WorldWideScience

Sample records for camelid anti-prp antibody

  1. Magnetosome Expression of Functional Camelid Antibody Fragments (Nanobodies) in Magnetospirillum gryphiswaldense▿†

    OpenAIRE

    Pollithy, Anna; Romer, Tina; Lang, Claus; Müller, Frank D.; Helma, Jonas; Leonhardt, Heinrich; Rothbauer, Ulrich; Schüler, Dirk

    2011-01-01

    Numerous applications of conventional and biogenic magnetic nanoparticles (MNPs), such as in diagnostics, immunomagnetic separations, and magnetic cell labeling, require the immobilization of antibodies. This is usually accomplished by chemical conjugation, which, however, has several disadvantages, such as poor efficiency and the need for coupling chemistry. Here, we describe a novel strategy to display a functional camelid antibody fragment (nanobody) from an alpaca (Lama pacos) on the surf...

  2. Alpaca (Lama pacos) as a convenient source of recombinant camelid heavy chain antibodies (VHHs)

    OpenAIRE

    Maass, David R.; Sepulveda, Jorge; Pernthaner, Anton; Shoemaker, Charles B.

    2007-01-01

    Recombinant single domain antibody fragments (VHHs) that derive from the unusual camelid heavy chain only IgG class (HCAbs) have many favourable properties compared with single-chain antibodies prepared from conventional IgG. As a result, VHHs have become widely used as binding reagents and are beginning to show potential as therapeutic agents. To date, the source of VHH genetic material has been camels and llamas despite their large size and limited availability. Here we demonstrate that the...

  3. Fully Human VH Single Domains That Rival the Stability and Cleft Recognition of Camelid Antibodies.

    Science.gov (United States)

    Rouet, Romain; Dudgeon, Kip; Christie, Mary; Langley, David; Christ, Daniel

    2015-05-01

    Human VH single domains represent a promising class of antibody fragments with applications as therapeutic modalities. Unfortunately, isolated human VH domains also generally display poor biophysical properties and a propensity to aggregate. This has encouraged the development of non-human antibody domains as alternative means of antigen recognition and, in particular, camelid (VHH) domains. Naturally devoid of light chain partners, these domains are characterized by favorable biophysical properties and propensity for cleft binding, a highly desirable characteristic, allowing the targeting of cryptic epitopes. In contrast, previously reported structures of human VH single domains had failed to recapitulate this property. Here we report the engineering and characterization of phage display libraries of stable human VH domains and the selection of binders against a diverse set of antigens. Unlike "camelized" human domains, the domains do not rely on potentially immunogenic framework mutations and maintain the structure of the VH/VL interface. Structure determination in complex with hen egg white lysozyme revealed an extended VH binding interface, with complementarity-determining region 3 deeply penetrating into the active site cleft, highly reminiscent of what has been observed for camelid domains. Taken together, our results demonstrate that fully human VH domains can be constructed that are not only stable and well expressed but also rival the cleft binding properties of camelid antibodies.

  4. Fully Human VH Single Domains That Rival the Stability and Cleft Recognition of Camelid Antibodies*

    Science.gov (United States)

    Rouet, Romain; Dudgeon, Kip; Christie, Mary; Langley, David; Christ, Daniel

    2015-01-01

    Human VH single domains represent a promising class of antibody fragments with applications as therapeutic modalities. Unfortunately, isolated human VH domains also generally display poor biophysical properties and a propensity to aggregate. This has encouraged the development of non-human antibody domains as alternative means of antigen recognition and, in particular, camelid (VHH) domains. Naturally devoid of light chain partners, these domains are characterized by favorable biophysical properties and propensity for cleft binding, a highly desirable characteristic, allowing the targeting of cryptic epitopes. In contrast, previously reported structures of human VH single domains had failed to recapitulate this property. Here we report the engineering and characterization of phage display libraries of stable human VH domains and the selection of binders against a diverse set of antigens. Unlike “camelized” human domains, the domains do not rely on potentially immunogenic framework mutations and maintain the structure of the VH/VL interface. Structure determination in complex with hen egg white lysozyme revealed an extended VH binding interface, with complementarity-determining region 3 deeply penetrating into the active site cleft, highly reminiscent of what has been observed for camelid domains. Taken together, our results demonstrate that fully human VH domains can be constructed that are not only stable and well expressed but also rival the cleft binding properties of camelid antibodies. PMID:25737448

  5. Isolation and optimization of camelid single-domain antibodies:Dirk Saerens’work on nanobodies

    Institute of Scientific and Technical Information of China (English)

    Dirk; Saerens

    2010-01-01

    It is well established that all camelids have unique antibodies circulating in their blood.Unlike antibodies from all other species,these special antibodies are devoid of light chains,and are composed of a heavy chain homodimer.These so-called heavy-chain antibodies(HCAbs)are expressed after a V-D-J rearrangement and require dedicated constant gamma genes. An immune response is raised in these HCAbs following a classical immunization protocol.These HCAbs are easily purified from serum,and their antigen-binding fragment interacts with parts of the target that are less antigenic to conventional antibodies.The antigen binding site of the dromedary HCAb comprises one single domain,referred to as VHH or nanobody(Nb),therefore,a strategy was designed to clone the Nb repertoire of an immunized dromedary and to select the Nb with specificity for our target antigens.The monoclonal Nb is produced well in bacteria,is very stable and highly soluble,and it binds the antigen with high affinity and specificity.Currently,the recombinant Nb has been developed successfully for research purposes, as a probe in biosensors,to diagnose infections,or to treat diseases such as cancer or trypanosomiasis.

  6. Alpaca (Lama pacos) as a convenient source of recombinant camelid heavy chain antibodies (VHHs).

    Science.gov (United States)

    Maass, David R; Sepulveda, Jorge; Pernthaner, Anton; Shoemaker, Charles B

    2007-07-31

    Recombinant single domain antibody fragments (VHHs) that derive from the unusual camelid heavy chain only IgG class (HCAbs) have many favourable properties compared with single-chain antibodies prepared from conventional IgG. As a result, VHHs have become widely used as binding reagents and are beginning to show potential as therapeutic agents. To date, the source of VHH genetic material has been camels and llamas despite their large size and limited availability. Here we demonstrate that the smaller, more tractable and widely available alpaca is an excellent source of VHH coding DNA. Alpaca sera IgG consists of about 50% HCAbs, mostly of the short-hinge variety. Sequencing of DNA encoding more than 50 random VHH and hinge domains permitted the design of PCR primers that will amplify virtually all alpaca VHH coding DNAs for phage display library construction. Alpacas were immunized with ovine tumour necrosis factor alpha (TNFalpha) and a VHH phage display library was prepared from a lymph node that drains the sites of immunizations and successfully employed in the isolation of VHHs that bind and neutralize ovine TNFalpha. PMID:17568607

  7. Isolation of Camelid Single-Domain Antibodies Against Native Proteins Using Recombinant Multivalent Peptide Ligands.

    Science.gov (United States)

    Alturki, Norah A; Henry, Kevin A; MacKenzie, C Roger; Arbabi-Ghahroudi, Mehdi

    2015-01-01

    Generation of antibodies against desired epitopes on folded proteins may be hampered by various characteristics of the target protein, including antigenic and immunogenic dominance of irrelevant epitopes and/or steric occlusion of the desired epitope. In such cases, peptides encompassing linear epitopes of the native protein represent attractive alternative reagents for immunization and screening. Peptide antigens are typically prepared by fusing or conjugating the peptide of interest to a carrier protein. The utility of such antigens depends on many factors including the peptide's amino acid sequence, display valency, display format (synthetic conjugate vs. recombinant fusion) and characteristics of the carrier. Here we provide detailed protocols for: (1) preparation of DNA constructs encoding peptides fused to verotoxin (VT) multimerization domain; (2) expression, purification, and characterization of the multivalent peptide-VT ligands; (3) concurrent panning of a non-immune phage-displayed camelid VHH library against the peptide-VT ligands and native protein; and (4) identification of VHHs enriched via panning using next-generation sequencing techniques. These methods are simple, rapid and can be easily adapted to yield custom peptide-VT ligands that appear to maintain the antigenic structures of the peptide. However, we caution that peptide sequences should be chosen with great care, taking into account structural, immunological, and biophysical information on the protein of interest.

  8. Development and utilization of camelid VHH antibodies from alpaca for 2,2',4,4'-tetrabrominated diphenyl ether detection.

    Science.gov (United States)

    Bever, Candace R S; Majkova, Zuzana; Radhakrishnan, Rajeswaran; Suni, Ian; McCoy, Mark; Wang, Yanru; Dechant, Julie; Gee, Shirley; Hammock, Bruce D

    2014-08-01

    An antibody-based analytical method for the detection of a chemical flame retardant using antibody fragments isolated from an alpaca has been developed. One specific chemical flame retardant congener, 2,2',4,4'-tetrabrominated diphenyl ether (BDE-47), is often the major poly-BDE (PBDE) congener present in human and environmental samples and that which is the most frequently detected. An alpaca was immunized with a surrogate of BDE-47 covalently attached to a carrier protein. The resulting mRNA coding for the variable domain of heavy-chain antibodies (VHH) were isolated, transcribed to cDNA, and cloned into a phagemid vector for phage display library construction. Selection of VHHs recognizing BDE-47 was achieved by panning under carefully modified conditions. The assay sensitivity for detecting BDE-47 was down to the part-per-billion (microgram per liter) level. Cross-reactivity analyses confirmed that this method was highly selective for BDE-47 and selected hydroxylated metabolites. When exposed to elevated temperatures, the camelid VHH antibodies retained more reactivity than a polyclonal antibody developed to the same target analyte. The use of this VHH antibody reagent immobilized onto a Au electrode for impedance biosensing demonstrates the increased versatility of VHH antibodies. PMID:25005746

  9. Novel Camelid Antibody Fragments Targeting Recombinant Nucleoprotein of Araucaria hantavirus: A Prototype for an Early Diagnosis of Hantavirus Pulmonary Syndrome

    Science.gov (United States)

    Pereira, Soraya S.; Moreira-Dill, Leandro S.; Morais, Michelle S. S.; Prado, Nidiane D. R.; Barros, Marcos L.; Koishi, Andrea C.; Mazarrotto, Giovanny A. C. A.; Gonçalves, Giselle M.; Zuliani, Juliana P.; Calderon, Leonardo A.; Soares, Andreimar M.; Pereira da Silva, Luiz H.; Duarte dos Santos, Claudia N.; Fernandes, Carla F. C.; Stabeli, Rodrigo G.

    2014-01-01

    In addition to conventional antibodies, camelids produce immunoglobulins G composed exclusively of heavy chains in which the antigen binding site is formed only by single domains called VHH. Their particular characteristics make VHHs interesting tools for drug-delivery, passive immunotherapy and high-throughput diagnosis. Hantaviruses are rodent-borne viruses of the Bunyaviridae family. Two clinical forms of the infection are known. Hemorrhagic Fever with Renal Syndrome (HFRS) is present in the Old World, while Hantavirus Pulmonary Syndrome (HPS) is found on the American continent. There is no specific treatment for HPS and its diagnosis is carried out by molecular or serological techniques, using mainly monoclonal antibodies or hantavirus nucleoprotein (N) to detect IgM and IgG in patient serum. This study proposes the use of camelid VHHs to develop alternative methods for diagnosing and confirming HPS. Phage display technology was employed to obtain VHHs. After immunizing one Lama glama against the recombinant N protein (prNΔ85) of a Brazilian hantavirus strain, VHH regions were isolated to construct an immune library. VHHs were displayed fused to the M13KO7 phage coat protein III and the selection steps were performed on immobilized prNΔ85. After selection, eighty clones recognized specifically the N protein. These were sequenced, grouped based mainly on the CDRs, and five clones were analyzed by western blot (WB), surface plasmon resonance (SPR) device, and ELISA. Besides the ability to recognize prNΔ85 by WB, all selected clones showed affinity constants in the nanomolar range. Additionaly, the clone KC329705 is able to detect prNΔ85 in solution, as well as the native viral antigen. Findings support the hypothesis that selected VHHs could be a powerful tool in the development of rapid and accurate HPS diagnostic assays, which are essential to provide supportive care to patients and reduce the high mortality rate associated with hantavirus infections. PMID

  10. Novel camelid antibody fragments targeting recombinant nucleoprotein of Araucaria hantavirus: a prototype for an early diagnosis of Hantavirus Pulmonary Syndrome.

