)I-labeled mCC49 Fab' fragments improves tumor MicroPET/CT imaging in mice. (United States)

    Ding, Haiming; Carlton, Michelle M; Povoski, Stephen P; Milum, Keisha; Kumar, Krishan; Kothandaraman, Shankaran; Hinkle, George H; Colcher, David; Brody, Rich; Davis, Paul D; Pokora, Alex; Phelps, Mitchell; Martin, Edward W; Tweedle, Michael F


    The tumor-associated glycoprotein-72 (TAG-72) antigen is highly overexpressed in various human adenocarcinomas and anti-TAG-72 monoclonal antibodies, and fragments are therefore useful as pharmaceutical targeting vectors. In this study, we investigated the effects of site-specific PEGylation with MW 2-4 kDa discrete, branched PEGylation reagents on mCC49 Fab' (MW 50 kDa) via in vitro TAG72 binding, and in vivo blood clearance kinetics, biodistribution, and mouse tumor microPET/CT imaging. mCC49Fab' (Fab'-NEM) was conjugated at a hinge region cysteine with maleimide-dPEG 12-(dPEG24COOH)3 acid (Mal-dPEG-A), maleimide-dPEG12-(dPEG12COOH)3 acid (Mal-dPEG-B), or maleimide-dPEG12-(m-dPEG24)3 (Mal-dPEG-C), and then radiolabeled with iodine-124 ((124)I) in vitro radioligand binding assays and in vivo studies used TAG-72 expressing LS174T human colon carcinoma cells and xenograft mouse tumors. Conjugation of mCC49Fab' with Mal-dPEG-A (Fab'-A) reduced the binding affinity of the non PEGylated Fab' by 30%; however, in vivo, Fab'-A significantly lengthened the blood retention vs Fab'-NEM (47.5 vs 28.1%/ID at 1 h, 25.1 vs 8.4%/ID at 5 h, p images due to higher tumor accumulation, and increased tumor concentration in excised tissues at 72 h by 130% (5.09 ± 0.83 vs 3.83 ± 1.50%ID/g, p imaging tumor signal intensity, and residual 72 h tumor concentration by 49% (3.83 ± 1.50 vs 1.97 ± 0.29%ID/g, p < 0.05) and 63% (3.83 ± 1.50 vs 1.42 ± 0.35%ID/g, p < 0.05), respectively. We conclude that remarkably subtle changes in the structure of the PEGylation reagent can create significantly altered biologic behavior. Further study is warranted of conjugates of the triple branched, negatively charged Mal-dPEG-A.

  16. Crystallization and X-ray diffraction analysis of the beta-ketoacyl-acyl carrier protein reductase FabG from Aquifex aeolicus VF5. (United States)

    Mao, Qilong; Duax, William L; Umland, Timothy C


    The gene product of fabG from Aquifex aeolicus has been heterologously expressed in Escherichia coli. Purification of the protein took place using anion-exchange and size-exclusion chromatography and the protein was then crystallized. Diffraction data were collected to a maximum resolution of 1.8 A and the initial phases were determined by molecular replacement. The A. aeolicus FabG protein is a putative beta-ketoacyl-acyl carrier protein reductase. Structure-function studies of this protein are being performed as part of a larger project investigating naturally occurring deviations from highly conserved residues within the short-chain oxidoreductase (SCOR) family.

  17. Structure of a human monoclonal antibody Fab fragment against gp41 of human immunodeficiency virus type 1 (United States)

    He, Xiao M.; Rueker, Florian; Casale, Elena; Carter, Daniel C.


    The three-dimensional structure of a human monoclonal antibody (Fab), which binds specifically to a major epitope of the transmembrane protein gp41 of the human immunodeficiency virus type 1, has been determined by crystallographic methods to a resolution of 2.7 A. It has been previously determined that this antibody recognizes the epitope SGKLICTTAVPWNAS, belongs to the subclass IgG1 (kappa), and exhibits antibody-dependent cellular cytotoxicity. The quaternary structure of the Fab is in an extended conformation with an elbow bend angle between the constant and variable domains of 175 deg. Structurally, four of the hypervariable loops can be classified according to previously recognized canonical structures. The third hypervariable loops of the heavy (H3) and light chain (L3) are structurally distinct. Hypervariable loop H3, residues 102H-109H, is unusually extended from the surface. The complementarity-determining region forms a hydrophobic binding pocket that is created primarily from hypervariable loops L3, H3, and H2.