    Directory of Open Access Journals (Sweden)

    Soraya S Pereira

    Full Text Available In addition to conventional antibodies, camelids produce immunoglobulins G composed exclusively of heavy chains in which the antigen binding site is formed only by single domains called VHH. Their particular characteristics make VHHs interesting tools for drug-delivery, passive immunotherapy and high-throughput diagnosis. Hantaviruses are rodent-borne viruses of the Bunyaviridae family. Two clinical forms of the infection are known. Hemorrhagic Fever with Renal Syndrome (HFRS is present in the Old World, while Hantavirus Pulmonary Syndrome (HPS is found on the American continent. There is no specific treatment for HPS and its diagnosis is carried out by molecular or serological techniques, using mainly monoclonal antibodies or hantavirus nucleoprotein (N to detect IgM and IgG in patient serum. This study proposes the use of camelid VHHs to develop alternative methods for diagnosing and confirming HPS. Phage display technology was employed to obtain VHHs. After immunizing one Lama glama against the recombinant N protein (prNΔ₈₅ of a Brazilian hantavirus strain, VHH regions were isolated to construct an immune library. VHHs were displayed fused to the M13KO7 phage coat protein III and the selection steps were performed on immobilized prNΔ₈₅. After selection, eighty clones recognized specifically the N protein. These were sequenced, grouped based mainly on the CDRs, and five clones were analyzed by western blot (WB, surface plasmon resonance (SPR device, and ELISA. Besides the ability to recognize prNΔ85 by WB, all selected clones showed affinity constants in the nanomolar range. Additionaly, the clone KC329705 is able to detect prNΔ₈₅ in solution, as well as the native viral antigen. Findings support the hypothesis that selected VHHs could be a powerful tool in the development of rapid and accurate HPS diagnostic assays, which are essential to provide supportive care to patients and reduce the high mortality rate associated with

  11. Analysis of camelid antibodies for antivenom development: Neutralisation of venom-induced pathology.

    Science.gov (United States)

    Cook, Darren A N; Owen, Timothy; Wagstaff, Simon C; Kinne, Joerg; Wernery, Ulrich; Harrison, Robert A

    2010-09-01

    Camelid IgG has been reported to be less immunogenic, less able to activate the complement cascade and more thermostable than IgG from other mammals, and has the ability to bind antigens that are unreactive with other mammalian IgGs. We are investigating whether these attributes of camelid IgG translate into antivenom with immunological and venom-neutralising efficacy advantages over conventional equine and ovine antivenoms. The objective of this study was to determine the preclinical venom-neutralising effectiveness of IgG from camels immunised with venoms, individually or in combination, of the saw-scaled viper, Echis ocellatus, the puff adder, Bitis arietans and the spitting cobra, Naja nigricollis - the most medically-important snake species in West Africa. Neutralisation of the pathological effects of venoms from E. ocellatus, B. arietans and N. nigricollis by IgG from the venom-immunised camels, or commercial antivenom, was compared using assays of venom lethality (ED(50)), haemorrhage (MHD) and coagulopathy (MCD). The E. ocellatus venom ED(50), MHD and MCD results of the E. ocellatus monospecific camel IgG antivenom were broadly equivalent to comparable ovine (EchiTAbG, MicroPharm Ltd, Wales) and equine (SAIMR Echis, South African Vaccine Producer, South Africa) antivenoms, although the equine antivenom required half the amount of IgG. The B. arietans monospecific camel IgG neutralised the lethal effects of B. arietans venom at one fourth the concentration of the SAIMR polyspecific antivenom (a monospecific B. arietans antivenom is not available). The N. nigricollis camel IgG antivenom was ineffective (at the maximum permitted dose, 100 mul) against the lethal effects of N. nigricollis venom. All the equine polyspecific antivenoms required more than 100 microl to be effective against this venom. The polyspecific camel IgG antivenom, prepared from five camels, was effective against the venom-induced effects of E. ocellatus but not against that of B. arietans

  12. An anti-hapten camelid antibody reveals a cryptic binding site with significant energetic contributions from a nonhypervariable loop

    Energy Technology Data Exchange (ETDEWEB)

    Fanning, Sean W.; Horn, James R. (NIU)

    2014-03-05

    Conventional anti-hapten antibodies typically bind low-molecular weight compounds (haptens) in the crevice between the variable heavy and light chains. Conversely, heavy chain-only camelid antibodies, which lack a light chain, must rely entirely on a single variable domain to recognize haptens. While several anti-hapten VHHs have been generated, little is known regarding the underlying structural and thermodynamic basis for hapten recognition. Here, an anti-methotrexate VHH (anti-MTX VHH) was generated using grafting methods whereby the three complementarity determining regions (CDRs) were inserted onto an existing VHH framework. Thermodynamic analysis of the anti-MTX VHH CDR1-3 Graft revealed a micromolar binding affinity, while the crystal structure of the complex revealed a somewhat surprising noncanonical binding site which involved MTX tunneling under the CDR1 loop. Due to the close proximity of MTX to CDR4, a nonhypervariable loop, the CDR4 loop sequence was subsequently introduced into the CDR1-3 graft, which resulted in a dramatic 1000-fold increase in the binding affinity. Crystal structure analysis of both the free and complex anti-MTX CDR1-4 graft revealed CDR4 plays a significant role in both intermolecular contacts and binding site conformation that appear to contribute toward high affinity binding. Additionally, the anti-MTX VHH possessed relatively high specificity for MTX over closely related compounds aminopterin and folate, demonstrating that VHH domains are capable of binding low-molecular weight ligands with high affinity and specificity, despite their reduced interface.

  13. One-step immunoassay for tetrabromobisphenol a using a camelid single domain antibody-alkaline phosphatase fusion protein.

    Science.gov (United States)

    Wang, Jia; Majkova, Zuzana; Bever, Candace R S; Yang, Jun; Gee, Shirley J; Li, Ji; Xu, Ting; Hammock, Bruce D

    2015-01-01

    Tetrabromobisphenol A (TBBPA) is a ubiquitous brominated flame retardant, showing widespread environmental and human exposures. A variable domain of the heavy chain antibody (VHH), naturally occurring in camelids, approaches the lower size limit of functional antigen-binding entities. The ease of genetic manipulation makes such VHHs a superior choice to use as an immunoreagent. In this study, a highly selective anti-TBBPA VHH T3-15 fused with alkaline phosphatase (AP) from E. coli was expressed, showing both an integrated TBBPA-binding capacity and enzymatic activity. A one-step immunoassay based on the fusion protein T3-15-AP was developed for TBBPA in 5% dimethyl sulfoxide (DMSO)/phosphate buffered saline (PBS, pH 7.4), with a half-maximum signal inhibition concentration (IC50) of 0.20 ng mL(-1). Compared to the parental VHH T3-15, T3-15-AP was able to bind to a wider variety of coating antigens and the assay sensitivity was slightly improved. Cross-reactivity of T3-15-AP with a set of important brominated analogues was negligible (<0.1%). Although T3-15-AP was susceptible to extreme heat (90 °C), much higher binding stability at ambient temperature was observed in the T3-15-AP-based assay for at least 70 days. A simple pretreatment method of diluting urine samples with DMSO was developed for a one-step assay. The recoveries of TBBPA from urine samples via this one-step assay ranged from 96.7% to 109.9% and correlated well with a high-performance liquid chromatography-tandem mass spectroscopy (HPLC-MS/MS) method. It is expected that the dimerized fusion protein, VHH-AP, will show promising applications in human exposure and environmental monitoring.

  14. Design and Selection of a Camelid Single-Chain Antibody Yeast Two-Hybrid Library Produced De Novo for the Cap Protein of Porcine Circovirus Type 2 (PCV2)

    OpenAIRE

    Xiangjing Fu; Xiaolong Gao; Shengfang He; Di Huang; Peng Zhang; Xinglong Wang; Shuxia Zhang; Ruyi Dang; Shuanghui Yin; Enqi Du; Zengqi Yang

    2013-01-01

    Nanobodies (or variable domain of the heavy chain of the heavy-chain antibodies, VHHs) are single-domain antigen-binding fragments derived from camelid heavy chain antibodies. Their comparatively small size, monomeric behavior, high stability, high solubility, and ability to bind epitopes inaccessible to conventional antibodies make them especially suitable for many therapeutic and biotechnological applications. In this paper, for the first time, we created the immunized Camelus Bactrianus VH...

  15. Development of protective agent against Hottentotta saulcyi venom using camelid single-domain antibody.

    Science.gov (United States)

    Darvish, Maryam; Behdani, Mahdi; Shokrgozar, Mohammad Ali; Pooshang-Bagheri, Kamran; Shahbazzadeh, Delavar

    2015-12-01

    Hottentotta saulcyi, medically important scorpion species, causes some of harmful toxic exposure in Iran. Administrated, conventional antivenom-based immunotherapy is still limited and hardly meet ideal characteristic of effective treatment for scorpion envenomation. In this study we aimed to develop a neutralizing agent directed against scorpion venom based on VHH, variable domain of the Camelidae heavy chain antibody or Nanobody. This promising biomolecule is well-established as an advantageous tool for therapeutic purposes due to its small size, stability, monomeric performance and less immunogenicity. In this study, a large Nb library was constructed and phage displayed after successful camel immunization using H. saulcyi scorpion crude venom. After a series of biopanning rounds on Sephadex G50 purified venom fraction and screening by monoclonal phage ELISA, the best reactive Nb was retrieved and designated Nb12. The selected Nb was then expressed as soluble protein in Escherichia coli, purified and confirmed by SDS-PAGE analysis and western blotting. The lead candidate Nb12 bound scorpion venom with Kaff value of 5×10(7)M(-1). Nb12 was shown to be capable of neutralizing 2 LD50 of whole venom of scorpion toxin when injected in the ratio of the Nb/toxin of 1.4:1 into C57BL/6 mice. In challenge experiment, Nb succeeded to rescue all i.p. lethal dose injected mice even when administrated i.v., 20min after envenoming. These results with ease of production and superior neutralizing activity make Nb a suitable anti-toxin candidate for treatment of scorpion envenoming. PMID:26468036

  16. Inhibition of the Myotoxicity Induced by Bothrops jararacussu Venom and Isolated Phospholipases A2 by Specific Camelid Single-Domain Antibody Fragments.

    Directory of Open Access Journals (Sweden)

    Nidiane D R Prado

    Full Text Available Antivenoms, produced using animal hyperimmune plasma, remains the standard therapy for snakebites. Although effective against systemic damages, conventional antivenoms have limited efficacy against local tissue damage. Additionally, the hypersensitivity reactions, often elicited by antivenoms, the high costs for animal maintenance, the difficulty of producing homogeneous lots, and the instability of biological products instigate the search for innovative products for antivenom therapy. In this study, camelid antibody fragments (VHH with specificity to Bothropstoxin I and II (BthTX-I and BthTX-II, two myotoxic phospholipases from Bothrops jararacussu venom, were selected from an immune VHH phage display library. After biopanning, 28 and 6 clones recognized BthTX-I and BthTX-II by ELISA, respectively. Complementarity determining regions (CDRs and immunoglobulin frameworks (FRs of 13 VHH-deduced amino acid sequences were identified, as well as the camelid hallmark amino acid substitutions in FR2. Three VHH clones (KF498607, KF498608, and KC329718 were capable of recognizing BthTX-I by Western blot and showed affinity constants in the nanomolar range against both toxins. VHHs inhibited the BthTX-II phospholipase A2 activity, and when tested for cross-reactivity, presented specificity to the Bothrops genus in ELISA. Furthermore, two clones (KC329718 and KF498607 neutralized the myotoxic effects induced by B. jararacussu venom, BthTX-I, BthTX-II, and by a myotoxin from Bothrops brazili venom (MTX-I in mice. Molecular docking revealed that VHH CDRs are expected to bind the C-terminal of both toxins, essential for myotoxic activity, and to epitopes in the BthTX-II enzymatic cleft. Identified VHHs could be a biotechnological tool to improve the treatment for snake envenomation, an important and neglected world public health problem.

  17. Inhibition of the Myotoxicity Induced by Bothrops jararacussu Venom and Isolated Phospholipases A2 by Specific Camelid Single-Domain Antibody Fragments

    Science.gov (United States)

    Prado, Nidiane D. R.; Pereira, Soraya S.; da Silva, Michele P.; Morais, Michelle S. S.; Kayano, Anderson M.; Moreira-Dill, Leandro S.; Luiz, Marcos B.; Zanchi, Fernando B.; Fuly, André L.; E. F. Huacca, Maribel; Fernandes, Cleberson F.; Calderon, Leonardo A.; Zuliani, Juliana P.; Soares, Andreimar M.; Stabeli, Rodrigo G.; F. C. Fernandes, Carla

    2016-01-01

    Antivenoms, produced using animal hyperimmune plasma, remains the standard therapy for snakebites. Although effective against systemic damages, conventional antivenoms have limited efficacy against local tissue damage. Additionally, the hypersensitivity reactions, often elicited by antivenoms, the high costs for animal maintenance, the difficulty of producing homogeneous lots, and the instability of biological products instigate the search for innovative products for antivenom therapy. In this study, camelid antibody fragments (VHH) with specificity to Bothropstoxin I and II (BthTX-I and BthTX-II), two myotoxic phospholipases from Bothrops jararacussu venom, were selected from an immune VHH phage display library. After biopanning, 28 and 6 clones recognized BthTX-I and BthTX-II by ELISA, respectively. Complementarity determining regions (CDRs) and immunoglobulin frameworks (FRs) of 13 VHH-deduced amino acid sequences were identified, as well as the camelid hallmark amino acid substitutions in FR2. Three VHH clones (KF498607, KF498608, and KC329718) were capable of recognizing BthTX-I by Western blot and showed affinity constants in the nanomolar range against both toxins. VHHs inhibited the BthTX-II phospholipase A2 activity, and when tested for cross-reactivity, presented specificity to the Bothrops genus in ELISA. Furthermore, two clones (KC329718 and KF498607) neutralized the myotoxic effects induced by B. jararacussu venom, BthTX-I, BthTX-II, and by a myotoxin from Bothrops brazili venom (MTX-I) in mice. Molecular docking revealed that VHH CDRs are expected to bind the C-terminal of both toxins, essential for myotoxic activity, and to epitopes in the BthTX-II enzymatic cleft. Identified VHHs could be a biotechnological tool to improve the treatment for snake envenomation, an important and neglected world public health problem. PMID:27028872

  18. Inhibition of the Myotoxicity Induced by Bothrops jararacussu Venom and Isolated Phospholipases A2 by Specific Camelid Single-Domain Antibody Fragments.

    Science.gov (United States)

    Prado, Nidiane D R; Pereira, Soraya S; da Silva, Michele P; Morais, Michelle S S; Kayano, Anderson M; Moreira-Dill, Leandro S; Luiz, Marcos B; Zanchi, Fernando B; Fuly, André L; Huacca, Maribel E F; Fernandes, Cleberson F; Calderon, Leonardo A; Zuliani, Juliana P; Pereira da Silva, Luiz H; Soares, Andreimar M; Stabeli, Rodrigo G; Fernandes, Carla F C

    2016-01-01

    Antivenoms, produced using animal hyperimmune plasma, remains the standard therapy for snakebites. Although effective against systemic damages, conventional antivenoms have limited efficacy against local tissue damage. Additionally, the hypersensitivity reactions, often elicited by antivenoms, the high costs for animal maintenance, the difficulty of producing homogeneous lots, and the instability of biological products instigate the search for innovative products for antivenom therapy. In this study, camelid antibody fragments (VHH) with specificity to Bothropstoxin I and II (BthTX-I and BthTX-II), two myotoxic phospholipases from Bothrops jararacussu venom, were selected from an immune VHH phage display library. After biopanning, 28 and 6 clones recognized BthTX-I and BthTX-II by ELISA, respectively. Complementarity determining regions (CDRs) and immunoglobulin frameworks (FRs) of 13 VHH-deduced amino acid sequences were identified, as well as the camelid hallmark amino acid substitutions in FR2. Three VHH clones (KF498607, KF498608, and KC329718) were capable of recognizing BthTX-I by Western blot and showed affinity constants in the nanomolar range against both toxins. VHHs inhibited the BthTX-II phospholipase A2 activity, and when tested for cross-reactivity, presented specificity to the Bothrops genus in ELISA. Furthermore, two clones (KC329718 and KF498607) neutralized the myotoxic effects induced by B. jararacussu venom, BthTX-I, BthTX-II, and by a myotoxin from Bothrops brazili venom (MTX-I) in mice. Molecular docking revealed that VHH CDRs are expected to bind the C-terminal of both toxins, essential for myotoxic activity, and to epitopes in the BthTX-II enzymatic cleft. Identified VHHs could be a biotechnological tool to improve the treatment for snake envenomation, an important and neglected world public health problem. PMID:27028872

  19. A Camelid-derived Antibody Fragment Targeting the Active Site of a Serine Protease Balances between Inhibitor and Substrate Behavior

    DEFF Research Database (Denmark)

    Kromann-Hansen, Tobias; Oldenburg, Emil; Yung, Kristen Wing Yu;

    2016-01-01

    -ray crystal structure of a nanobody in complex with a serine protease. The nanobody displays a new type of interaction between an antibody and a serine protease as it inserts its CDR-H3 loop into the active site of the protease in a substrate-like manner. The unique binding mechanism causes the nanobody to...

  20. A Camelid-derived Antibody Fragment Targeting the Active Site of a Serine Protease Balances between Inhibitor and Substrate Behavior.

    Science.gov (United States)

    Kromann-Hansen, Tobias; Oldenburg, Emil; Yung, Kristen Wing Yu; Ghassabeh, Gholamreza H; Muyldermans, Serge; Declerck, Paul J; Huang, Mingdong; Andreasen, Peter A; Ngo, Jacky Chi Ki

    2016-07-15

    A peptide segment that binds the active site of a serine protease in a substrate-like manner may behave like an inhibitor or a substrate. However, there is sparse information on which factors determine the behavior a particular peptide segment will exhibit. Here, we describe the first x-ray crystal structure of a nanobody in complex with a serine protease. The nanobody displays a new type of interaction between an antibody and a serine protease as it inserts its complementary determining region-H3 loop into the active site of the protease in a substrate-like manner. The unique binding mechanism causes the nanobody to behave as a strong inhibitor as well as a poor substrate. Intriguingly, its substrate behavior is incomplete, as 30-40% of the nanobody remained intact and inhibitory after prolonged incubation with the protease. Biochemical analysis reveals that an intra-loop interaction network within the complementary determining region-H3 of the nanobody balances its inhibitor versus substrate behavior. Collectively, our results unveil molecular factors, which may be a general mechanism to determine the substrate versus inhibitor behavior of other protease inhibitors. PMID:27226628

  1. Generation of a phage-display library of single-domain camelid VH H antibodies directed against Chlamydomonas reinhardtii antigens, and characterization of VH Hs binding cell-surface antigens.

    Science.gov (United States)

    Jiang, Wenzhi; Rosenberg, Julian N; Wauchope, Akelia D; Tremblay, Jacqueline M; Shoemaker, Charles B; Weeks, Donald P; Oyler, George A

    2013-11-01

    Single-domain antibodies (sdAbs) are powerful tools for the detection, quantification, purification and subcellular localization of proteins of interest in biological research. We have generated camelid (Lama pacos) heavy chain-only variable VH domain (VH H) libraries against antigens in total cell lysates from Chlamydomonas reinhardtii. The sdAbs in the sera from immunized animals and VH H antibody domains isolated from the library show specificity to C. reinhardtii and lack of reactivity to antigens from four other algae: Chlorella variabilis, Coccomyxa subellipsoidea, Nannochloropsis oceanica and Thalassiosira pseudonana. Antibodies were produced against a diverse representation of antigens as evidenced by sera ELISA and protein-blot analyses. A phage-display library consisting of the VH H region contained at least 10(6) individual transformants, and thus should represent a wide range of C. reinhardtii antigens. The utility of the phage library was demonstrated by using live C. reinhardtii cells to pan for VH H clones with specific recognition of cell-surface epitopes. The lead candidate VH H clones (designated B11 and H10) bound to C. reinhardtii with EC50 values ≤ 0.5 nm. Treatment of cells with VH H B11 fused to the mCherry or green fluorescent proteins allowed brilliant and specific staining of the C. reinhardtii cell wall and analysis of cell-wall genesis during cell division. Such high-complexity VH H antibody libraries for algae will be valuable tools for algal researchers and biotechnologists.

  2. Design and selection of a camelid single-chain antibody yeast two-hybrid library produced de novo for the cap protein of porcine circovirus type 2 (PCV2.

    Directory of Open Access Journals (Sweden)

    Xiangjing Fu

    Full Text Available Nanobodies (or variable domain of the heavy chain of the heavy-chain antibodies, VHHs are single-domain antigen-binding fragments derived from camelid heavy chain antibodies. Their comparatively small size, monomeric behavior, high stability, high solubility, and ability to bind epitopes inaccessible to conventional antibodies make them especially suitable for many therapeutic and biotechnological applications. In this paper, for the first time, we created the immunized Camelus Bactrianus VHH yeast two-hybrid (Y2H library according to the Clontech Mate & Plate library construction system. The transformation efficiency and titer of the VHH Y2H library were 7.26×10(6 cfu/3 µg and 2×10(9 cfu/ml, which met the demand for Y2H library screening. Using as an example the porcine circovirus type 2 (PCV2 Cap protein as bait, we screened 21 positive Cap-specific VHH sequences. Among these sequences, 7 of 9 randomly selected clones were strongly positive as indicated by enzyme-linked immunosorbent assay, either using PCV2 viral lysis or purified Cap protein as coated antigen. Additionally, the immunocytochemistry results further indicated that the screened VHHs could specifically detected PCV2 in the infected cells. All this suggests the feasibility of in vivo VHH throughput screening based on Y2H strategy.

  3. Design and selection of a camelid single-chain antibody yeast two-hybrid library produced de novo for the cap protein of porcine circovirus type 2 (PCV2).

    Science.gov (United States)

    Fu, Xiangjing; Gao, Xiaolong; He, Shengfang; Huang, Di; Zhang, Peng; Wang, Xinglong; Zhang, Shuxia; Dang, Ruyi; Yin, Shuanghui; Du, Enqi; Yang, Zengqi

    2013-01-01

    Nanobodies (or variable domain of the heavy chain of the heavy-chain antibodies, VHHs) are single-domain antigen-binding fragments derived from camelid heavy chain antibodies. Their comparatively small size, monomeric behavior, high stability, high solubility, and ability to bind epitopes inaccessible to conventional antibodies make them especially suitable for many therapeutic and biotechnological applications. In this paper, for the first time, we created the immunized Camelus Bactrianus VHH yeast two-hybrid (Y2H) library according to the Clontech Mate & Plate library construction system. The transformation efficiency and titer of the VHH Y2H library were 7.26×10(6) cfu/3 µg and 2×10(9) cfu/ml, which met the demand for Y2H library screening. Using as an example the porcine circovirus type 2 (PCV2) Cap protein as bait, we screened 21 positive Cap-specific VHH sequences. Among these sequences, 7 of 9 randomly selected clones were strongly positive as indicated by enzyme-linked immunosorbent assay, either using PCV2 viral lysis or purified Cap protein as coated antigen. Additionally, the immunocytochemistry results further indicated that the screened VHHs could specifically detected PCV2 in the infected cells. All this suggests the feasibility of in vivo VHH throughput screening based on Y2H strategy. PMID:23469171

  4. Development and Utilization of Camelid VHH Antibodies from Alpaca for 2,2′,4,4′-Tetrabrominated Diphenyl Ether Detection

    OpenAIRE

    Bever, Candace R. S.; Majkova, Zuzana; Radhakrishnan, Rajeswaran; Suni, Ian; McCoy, Mark; Wang, Yanru; Dechant, Julie; Gee, Shirley; Hammock, Bruce D.