  18. The role of indium-111 antimyosin (Fab) imaging as a noninvasive surveillance method of human heart transplant rejection

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    De Nardo, D.; Scibilia, G.; Macchiarelli, A.G.; Cassisi, A.; Tonelli, E.; Papalia, U.; Gallo, P.; Antolini, M.; Pitucco, G.; Reale, A. (Universita degli Studi di Roma I La Sapienza Policlinico Umberto I (Italy))


    The identification of rejection after heart transplantation in patients receiving cyclosporine immunosuppressive therapy requires the endomyocardial biopsy, an invasive method associated with a finite morbidity. To evaluate the role of indium-111 antimyosin (Fab) scintigraphy as a noninvasive surveillance method of heart transplant rejection, the Fab fragment of murine monoclonal antimyosin antibodies labeled with indium-111 was administered intravenously in 30 scintigraphic studies to 10 consecutive heart transplant recipients. Endomyocardial biopsy specimens were obtained 72 hours after each scintigraphic study. Nineteen scintigraphic studies had negative findings; no false negative finding was obtained. Eleven antimyosin scintigraphic studies had positive findings, and in these studies endomyocardial biopsy revealed mild rejection in two cases, moderate acute rejection with myocyte necrosis in two cases, myocyte necrosis as a consequence of ischemic injury in six cases, and possibly cytotoxic damage in one case. Antimyosin scintigraphy may represent a reliable screening method for the surveillance of heart transplant patients. In the presence of a negative finding from antimyosin scintigraphy, it may be possible to avoid endomyocardial biopsy. Conversely, in patients who have a positive finding from antimyosin scintigraphy, the endomyocardial biopsy is mandatory to establish the definitive diagnosis by histologic examination of the myocardium.

  19. Crystallization and preliminary X-ray crystallographic analysis of the sclerostin-neutralizing Fab AbD09097. (United States)

    Boschert, Verena; Muth, Eva Maria; Knappik, Achim; Frisch, Christian; Mueller, Thomas D


    The secreted cystine-knot protein sclerostin was first identified from genetic screening of patients suffering from the rare bone-overgrowth diseases sclerosteosis and van Buchem disease. Sclerostin acts a negative regulator of bone growth through inhibiting the canonical Wnt signalling cascade by binding to and blocking the Wnt co-receptor LRP5/6. Its function in blocking osteoblastogenesis makes it an important target for osteoanabolic therapy approaches to treat osteoporosis, which is characterized by a progressive decrease in bone mass and density. In this work, the production, crystallization and preliminary X-ray diffraction data analysis of a sclerostin-neutralizing human Fab antibody fragment, AbD09097, obtained from a naive antibody library are reported. Crystals of the Fab AbD09097 belonged to space group P21, with unit-cell parameters a = 45.19, b = 78.49, c = 59.20 Å, β = 95.71° and diffracted X-rays to a resolution of 1.8 Å.

  20. Crystal Structure of the Fab Fragment of an Anti-ofloxacin Antibody and Exploration of Its Specific Binding. (United States)

    He, Kuo; Du, Xinjun; Sheng, Wei; Zhou, Xiaonan; Wang, Junping; Wang, Shuo


    The limited knowledge on the mechanism of interactions between small contaminants and the corresponding antibodies greatly inhibits the development of enzyme-linked immunosorbent assay methods. In this study, the crystal structure of a Fab fragment specific for ofloxacin was obtained. On the basis of the crystal characteristics, the modeling of the interactions between ofloxacin and the Fab revealed that TYR31 and HIS99 of the heavy chain and MET20 and GLN79 of the light chain formed a hydrophobic region and that SER52 and ALA97 of the heavy chain and TYR35 of the light chain formed a salt bridge and two hydrogen bonds for specific binding. The key roles of SER52 and ALA97 were further confirmed by site-directed mutation. A specificity analysis using 14 ofloxacin analogues indicates that the length of the bond formed between the piperazine ring and the antibody plays key roles in specific recognition. This work helps to clarify the mechanisms through which antibodies recognize small molecules and improve immune detection methods.