    2014-01-01

    An antibody-based analytical method for the detection of a chemical flame retardant using antibody fragments isolated from an alpaca has been developed. One specific chemical flame retardant congener, 2,2′,4,4′-tetrabrominated diphenyl ether (BDE-47), is often the major poly-BDE (PBDE) congener present in human and environmental samples and that which is the most frequently detected. An alpaca was immunized with a surrogate of BDE-47 covalently attached to a carrier protein. The resulting mRN...

  5. Algal chloroplast produced camelid VH H antitoxins are capable of neutralizing botulinum neurotoxin.

    Science.gov (United States)

    Barrera, Daniel J; Rosenberg, Julian N; Chiu, Joanna G; Chang, Yung-Nien; Debatis, Michelle; Ngoi, Soo-Mun; Chang, John T; Shoemaker, Charles B; Oyler, George A; Mayfield, Stephen P

    2015-01-01

    We have produced three antitoxins consisting of the variable domains of camelid heavy chain-only antibodies (VH H) by expressing the genes in the chloroplast of green algae. These antitoxins accumulate as soluble proteins capable of binding and neutralizing botulinum neurotoxin. Furthermore, they accumulate at up to 5% total soluble protein, sufficient expression to easily produce these antitoxins at scale from algae. The genes for the three different antitoxins were transformed into Chlamydomonas reinhardtii chloroplasts and their products purified from algae lysates and assayed for in vitro biological activity using toxin protection assays. The produced antibody domains bind to botulinum neurotoxin serotype A (BoNT/A) with similar affinities as camelid antibodies produced in Escherichia coli, and they are similarly able to protect primary rat neurons from intoxication by BoNT/A. Furthermore, the camelid antibodies were produced in algae without the use of solubilization tags commonly employed in E. coli. These camelid antibody domains are potent antigen-binding proteins and the heterodimer fusion protein containing two VH H domains was capable of neutralizing BoNT/A at near equimolar concentrations with the toxin. Intact antibody domains were detected in the gastrointestinal (GI) tract of mice treated orally with antitoxin-producing microalgae. These findings support the use of orally delivered antitoxins produced in green algae as a novel treatment for botulism.

  6. Construction of naïve camelids VHH repertoire in phage display-based library.

    Science.gov (United States)

    Sabir, Jamal S M; Atef, Ahmed; El-Domyati, Fotouh M; Edris, Sherif; Hajrah, Nahid; Alzohairy, Ahmed M; Bahieldin, Ahmed

    2014-04-01

    Camelids have unique antibodies, namely HCAbs (VHH) or commercially named Nanobodies(®) (Nb) that are composed only of a heavy-chain homodimer. As libraries based on immunized camelids are time-consuming, costly and likely redundant for certain antigens, we describe the construction of a naïve camelid VHHs library from blood serum of non-immunized camelids with affinity in the subnanomolar range and suitable for standard immune applications. This approach is rapid and recovers VHH repertoire with the advantages of being more diverse, non-specific and devoid of subpopulations of specific antibodies, which allows the identification of binders for any potential antigen (or pathogen). RNAs from a number of camelids from Saudi Arabia were isolated and cDNAs of the diverse vhh gene were amplified; the resulting amplicons were cloned in the phage display pSEX81 vector. The size of the library was found to be within the required range (10(7)) suitable for subsequent applications in disease diagnosis and treatment. Two hundred clones were randomly selected and the inserted gene library was either estimated for redundancy or sequenced and aligned to the reference camelid vhh gene (acc. No. ADE99145). Results indicated complete non-specificity of this small library in which no single event of redundancy was detected. These results indicate the efficacy of following this approach in order to yield a large and diverse enough gene library to secure the presence of the required version encoding the required antibodies for any target antigen. This work is a first step towards the construction of phage display-based biosensors useful in disease (e.g., TB or tuberculosis) diagnosis and treatment. PMID:24702893

  7. 驼源天然单域重链抗体库的构建与鉴定%Construction and Biopanning of Camelid Naive Single-domain Antibody Phage Display Library

    Institute of Scientific and Technical Information of China (English)

    涂追; 许杨; 刘夏; 何庆华; 陶勇

    2011-01-01

    从未经主动免疫的健康羊驼(Lamapacos)外周血淋巴细胞中提取总RNA,反转录后作为第一轮PCR的模板.根据重链抗体保守区域设计引物,经巢式PCR法扩增获得了全套重链抗体可变区基因,将其克隆至噬菌粒 pHENl,电转化大肠杆菌TG1得到初级抗体库NAL,含有2×10个独立克隆,菌落PCR和Hinf Ⅰ酶切分析结果显示,克隆效率大于97%,文库的多样性良好.辅助噬菌体救援后,得到噬菌体展示文库命名为NA-PDL,滴度达10CFU/ml.以真菌毒素人工抗原DON-MBSA为目标抗原,对NA-PDL进行了淘选,第二轮洗脱物中,阳性克隆率达36.4%,提示针对目标抗原的噬菌体颗粒得到了有效富集,文库NA-PDL多样性较好,为后续淘选针对特定抗原的单域重链抗体奠定了基础.%The objective is to construct a camelid na(i)ve single-domain heavy chain antibody phage display library. Total RNA was purified from 30ml blood of two healthy non-immune alpacas ( Lama pacos) and directly used for complementary DNA (cDNA) synthesis. Three sets of primers were designed based on the conserved region of heavy-chain antibody. The repertoire of VHH coding sequence was amplified by nested PCR, and the PCR products were cloned into a phagemid vector pHEN1. By electroporation of E. coli TG1 , the primary library (designate NAL) was obtained containing more than 107 independence clones. After helper phage rescue, the phage display library ( designate SNA-PDL) was generated with a titre up to 1013 CFU/ml. The library exhibited high diversity as judged by the Hinf Ⅰ restriction pattern. Solid phage biopanning against artificial antigen DONMBSA showed significant enrichment of binding phage particles. The positive rate of panning round two was 36.4% . The data indicated that a na(i)ve single-domain antibody phage display library was constructed. which has good diversity and would be useful for generating VHHs with specific binding affinity.

  8. Algal chloroplast produced camelid VHH antitoxins are capable of neutralizing botulinum neurotoxin

    OpenAIRE

    Daniel J Barrera; Rosenberg, Julian N.; Chiu, Joanna G.; Chang, Yung-Nien; Debatis, Michelle; Ngoi, Soo-Mun; Chang, John T.; Shoemaker, Charles B.; George A Oyler; Mayfield, Stephen P

    2014-01-01

    We have produced three antitoxins consisting of the variable domains of camelid heavy chain-only antibodies (VHH) by expressing the genes in the chloroplast of green algae. These antitoxins accumulate as soluble proteins capable of binding and neutralizing botulinum neurotoxin. Furthermore, they accumulate at up to 5% total soluble protein, sufficient expression to easily produce these antitoxins at scale from algae. The genes for the three different antitoxins were transformed into Chlamydom...

  9. Seminal plasma components in camelids and comparisons with other species.

    Science.gov (United States)

    Kershaw-Young, C M; Maxwell, W M C

    2012-08-01

    Camelid semen is characterized by a highly viscous, low-volume ejaculate with a low concentration of spermatozoa that exhibit low progressive motility. The viscous seminal plasma is currently the major impediment to the development of assisted reproductive technologies (ARTs) in camelids. To advance ARTs such as sperm cryopreservation and artificial insemination in camelids, it is necessary to identify the cause of the viscosity and gain an understanding of the role of seminal plasma components on sperm function and fertility. Numerous compounds and proteins have been identified as mediators of sperm function and predictors of fertility in other livestock species, and understanding the importance of specific proteins has progressed the success of ARTs in these species. Current knowledge on the components of camelid seminal plasma is outlined, together with the implications of these components for the development of ARTs in camelids. The cause of semen viscosity, as well as proteins that are present in camelid seminal plasma, is described for the first time. Seminal plasma components are compared with those of other species to hypothesize their role in sperm function and fertility. PMID:22827394

  10. Camelid antivenom development and potential in vivo neutralization of Hottentotta saulcyi scorpion venom.

    Science.gov (United States)

    Darvish, Maryam; Ebrahimi, Soltan Ahmad; Shahbazzadeh, Delavar; Bagheri, Kamran-Pooshang; Behdani, Mahdi; Shokrgozar, Mohammad Ali

    2016-04-01

    Scorpion envenoming is a serious health problem which can cause a variety of clinical toxic effects. Of the many scorpion species native to Iran, Hottentotta saulcyi is important because its venom can produce toxic effects in man. Nowadays, antivenom derived from hyper immune horses is the only effective treatment for sever scorpion stings. Current limitations of immunotherapy urgently require an efficient alternative with high safety, target affinity and more promising venom neutralizing capability. Recently, heavy chain-only antibodies (HC-Abs) found naturally in camelid serum met the above mentioned advantages. In this study, immuno-reactivities of polyclonal antibodies were tested after successful immunization of camel using H. saulcyi scorpion crude venom. The lethal potency of scorpion venom in C57BL/6 mice injected intraperitoneally was determined to be 2.7 mg/kg. These results were followed by the efficient neutralization of lethal activity of H. saulcyi scorpion venom by injection of antivenom and purified IgG fractions into mice intraperitonelly or intravenously, respectively. HC-Ab camelid antivenom could be considered as a useful serotherapeutics instead of present treatment for scorpion envenomation. PMID:26809016

  11. Advances in sarcocystiosis diagnosis in South American camelids in Peru

    International Nuclear Information System (INIS)

    Full text: Sarcocystiosis in south-American camelids has a high prevalence, about 94% and affects alpacas and llamas by producing cysts either in skeletal muscle or heart muscle and is caused aucheniae and S. lamacanis. The approach of this investigation was to identify the species involves their pathogenicity, protein structure and the antibody kinetics of animals raised under field conditions, in isolation and under experimental infection and immunization by ELISA and Western blot analysis. Also a PCR assay was developed for diagnosis of the disease in live animals, because so far this parasitic disease is diagnosed at the post mortem examination in the slaughterhouse. PCR product was sequenced and registered in the GeneBank for S. lamacanis. To learn about antibody kinetics under field conditions a group of 30 alpacas and their offspring raised in the Andean region of Huancavelica was sampled. Blood samples were collected and assayed by ELISA for 6 months after lambing. Similarly, a group of alpaca crias kept in isolation in Lima was monitored for antibodies. A group of 20 pregnant alpacas raised in the Andean region of Puno was transferred to our station in Lima and raised in sarcocystis-free pens. These alpacas were monitored for antibody to sarcocystis by ELISA and blood parasitemia by PCR for 2 years. A group of alpaca crias was immunized using a bradyzoite protein suspension, and blood samples were collected for monitoring antibody by ELISA and western blot. In order to identify genes coding surface antigens, bradkyzoites of S. aucheniae were collected for total RNA and mRNA isolation. From this mRNA a cDNA library will be constructed, using ligation, transformation and sequencing. Alpacas from Huancavelica assayed by ELISA showed antibody (93.3%) up to 4 months after parturition, and then 100% of them were positive. This result indicates the high rate of the infection in field conditions. A 30% of their offspring showed antibody up to 2 months of age being

  12. Stability of llama heavy chain antibody fragments under extreme conditions

    NARCIS (Netherlands)

    Dolk, E.

    2004-01-01

    Camelids have next to their normal antibodies, a unique subset of antibodies lacking light chains. The resulting single binding domain, VHH, of these heavy chain antibodies consequently have unique properties. A high stability is one of these properties, which was investigated in this thesis. The a

  13. Biochemical adaptation of camelids during periods where feed is withheld

    Directory of Open Access Journals (Sweden)

    J. Wensvoort

    2001-03-01

    Full Text Available Biochemical changes during fasting or the withholding of feed for 5 day were studied in serum of camelids (dromedary camel, llama and ruminants (sheep, steers. Camels maintained low levels of 13-hydroxybutyrate (BHB and high levels of glucose but showed some increased levels of non-esterified fatty acid (NEFA and urea when fasting. Sheep and steers showed a rise in serum BHB and much higher increases of NEFA than camels and llamas. Sheep showed decreased serum glucose. The llama showed some increase in BHB but NEFA was lower than the other three species. The results indicate that camelids have a unique ability to control lipolytic and gluconeogenic activity to prevent or postpone the state of ketosis. Understanding and manipulation of these metabolic mechanisms in cattle and sheep could have great benefit to the livestock industry.

  14. Camelid-derived heavy-chain nanobody against Clostridium botulinum neurotoxin E in Pichia pastoris.

    Science.gov (United States)

    Baghban, Roghayyeh; Gargari, Seyed Latif Mousavi; Rajabibazl, Masoumeh; Nazarian, Shahram; Bakherad, Hamid

    2016-01-01

    Botulinum neurotoxins (BoNTs) result in severe and often fatal disease, botulism. Common remedial measures such as equine antitoxin and human botulism immunoglobulin in turn are problematic and time-consuming. Therefore, diagnosis and therapy of BoNTs are vital. The variable domain of heavy-chain antibodies (VHH) has unique features, such as the ability to identify and bind specifically to target epitopes and ease of production in bacteria and yeast. The Pichia pastoris is suitable for expression of recombinant antibody fragments. Disulfide bond formation and correct folds of protein with a high yield are some of the advantages of this eukaryotic host. In this study, we have expressed and purified the camelid VHH against BoNT/E in P. pastoris. The final yield of P. pastoris-expressed antibody was estimated to be 16 mg/l, which is higher than that expressed by Escherichia coli. The nanobody expressed in P. pastoris neutralized 4LD50 of the BoNT/E upon i.p. injection in 25% of mice. The nanobody expressed in E. coli extended the mice's survival to 1.5-fold compared to the control. This experiment indicated that the quality of expressed protein in the yeast is superior to that of the bacterial expression. Favorable protein folding by P. pastoris seems to play a role in its better toxin-binding property. PMID:24673401

  15. Biomechanical analysis of the camelid cervical intervertebral disc

    Directory of Open Access Journals (Sweden)

    Dean K. Stolworthy

    2015-01-01

    Full Text Available Chronic low back pain (LBP is a prevalent global problem, which is often correlated with degenerative disc disease. The development and use of good, relevant animal models of the spine may improve treatment options for this condition. While no animal model is capable of reproducing the exact biology, anatomy, and biomechanics of the human spine, the quality of a particular animal model increases with the number of shared characteristics that are relevant to the human condition. The purpose of this study was to investigate the camelid (specifically, alpaca and llama cervical spine as a model of the human lumbar spine. Cervical spines were obtained from four alpacas and four llamas and individual segments were used for segmental flexibility/biomechanics and/or morphology/anatomy studies. Qualitative and quantitative data were compared for the alpaca and llama cervical spines, and human lumbar specimens in addition to other published large animal data. Results indicate that a camelid cervical intervertebral disc (IVD closely approximates the human lumbar disc with regard to size, spinal posture, and biomechanical flexibility. Specifically, compared with the human lumbar disc, the alpaca and llama cervical disc size are approximately 62%, 83%, and 75% with regard to area, depth, and width, respectively, and the disc flexibility is approximately 133%, 173%, and 254%, with regard to range of motion (ROM in axial-rotation, flexion-extension, and lateral-bending, respectively. These results, combined with the clinical report of disc degeneration in the llama lower cervical spine, suggest that the camelid cervical spine is potentially well suited for use as an animal model in biomechanical studies of the human lumbar spine.

  16. Rotavirus strains in neglected animal species including lambs, goats and camelids

    OpenAIRE

    Papp, Hajnalka; Yashpal S. Malik; Farkas, Szilvia L.; Jakab, Ferenc; Martella, Vito; Bányai, Krisztián

    2014-01-01

    Surveillance of rotavirus infections and circulating strains in small ruminants (i.e. lambs, goats and camelids) has been a neglected research area in the past. However, recent years that have seen an intensification of surveillance in humans and livestock animals, where vaccines to reduce disease burden caused by Rotavirus A (RVA) are available, led to the efforts to better understand the epidemiology, ecology and evolution of RVA strains in other hosts, including lambs, goats and camelids. ...

  17. [The occurrence of "Candidatus Mycoplasma haemolamae" infections in clinically asymptomatic South American Camelids in Austria].

    Science.gov (United States)

    Franz, Sonja; Spergser, Joachim; Schwendenwein, Ilse; Stanitznig, Anna; Lambacher, Bianca; Tichy, Alexander; Wittek, Thomas

    2016-01-01

    Reports of CMhl infections in South American Camelids in Europe are only available from the United Kingdom and Switzerland. Knowing that CMhl infections can lead to severe disease resulting in death if combined with other diseases or stress, it was the aim of this study to assess prevalence data from camelids in Austria. In comparison to the previous studies a representative number of camelids was investigated nationwide. Data were assessed due to differences in geographical region, age, sex, species, and origin. A relatively high prevalence of 25.8% was recorded. CMhl was detected significantly more often in alpacas (Vicunja pacos) than in llamas (Lama glama) and more frequently in animals younger than 2 years. Additionally regional differences have been observed, which might be due to climatic differences and/or variations in insect vectors. In this study apperantly clinical healthy animals were shown to be infected with CMhl. Camelids infected with CMhl are a pathogen reservoir. The results of this study indicate different risk levels of infection between llamas and alpacas and between younger and older animals. The data presented underline the necessity of further studies on CMhlI infections in South American Camelids.

  18. Semen preservation and artificial insemination in domesticated South American camelids.

    Science.gov (United States)

    Bravo, P Walter; Alarcon, V; Baca, L; Cuba, Y; Ordoñez, C; Salinas, J; Tito, F

    2013-01-10

    Semen preservation and artificial insemination in South American camelids are reviewed giving emphasis to work done in Peru and by the authors. Reports on semen evaluation and the preservation process indicate that semen of alpacas and llamas can be manipulated by making it liquid first. Collagenase appears to be the best enzyme to eliminate viscosity. Tris buffer solution maintains a higher motility than egg-yolk citrate, phosphate buffered saline (PBS), Triladyl, and Merck-I extenders. Cooling of semen took 1h after collected, and equilibrated with 7% glycerol presented a better motility and spermatozoa survival at 1, 7, 15 and 30days after being slowly frozen in 0.25mL plastic straws. Trials of artificial insemination with freshly diluted semen and frozen-thawed semen are encouraging and needs to be tested extensively under field conditions. Recently, fertility rates varied from 3 to 67%. Semen preservation and most important, artificial insemination appear to be a reality, and could be used to improve the genetic quality of alpacas and llamas. PMID:23153624

  19. Rotavirus strains in neglected animal species including lambs, goats and camelids.

    Science.gov (United States)

    Papp, Hajnalka; Malik, Yashpal S; Farkas, Szilvia L; Jakab, Ferenc; Martella, Vito; Bányai, Krisztián

    2014-01-01

    Surveillance of rotavirus infections and circulating strains in small ruminants (i.e. lambs, goats and camelids) has been a neglected research area in the past. However, recent years that have seen an intensification of surveillance in humans and livestock animals, where vaccines to reduce disease burden caused by Rotavirus A (RVA) are available, led to the efforts to better understand the epidemiology, ecology and evolution of RVA strains in other hosts, including lambs, goats and camelids. The aim of this review is to provide an update of the epidemiology and strain diversity of RV strains in these species through searching for relevant information in public data bases. PMID:25674588

  20. Culicoides fauna and bluetongue virus serotype 8 infection in South American camelid herds in Germany

    OpenAIRE

    Schulz, Claudia

    2012-01-01

    Bluetongue (BT) is a Culicoides-born infectious disease caused by bluetongue virus (BTV). From 2006 to 2010, BTV serotype 8 (BTV-8) spread throughout Europe, causing severe disease in domestic and some wild ruminant species and in an alpaca. Compulsory vaccination of susceptible animals was the most effective strategy to control and eradicate the BTV-8 epizootic in Europe. However, South American camelids (SAC) were not included in the BTV-8 vaccination programmes in Europe. The presented...

  1. Competitive Selection from Single Domain Antibody Libraries Allows Isolation of High-Affinity Antihapten Antibodies That Are Not Favored in the llama Immune Response

    OpenAIRE

    Rosa, Sofia Tabares-da; Rossotti, Martin; Carleiza, Carmen; Carrión, Federico; Pritsch, Otto; Ahn, Ki Chang; Last, Jerold A.; Hammock, Bruce D.; González-Sapienza, Gualberto

    2011-01-01

    Single-domain antibodies (sdAbs) found in camelids, lack a light chain and their antigen-binding site sits completely in the heavy-chain variable domain (VHH). Their simplicity, thermostability, and ease in expression have made VHHs highly attractive. While this has been successfully exploited for macromolecular antigens, their application to the detection of small molecules is still limited to a very few reports, mostly describing low affinity VHHs. Using triclocarban (TCC) as a model hapten...

  2. Prospección serológica del virus parainfluenza 3 en camélidos sudamericanos en Chile A serological survey of parainfluenza-3 virus in South American camelids from Chile

    Directory of Open Access Journals (Sweden)

    CP Cepeda

    2011-01-01

    Full Text Available En un total de 370 sueros obtenidos de 87 alpacas (Lama pacos, 88 llamas (Llama glama, 89 guanacos (Lama guanicoe y 106 vicuñas (Vicugna vicugna se investigó la presencia de anticuerpos para el virus parainfluenza 3 (VPI-3. La prueba de inhibición de la hemoaglutinación mostró que 91 (24,6% de los camélidos sudamericanos (CSA distribuidos en 18 (20,7% alpacas, 36 (40,9% llamas, 27 (18,9% vicuñas y 10 (11,2% guanacos tenían anticuerpos. El rango de los títulos de las muestras positivas estuvo entre 8 y 256 con una media geométrica de 31. Los resultados establecen que los CSA en Chile están infectados con VPI-3.A total of 370 sera collected from 87 alpacas (Lama pacos, 88 llamas (Lama glama, 89 guanacos (Lama guanicoe and 106 vicunas (Vicugna vicugna were investigated for the presence of antibodies to parainfluenza 3 virus (PIV-3. Hemagglutination inhibition test indicated that a total of 91 (24.6% South American camelids (SAC had PIV-3 antibodies, including 18 (20.7% alpacas, 36 (40.9% llamas, 27 (18.9% vicunas and 10 (11.2% guanacos. Titers of the positive samples ranged from 8 to 256 with a geometric mean of 31. These results establish that SAC in Chile are infected with PIV-3.

  3. Genetically engineered T cells bearing chimeric nanoconstructed receptors harboring TAG-72-specific camelid single domain antibodies as targeting agents

    DEFF Research Database (Denmark)

    Sharifzadeh, Zahra; Rahbarizadeh, Fatemeh; Shokrgozar, Mohammad A;

    2013-01-01

    Despite the preclinical success of adoptive therapy with T cells bearing chimeric nanoconstructed antigen receptors (CARs), certain limitations of this therapeutic approach such as the immunogenicity of the antigen binding domain, the emergence of tumor cell escape variants and the blocking...

  4. Optimizing selection of large animals for antibody production by screening immune response to standard vaccines.

    Science.gov (United States)

    Thompson, Mary K; Fridy, Peter C; Keegan, Sarah; Chait, Brian T; Fenyö, David; Rout, Michael P

    2016-03-01

    Antibodies made in large animals are integral to many biomedical research endeavors. Domesticated herd animals like goats, sheep, donkeys, horses and camelids all offer distinct advantages in antibody production. However, their cost of use is often prohibitive, especially where poor antigen response is commonplace; choosing a non-responsive animal can set a research program back or even prevent experiments from moving forward entirely. Over the course of production of antibodies from llamas, we found that some animals consistently produced a higher humoral antibody response than others, even to highly divergent antigens, as well as to their standard vaccines. Based on our initial data, we propose that these "high level responders" could be pre-selected by checking antibody titers against common vaccines given to domestic farm animals. Thus, time and money can be saved by reducing the chances of getting poor responding animals and minimizing the use of superfluous animals.

  5. Optimizing selection of large animals for antibody production by screening immune response to standard vaccines.

    Science.gov (United States)

    Thompson, Mary K; Fridy, Peter C; Keegan, Sarah; Chait, Brian T; Fenyö, David; Rout, Michael P

    2016-03-01

    Antibodies made in large animals are integral to many biomedical research endeavors. Domesticated herd animals like goats, sheep, donkeys, horses and camelids all offer distinct advantages in antibody production. However, their cost of use is often prohibitive, especially where poor antigen response is commonplace; choosing a non-responsive animal can set a research program back or even prevent experiments from moving forward entirely. Over the course of production of antibodies from llamas, we found that some animals consistently produced a higher humoral antibody response than others, even to highly divergent antigens, as well as to their standard vaccines. Based on our initial data, we propose that these "high level responders" could be pre-selected by checking antibody titers against common vaccines given to domestic farm animals. Thus, time and money can be saved by reducing the chances of getting poor responding animals and minimizing the use of superfluous animals. PMID:26775851

  6. Leptospirosis en camélidos sudamericanos. Estudio de prevalencia serológica en distintas regiones de la Argentina Leptospirosis in south-american camelids. A study on the serological prevalence in different regions of Argentina

    Directory of Open Access Journals (Sweden)

    P. LLORENTE

    2002-01-01

    Full Text Available Se evaluó la seroprevalencia en Camélidos Sudamericanos de Leptospirosis, zoonosis de distribución mundial, producida por espiroquetas patógenas del género Leptospira. Se examinaron 494 animales (llamas, guanacos y vicuñas, clínicamente sanos sin vacunar, de diferentes regiones geográficas de la República Argentina. Se utilizó la técnica, serovar específica, de microaglutinación (MAT. El estudio reveló prevalencias entre 47.3 y 96.2% en llamas, entre 0 y 13% en guanacos y entre 9 y 62.8% en vicuñas. De los serovares que se usaron como antígeno en las determinaciones, los que más frecuentemente reaccionaron con los anticuerpos séricos de los camélidos, fueron copenhageni y castellonisLeptospirosis is a zoonotic infectious disease, affecting wild and domestic animals and human beings, caused by pathogenic spirochetes, spread world wide, belonging to the genus Leptospira. It is transmited by direct contact with infected animal urine or tissues, and indirectly through contaminated water and soil. Leptospirosis has a negative economic impact on porcine and bovine productions. It causes abortions, stillbirths, placental retention, infertility and chronical renal deficiency, causing disturbance of flow milk and quality in dairy cattle. Studies on south-american camelids productive aspects, have increased during the last decades, in order to promote alternative regional economies. There exist three species in Argentina, llama (Lama glama, guanaco (Lama guanicoe and vicuña (Vicugna vicugna. The knowledge of physiological parameters and susceptibility and immune response to infectious agents of these animals, are required to improve their breeding efficiency. Leptospirosis is an infectious disease, which may affect reproduction efficiency. Leptospira antibody prevalence in 494 sera obtained from healthy non vaccinated llamas, vicuñas and guanacos from different geographic zones in Argentina, was evaluated. The serovar specific

  7. Persistent Bovine Viral Diarrhea Virus infection in domestic and wild small ruminants and camelids including the mountain goat (Oreamnos americanus

    Directory of Open Access Journals (Sweden)

    Danielle Darracq Nelson

    2016-01-01

    Full Text Available Bovine viral diarrhea virus (BVDV is a Pestivirus best known for causing a variety of disease syndromes in cattle, including gastrointestinal disease, reproductive insufficiency, immunosuppression, mucosal disease, and hemorrhagic syndrome. The virus can be spread by transiently infected individuals and by persistently infected animals that may be asymptomatic while shedding large amounts of virus throughout their lifetime. BVDV has been reported in over 40 domestic and free-ranging species, and persistent infection has been described in eight of those species: white-tailed deer, mule deer, eland, mousedeer, mountain goats, alpacas, sheep, and domestic swine. This paper reviews the various aspects of BVDV transmission, disease syndromes, diagnosis, control, and prevention, as well as examines BVDV infection in domestic and wild small ruminants and camelids including mountain goats (Oreamnos americanus.

  8. Antibody Persistence in Young Children 5 Years after Vaccination with a Combined Haemophilus influenzae Type b-Neisseria meningitidis Serogroup C Conjugate Vaccine Coadministered with Diphtheria-Tetanus-Acellular Pertussis-Based and Pneumococcal Conjugate Vaccines

    Science.gov (United States)

    Tejedor, Juan Carlos; Brzostek, Jerzy; Konior, Ryszard; Grunert, Detlef; Kolhe, Devayani; Baine, Yaela

    2016-01-01

    We evaluated antibody persistence in children up to 5 years after administration of a combined Haemophilus influenzae type b (Hib)-Neisseria meningitidis serogroup C (MenC)-tetanus toxoid (TT) conjugate vaccine coadministered with a pneumococcal conjugate vaccine. This is the follow-up study of a randomized trial (ClinicalTrials.gov registration no. NCT00334334/00463437) in which healthy children were vaccinated (primary vaccinations at 2, 4, and 6 months of age and booster vaccination at 11 to 18 months of age) with Hib-MenC-TT or a control MenC conjugate vaccine, coadministered with diphtheria-tetanus-acellular pertussis (DTPa)-based combination vaccines (DTPa/Hib for control groups) and a pneumococcal conjugate vaccine (10-valent pneumococcal nontypeable H. influenzae protein D conjugate vaccine [PHiD-CV] or 7-valent cross-reacting material 197 [CRM197] conjugate vaccine [7vCRM]). MenC antibody titers were measured with a serum bactericidal antibody (SBA) assay using rabbit complement (i.e., rabbit SBA [rSBA]), and antibodies against Hib polyribosylribitol phosphate (PRP) were measured with an enzyme-linked immunosorbent assay. Antibody persistence up to 5 years after booster vaccination is reported for 530 children ∼6 years of age. The percentages of children with seroprotective rSBA-MenC titers were between 24.2% and 40.1% in all groups approximately 5 years after booster vaccination. More than 98.5% of children in each group retained seroprotective anti-PRP concentrations. No vaccine-related serious adverse events and no events related to a lack of vaccine efficacy were reported. Approximately 5 years after booster vaccination, the majority of children retained seroprotective anti-PRP antibody concentrations. The percentage of children retaining seroprotective rSBA-MenC titers was low (≤40%), suggesting that a significant proportion of children may be unprotected against MenC disease. (This study has been registered at ClinicalTrials.gov under

  9. Antibody Persistence in Young Children 5 Years after Vaccination with a Combined Haemophilus influenzae Type b-Neisseria meningitidis Serogroup C Conjugate Vaccine Coadministered with Diphtheria-Tetanus-Acellular Pertussis-Based and Pneumococcal Conjugate Vaccines.

    Science.gov (United States)

    Tejedor, Juan Carlos; Brzostek, Jerzy; Konior, Ryszard; Grunert, Detlef; Kolhe, Devayani; Baine, Yaela; Van Der Wielen, Marie

    2016-07-01

    We evaluated antibody persistence in children up to 5 years after administration of a combined Haemophilus influenzae type b (Hib)-Neisseria meningitidis serogroup C (MenC)-tetanus toxoid (TT) conjugate vaccine coadministered with a pneumococcal conjugate vaccine. This is the follow-up study of a randomized trial (ClinicalTrials.gov registration no. NCT00334334/00463437) in which healthy children were vaccinated (primary vaccinations at 2, 4, and 6 months of age and booster vaccination at 11 to 18 months of age) with Hib-MenC-TT or a control MenC conjugate vaccine, coadministered with diphtheria-tetanus-acellular pertussis (DTPa)-based combination vaccines (DTPa/Hib for control groups) and a pneumococcal conjugate vaccine (10-valent pneumococcal nontypeable H. influenzae protein D conjugate vaccine [PHiD-CV] or 7-valent cross-reacting material 197 [CRM197] conjugate vaccine [7vCRM]). MenC antibody titers were measured with a serum bactericidal antibody (SBA) assay using rabbit complement (i.e., rabbit SBA [rSBA]), and antibodies against Hib polyribosylribitol phosphate (PRP) were measured with an enzyme-linked immunosorbent assay. Antibody persistence up to 5 years after booster vaccination is reported for 530 children ∼6 years of age. The percentages of children with seroprotective rSBA-MenC titers were between 24.2% and 40.1% in all groups approximately 5 years after booster vaccination. More than 98.5% of children in each group retained seroprotective anti-PRP concentrations. No vaccine-related serious adverse events and no events related to a lack of vaccine efficacy were reported. Approximately 5 years after booster vaccination, the majority of children retained seroprotective anti-PRP antibody concentrations. The percentage of children retaining seroprotective rSBA-MenC titers was low (≤40%), suggesting that a significant proportion of children may be unprotected against MenC disease. (This study has been registered at ClinicalTrials.gov under

  10. Antibody Persistence in Young Children 5 Years after Vaccination with a Combined Haemophilus influenzae Type b-Neisseria meningitidis Serogroup C Conjugate Vaccine Coadministered with Diphtheria-Tetanus-Acellular Pertussis-Based and Pneumococcal Conjugate Vaccines.

    Science.gov (United States)

    Tejedor, Juan Carlos; Brzostek, Jerzy; Konior, Ryszard; Grunert, Detlef; Kolhe, Devayani; Baine, Yaela; Van Der Wielen, Marie

    2016-07-01

    We evaluated antibody persistence in children up to 5 years after administration of a combined Haemophilus influenzae type b (Hib)-Neisseria meningitidis serogroup C (MenC)-tetanus toxoid (TT) conjugate vaccine coadministered with a pneumococcal conjugate vaccine. This is the follow-up study of a randomized trial (ClinicalTrials.gov registration no. NCT00334334/00463437) in which healthy children were vaccinated (primary vaccinations at 2, 4, and 6 months of age and booster vaccination at 11 to 18 months of age) with Hib-MenC-TT or a control MenC conjugate vaccine, coadministered with diphtheria-tetanus-acellular pertussis (DTPa)-based combination vaccines (DTPa/Hib for control groups) and a pneumococcal conjugate vaccine (10-valent pneumococcal nontypeable H. influenzae protein D conjugate vaccine [PHiD-CV] or 7-valent cross-reacting material 197 [CRM197] conjugate vaccine [7vCRM]). MenC antibody titers were measured with a serum bactericidal antibody (SBA) assay using rabbit complement (i.e., rabbit SBA [rSBA]), and antibodies against Hib polyribosylribitol phosphate (PRP) were measured with an enzyme-linked immunosorbent assay. Antibody persistence up to 5 years after booster vaccination is reported for 530 children ∼6 years of age. The percentages of children with seroprotective rSBA-MenC titers were between 24.2% and 40.1% in all groups approximately 5 years after booster vaccination. More than 98.5% of children in each group retained seroprotective anti-PRP concentrations. No vaccine-related serious adverse events and no events related to a lack of vaccine efficacy were reported. Approximately 5 years after booster vaccination, the majority of children retained seroprotective anti-PRP antibody concentrations. The percentage of children retaining seroprotective rSBA-MenC titers was low (≤40%), suggesting that a significant proportion of children may be unprotected against MenC disease. (This study has been registered at ClinicalTrials.gov under

  11. Thyroid Antibodies

    Science.gov (United States)

    ... be limited. Home Visit Global Sites Search Help? Thyroid Antibodies Share this page: Was this page helpful? Also known as: Thyroid Autoantibodies; Antithyroid Antibodies; Antimicrosomal Antibody; Thyroid Microsomal Antibody; ...

  12. Selection and characterization of camelid nanobodies towards urokinase-type plasminogen activator

    DEFF Research Database (Denmark)

    Kaczmarek, Jakub; Skottrup, Peter Durand

    2015-01-01

    , increased cancer malignancy and poor survival prognosis. For these reasons uPA is considered an important target for anticancer drug therapy. In this study we isolated two camel single domain antibodies (nanobodies) from a naïve library by phage display. The nanobody sequences were sequence...

  13. Importance of Hypervariable Region 2 for Stability and Affinity of a Shark Single-Domain Antibody Specific for Ebola Virus Nucleoprotein.

    Science.gov (United States)

    Anderson, George P; Teichler, Daniel D; Zabetakis, Dan; Shriver-Lake, Lisa C; Liu, Jinny L; Lonsdale, Stephen G; Goodchild, Sarah A; Goldman, Ellen R

    2016-01-01

    Single-domain antibodies derived from the unique New Antigen Receptor found in sharks have numerous potential applications, ranging from diagnostic reagents to therapeutics. Shark-derived single-domain antibodies possess the same characteristic ability to refold after heat denaturation found in single-domain antibodies derived from camelid heavy-chain-only antibodies. Recently, two shark derived single-domain antibodies specific for the nucleoprotein of Ebola virus were described. Our evaluation confirmed their high affinity for the nucleoprotein, but found their melting temperatures to be low relative to most single-domain antibodies. Our first approach towards improving their stability was grafting antigen-binding regions (complementarity determining regions) of one of these single-domain antibodies onto a high melting temperature shark single-domain antibody. This resulted in two variants: one that displayed excellent affinity with a low melting temperature, while the other had poor affinity but a higher melting temperature. These new proteins, however, differed in only 3 amino acids within the complementarity determining region 2 sequence. In shark single-domain antibodies, the complementarity determining region 2 is often referred to as hypervariable region 2, as this segment of the antibody domain is truncated compared to the sequence in camelid single-domain antibodies and conventional heavy chain variable domains. To elucidate which of the three amino acids or combinations thereof were responsible for the affinity and stability we made the 6 double and single point mutants that covered the intermediates between these two clones. We found a single amino acid change that achieved a 10°C higher melting temperature while maintaining sub nM affinity. This research gives insights into the impact of the shark sdAb hypervariable 2 region on both stability and affinity. PMID:27494523

  14. Importance of Hypervariable Region 2 for Stability and Affinity of a Shark Single-Domain Antibody Specific for Ebola Virus Nucleoprotein

    Science.gov (United States)

    Anderson, George P.; Teichler, Daniel D.; Zabetakis, Dan; Shriver-Lake, Lisa C.; Liu, Jinny L.; Lonsdale, Stephen G.; Goodchild, Sarah A.; Goldman, Ellen R.

    2016-01-01

    Single-domain antibodies derived from the unique New Antigen Receptor found in sharks have numerous potential applications, ranging from diagnostic reagents to therapeutics. Shark-derived single-domain antibodies possess the same characteristic ability to refold after heat denaturation found in single-domain antibodies derived from camelid heavy-chain-only antibodies. Recently, two shark derived single-domain antibodies specific for the nucleoprotein of Ebola virus were described. Our evaluation confirmed their high affinity for the nucleoprotein, but found their melting temperatures to be low relative to most single-domain antibodies. Our first approach towards improving their stability was grafting antigen-binding regions (complementarity determining regions) of one of these single-domain antibodies onto a high melting temperature shark single-domain antibody. This resulted in two variants: one that displayed excellent affinity with a low melting temperature, while the other had poor affinity but a higher melting temperature. These new proteins, however, differed in only 3 amino acids within the complementarity determining region 2 sequence. In shark single-domain antibodies, the complementarity determining region 2 is often referred to as hypervariable region 2, as this segment of the antibody domain is truncated compared to the sequence in camelid single-domain antibodies and conventional heavy chain variable domains. To elucidate which of the three amino acids or combinations thereof were responsible for the affinity and stability we made the 6 double and single point mutants that covered the intermediates between these two clones. We found a single amino acid change that achieved a 10°C higher melting temperature while maintaining sub nM affinity. This research gives insights into the impact of the shark sdAb hypervariable 2 region on both stability and affinity. PMID:27494523

  15. Successful use of camelid (alpaca) antivenom to treat a potentially lethal tiger snake (Notechis scutatus) envenomation in a dog.

    Science.gov (United States)

    Padula, Andrew M; Winkel, Kenneth D

    2016-05-01

    This report describes a confirmed clinical case of tiger snake (Notechis scutatus) envenomation in a domestic dog that was successfully treated with a novel polyvalent camelid (alpaca; Llama pacos) antivenom. Samples collected from the dog were assayed for tiger snake venom (TSV) using a highly sensitive and specific ELISA. The TSV concentration in serum and urine at initial presentation was 365 ng/mL and 11,640 ng/mL respectively. At the time of initial presentation whole blood collected from the dog did not clot and the Prothrombin Time was abnormally increased (>300 s). Serum was also visibly hemolysed. The dog was administered antihistamine, dexamethasone and 4000 Units (sufficient to neutralise 40 mg of TSV) of a novel polyvalent alpaca antivenom diluted in 0.9% NaCl. At 4 h post-antivenom treatment the dog's clinical condition had improved markedly with serum TSV concentrations below the limit of detection (antivenom. Coagulation parameters had begun to improve by 4 h and had fully normalised by 16 h post-antivenom. Venom concentrations in both serum and urine remained undetectable at 16 h post-antivenom. The dog made a complete recovery, without complications, suggesting that the alpaca-based antivenom is both clinically safe and effective. PMID:26930223

  16. OSTEOMETRÍA Y GENÉTICA DE LOS CAMÉLIDOS MOCHICA, COSTA NORTE DEL PERÚ / Mochica camelids osteometry and genetics, north coast of Peru

    Directory of Open Access Journals (Sweden)

    Víctor Félix Vásquez Sánchez

    2009-12-01

    Full Text Available El objetivo de este trabajo es evaluar los resultados de la osteometría a dos muestras óseas de camélidos de sitios de la época Mochica (siglos I y VII d.C. en la costa norte de Perú. Los huesos analizados en este estudio son primeras falanges que provienen de dos sitios arqueológicos Mochica (Cerro Chepén y Zona Urbana Moche. Se realizaron análisis discriminantes a una muestra de 44 primeras falanges, tomando en cuenta las cinco variables que son medidas en este hueso. Los resultados obtenidos mostraron la identificación de dos especies de camélidos domésticos en ambos sitios, Lama glama “llama” y Lama pacos “alpaca”. La interpretación de los resultados es discutida mediante la explicación de un modelo de especiación geográfica o vicariante de los camélidos que habitaron la costa peruana en la época Mochica. Este modelo toma en cuenta que actualmente en esta ecología no hay crianza de camélidos, así como factores genéticos y evolutivos de una nueva forma doméstica, incluyendo el efecto de la Regla de Bergmann y los nuevos pastos en la morfología de esta nueva forma de camélido doméstico que vivió en la época Mochica.Palabras claves: osteometría, genética, camélidos, especiación, Mochica  AbstractThe objective of this study was to evaluate the results of osteometric analyses applied to camelid bones samples from Moche sites (I and VII AD located on the north coast of Peru. The bones analyzed in this study correspond to first phalanges from two Moche archaeological sites (Cerro Chepén and Moche Urban Zone. Discriminant analysis was conducted on a sample of 44 first phalanges, taking into account five measurments in each bone. The results showed the identification in both sites of two species of domestic camelids, Lama glama “llama” and Lama pacos “alpaca.” The interpretation of the results is discussed by explaining a model of geographic or vicariant speciation of camelids that inhabited the coast

  17. Structural Mimicry of Receptor Interaction by Antagonistic Interleukin-6 (IL-6) Antibodies.

    Science.gov (United States)

    Blanchetot, Christophe; De Jonge, Natalie; Desmyter, Aline; Ongenae, Nico; Hofman, Erik; Klarenbeek, Alex; Sadi, Ava; Hultberg, Anna; Kretz-Rommel, Anke; Spinelli, Silvia; Loris, Remy; Cambillau, Christian; de Haard, Hans

    2016-06-24

    Interleukin 6 plays a key role in mediating inflammatory reactions in autoimmune diseases and cancer, where it is also involved in metastasis and tissue invasion. Neutralizing antibodies against IL-6 and its receptor have been approved for therapeutic intervention or are in advanced stages of clinical development. Here we describe the crystal structures of the complexes of IL-6 with two Fabs derived from conventional camelid antibodies that antagonize the interaction between the cytokine and its receptor. The x-ray structures of these complexes provide insights into the mechanism of neutralization by the two antibodies and explain the very high potency of one of the antibodies. It effectively competes for binding to the cytokine with IL-6 receptor (IL-6R) by using side chains of two CDR residues filling the site I cavities of IL-6, thus mimicking the interactions of Phe(229) and Phe(279) of IL-6R. In the first antibody, a HCDR3 tryptophan binds similarly to hot spot residue Phe(279) Mutation of this HCDR3 Trp residue into any other residue except Tyr or Phe significantly weakens binding of the antibody to IL-6, as was also observed for IL-6R mutants of Phe(279) In the second antibody, the side chain of HCDR3 valine ties into site I like IL-6R Phe(279), whereas a LCDR1 tyrosine side chain occupies a second cavity within site I and mimics the interactions of IL-6R Phe(229). PMID:27129274

  18. Ricin Detection Using Phage Displayed Single Domain Antibodies

    Directory of Open Access Journals (Sweden)

    Ellen R. Goldman

    2009-01-01

    Full Text Available Phage-displayed single domain antibodies (sdAb were compared to monomeric solubly expressed sdAb and llama polyclonal antibodies for the detection of ricin. SdAb are comprised of the variable domain derived from camelid heavy chain only antibodies (HcAb. Although HcAb lack variable light chains, they as well as their derivative sdAb are able to bind antigens with high affinity. The small size of sdAb (~16 kDa, while advantageous in many respects, limits the number of labels that can be incorporated. The ability to incorporate multiple labels is a beneficial attribute for reporter elements. Opportunely, sdAb are often selected using phage display methodology. Using sdAb displayed on bacteriophage M13 as the reporter element gives the potential for incorporating a very high number of labels. We have demonstrated the use of both sdAb and phage- displayed sdAb for the detection of ricin using both enzyme linked immunosorbent assays (ELISAs and Luminex fluid array assays. The phage-displayed sdAb led to five to ten fold better detection of ricin in both the ELISA and Luminex assays, resulting in limits of detection of 1 ng/mL and 64 pg/mL respectively. The phage-displayed sdAb were also dramatically more effective for the visualization of binding to target in nitrocellulose dot blot assays, a method frequently used for epitope mapping.

  19. Culicoides vector species on three South American camelid farms seropositive for bluetongue virus serotype 8 in Germany 2008/2009.

    Science.gov (United States)

    Schulz, Claudia; Ziller, Mario; Kampen, Helge; Gauly, Matthias; Beer, Martin; Grevelding, Christoph G; Hoffmann, Bernd; Bauer, Christian; Werner, Doreen

    2015-12-15

    Palearctic species of Culicoides (Diptera, Ceratopogonidae), in particular of the Obsoletus and Pulicaris complexes, were identified as putative vectors of bluetongue virus serotype 8 (BTV-8) on ruminant farms during the epizootic in Germany from 2006 to 2009. BTV may cause severe morbidity and mortality in ruminants and sporadically in South American camelids (SAC). However, the fauna of Culicoides spp. on SAC farms has not been investigated. Therefore, the ceratopogonid fauna was monitored on three farms with BTV-seropositive SAC in Germany. Black-light traps were set up on pastures and in stables from summer 2008 to autumn 2009. Additionally, ceratopogonids were caught in emergence traps mounted on llama dung and dung-free pasture from spring to autumn 2009. After morphological identification, selected Culicoides samples were analysed for BTV-RNA by real-time RT-PCR. The effects of the variables 'location', 'temperature' and 'humidity' on the number of Culicoides caught in black-light traps were modelled using multivariable Poisson regression. In total, 26 species of Culicoides and six other genera of biting midges were identified. The most abundant Culicoides spp. collected both outdoors and indoors with black-light traps belonged to the Obsoletus (77.4%) and Pulicaris (16.0%) complexes. The number of Culicoides peaked in summer, while no biting midges were caught during the winter months. Daily collections of Culicoides were mainly influenced by the location and depended on the interaction of temperature and humidity. In the emergence traps, species of the Obsoletus complex predominated the collections. In summary, the absence of BTV-RNA in any of the analysed Culicoides midges and in the BTV-seropositive SAC on the three farms together with the differences in the pathogenesis of BTV-8 in SAC compared to ruminants suggests a negligible role of SAC in the spread of the virus. Although SAC farms may provide similar suitable habitats for putative Culicoides

  20. Presence, distribution and steroidogenic effect of the peptides orexin A and receptor 1 for orexins in the testis of the South American camelid alpaca (Vicugna pacos).

    Science.gov (United States)

    Liguori, Giovanna; Assisi, Loredana; Squillacioti, Caterina; Paino, Salvatore; Mirabella, Nicola; Vittoria, Alfredo

    2012-10-01

    The orexins A (oxA) and B are peptides discovered in the rat hypothalamus and successively found in some peripheral organs of the mammalian body. They binds two protein G-coupled receptors defined receptor 1 (ox1r) and 2 for orexins, the first of which is highly specific for oxA while the second binds both the peptides with equal affinity. This work aimed to detect the presence of oxA and ox1r in the testis of the South American camelid alpaca (Vicugna pacos) and investigate the role played by them on Leydig cell steroidogenesis. The species alpaca acquired, in the last years, increasing zootechnical interest for the quality of the wool produced and its breeding spread from the country of origin to USA, Australia and Europe. Immunohistochemistry allowed us to detect oxA in Leydig and Sertoli cells, spermatogonia, resting spermatocytes, round and oval spermatids. Ox1r-immunoreactivity was found in Leydig cells and round, oval and elongated spermatids. The expression of the two peptides in tissue extracts was established by using Western blotting technique. Such results demonstrated that in the alpaca testis exists in a cellular complex able to produce and/or internalize oxA. Finally, the effect of oxA on steroidogenesis was investigated by means of in vitro cultured thin testis slices which were added with oxA or/and Müllerian Inhibiting Substance (MIS), a steroidolitic agent basally produced by the Sertoli cell. OxA evoked increase of testosterone production while MIS a decrease. The consecutive addition of oxA and MIS, or vice versa, highlighted an antagonistic interplay between the two substances which has been thought to be the main molecular event at the basis of the oxA-stimulated steroidogenesis mechanism.

  1. Monoclonal antibodies

    International Nuclear Information System (INIS)

    Monoclonal antibodies (MAbs) are antibodies having single specificity for a given antigen site (epitope). The development of hybridoma technology and the relative ease by which MAbs can be prepared has revolutionized many aspects of serological applications in diagnosis and differentiation of disease producing agents. The property of monospecificity offers advantages in diagnostic applications over polyclonal sera in that tests can be defined exactly with regard to the antigen detected and the affinity of reaction between the given antigenic site and the monoclonal reagent. In addition, MAbs offer better possibilities for test standardization, because the same reagent can be used in different laboratories. Such an MAb can be supplied by a central laboratory or 'grown' from hybridoma cells, ensuring that the resultant product is identical from laboratory to laboratory and that the part of the test involving the MAb reaction is the same. The methodologies for inoculation regimes, mice, cloning methods, selection of fusion partners, etc., have been validated extensively in developed country laboratories. The decision to establish a MAb production facility must be examined on a strict cost-benefit basis, since it is still expensive to produce a product. There are many MAbs available that should be sought to allow exploitation in developing tests. If a production facility is envisaged, it should produce reagents for national needs, i.e. there should be a clear problem oriented approach whereby exact needs are defined. In the field of veterinary applications, MAbs are the central reagent in many immunoassays based on the enzyme linked immunosorbent assay (ELISA). The development of specific tests for diagnosing diseases is dominated by MAbs and has been fuelled by a strong research base, mainly in developed countries allied to developing countries through the study of related diseases. Thus, there are very many assays dependent on MAbs, some of which form the basis of

  2. Monoclonal antibodies.

    Science.gov (United States)

    2009-01-01

    The ability to produce and exploit monoclonal antibodies (mAbs) has revolutionized many areas of biological sciences. The unique property of an mAb is that it is a single species of immunoglobulin (IG) molecule. This means that the specificity of the interaction of the paratopes on the IG, with the epitopes on an antigenic target, is the same on every molecule. This property can be used to great benefit in immunoassays to provide tests of defined specificity and sensitivity, which improve the possibilities of standardization. The performance of assays can often be determined relating the actual weight of antibody (hence the number of molecules) to the activity. Often the production of an mAb against a specific epitope is the only way that biological entities can be differentiated. This chapter outlines the areas involving the development of assays based on mAbs. The problems involved address include the physical aspects of mAbs and how they may affect assay design and also the implications of results based on monospecific reagents. Often these are not fully understood, leading to assays that are less than satisfactory, which does not justify the relatively high cost of preparing and screening of mAbs. There are many textbooks and reviews dealing with the preparation of mAbs, the principles involved, and various purification and manipulative methods for the preparation of fragments and conjugation. There has been little general information attempting to summarize the best approaches to assay design using mAbs. Much time can be wasted through bad planning, and this is particularly relevant to mAbs. A proper understanding of some basic principles is essential. It is beyond the scope of this chapter to discuss all aspects, but major areas are highlighted. PMID:19219589

  3. Improving the biophysical properties of anti-ricin single-domain antibodies

    Directory of Open Access Journals (Sweden)

    Kendrick B. Turner

    2015-06-01

    Full Text Available Single-domain antibodies (sdAbs derived from heavy-chain only antibodies produced in camelids are attractive immunoreagents due to their small size, high affinity, and ability to refold and retain binding activity after denaturation. It has been observed that some sdAbs, however, exhibit undesirable properties including reduced solubility when subjected to heating or upon long-term storage at production-relevant concentrations, which can limit their usefulness. Using a multi-step, rational design approach that included consensus-sequence driven sequence repairs, the alteration of net protein charge, and the introduction of non-native disulfide bonds, augmented solubility and increased melting temperatures were achieved. The improved sdAbs tolerated storage in solution at high concentration (10 mg/mL and were able to withstand multiple cycles of heating to high temperature (70 °C. This work demonstrates a pathway for improving the biophysical characteristics of sdAbs which is essential for expanding their utility for both diagnostic as well as therapeutic applications.

  4. Intramolecular trimerization, a novel strategy for making multispecific antibodies with controlled orientation of the antigen binding domains.

    Science.gov (United States)

    Alvarez-Cienfuegos, Ana; Nuñez-Prado, Natalia; Compte, Marta; Cuesta, Angel M; Blanco-Toribio, Ana; Harwood, Seandean Lykke; Villate, Maider; Merino, Nekane; Bonet, Jaume; Navarro, Rocio; Muñoz-Briones, Clara; Sørensen, Karen Marie Juul; Mølgaard, Kasper; Oliva, Baldo; Sanz, Laura; Blanco, Francisco J; Alvarez-Vallina, Luis

    2016-01-01

    Here, we describe a new strategy that allows the rapid and efficient engineering of mono and multispecific trivalent antibodies. By fusing single-domain antibodies from camelid heavy-chain-only immunoglobulins (VHHs) to the N-terminus of a human collagen XVIII trimerization domain (TIE(XVIII)) we produced monospecific trimerbodies that were efficiently secreted as soluble functional proteins by mammalian cells. The purified VHH-TIE(XVIII) trimerbodies were trimeric in solution and exhibited excellent antigen binding capacity. Furthermore, by connecting with two additional glycine-serine-based linkers three VHH-TIE(XVIII) modules on a single polypeptide chain, we present an approach for the rational design of multispecific tandem trimerbodies with defined stoichiometry and controlled orientation. Using this technology we report here the construction and characterization of a tandem VHH-based trimerbody capable of simultaneously binding to three different antigens: carcinoembryonic antigen (CEA), epidermal growth factor receptor (EGFR) and green fluorescence protein (GFP). Multispecific tandem VHH-based trimerbodies were well expressed in mammalian cells, had good biophysical properties and were capable of simultaneously binding their targeted antigens. Importantly, these antibodies were very effective in inhibiting the proliferation of human epidermoid carcinoma A431 cells. Multispecific VHH-based trimerbodies are therefore ideal candidates for future applications in various therapeutic areas. PMID:27345490

  5. [Antinuclear antibodies].

    Science.gov (United States)

    Cabiedes, Javier; Núñez-Álvarez, Carlos A

    2010-01-01

    Anti-nuclear antibodies (ANA) are immunoglobulin directed against autologous cell nuclear and cytoplasmic components. Besides the autoimmune ANA there are other ANA that can be detected in circulation, like natural and infectious ANA. Because of its high sensibility, detection of the ANA must be done by indirect immunofluorescence (IIF) as screening test and all of those positive samples are convenient to confirm its specificity by ELISA, western blot or other techniques. Positive ANA detected by IIF must be evaluated taking in to account the pattern and titer. The following recommended step is the specificity characterization (reactivity against extractable nuclear antigens [ENA], dsDNA, etc.) which is useful for the diagnosis and follow up of patients with autoimmune diseases, and by such reasoning, its detection must be performed in an orderly and reasonable way using guides or strategies focused to the good use and interpretation of the autoantibodies. The objective of this review is to present a compilation of the literature and our experience in the detection and study of the ANA.

  6. Generation and characterization of CD1d-specific single-domain antibodies with distinct functional features.

    Science.gov (United States)

    Lameris, Roeland; de Bruin, Renée C G; van Bergen En Henegouwen, Paul M P; Verheul, Henk M; Zweegman, Sonja; de Gruijl, Tanja D; van der Vliet, Hans J

    2016-09-01

    Ligation of the CD1d antigen-presenting molecule by monoclonal antibodies (mAbs) can trigger important biological functions. For therapeutic purposes camelid-derived variable domain of heavy-chain-only antibodies (VHH) have multiple advantages over mAbs because they are small, stable and have low immunogenicity. Here, we generated 21 human CD1d-specific VHH by immunizing Lama glama and subsequent phage display. Two clones induced maturation of dendritic cells, one clone induced early apoptosis in CD1d-expressing B lymphoblasts and multiple myeloma cells, and another clone blocked recognition of glycolipid-loaded CD1d by CD1d-restricted invariant natural killer T (iNKT) cells. In contrast to reported CD1d-specific mAbs, these CD1d-specific VHH have the unique characteristic that they induce specific and well-defined biological effects. This feature, combined with the above-indicated general advantages of VHH, make the CD1d-specific VHH generated here unique and useful tools to exploit both CD1d ligation as well as disruption of CD1d-iNKT interactions in the treatment of cancer or inflammatory disorders. PMID:27312006

  7. Monoclonal antibodies and cancer

    International Nuclear Information System (INIS)

    The usefulness of radiolabeled monoclonal antibodies for imaging and treatment of human (ovarian) cancer was investigated. A review of tumor imaging with monoclonal antibodies is presented. Special attention is given to factors that influence the localization of the antibodies in tumors, isotope choice and methods of radiolabeling of the monoclonal antibodies. Two monoclonal antibodies, OC125 and OV-TL3, with high specificity for human epithelial ovarian cancer are characterized. A simple radio-iodination technique was developed for clinical application of the monoclonal antibodies. The behavior of monoclonal antibodies in human tumor xenograft systems and in man are described. Imaging of tumors is complicated because of high background levels of radioactivity in other sites than the tumor, especially in the bloodpool. A technique was developed to improve imaging of human tumor xenographs in nude mice, using subtraction of a specific and a non-specific antibody, radiolabeled with 111In, 67Ga and 131I. To investigate the capability of the two monoclonal antibodies, to specifically localize in human ovarian carcinomas, distribution studies in mice bearing human ovarian carcinoma xenografts were performed. One of the antibodies, OC125, was used for distribution studies in ovarian cancer patients. OC125 was used because of availability and approval to use this antibody in patients. The same antibody was used to investigate the usefulness of radioimmunoimaging in ovarian cancer patients. The interaction of injected radiolabeled antibody OC125 with circulating antigen and an assay to measure the antibody response in ovarian cancer patients after injection of the antibody is described. 265 refs.; 30 figs.; 19 tabs

  8. [VGKC-complex antibodies].

    Science.gov (United States)

    Watanabe, Osamu

    2013-04-01

    Various antibodies are associated with voltage-gated potassium channels (VGKCs). Representative antibodies to VGKCs were first identified by radioimmunoassays using radioisotope-labeled alpha-dendrotoxin-VGKCs solubilized from rabbit brain. These antibodies were detected only in a proportion of patients with acquired neuromyotonia (Isaacs' syndrome). VGKC antibodies were also detected in patients with Morvan's syndrome and in those with a form of autoimmune limbic encephalitis. Recent studies indicated that the "VGKC" antibodies are mainly directed toward associated proteins (for example LGI-1 and CASPR-2) that complex with the VGKCs themselves. The "VGKC" antibodies are now commonly known as VGKC-complex antibodies. In general, LGI-1 antibodies are most commonly detected in patients with limbic encephalitis with syndrome of inappropriate secretion of antidiuretic hormone. CASPR-2 antibodies are present in the majority of patients with Morvan's syndrome. These patients develop combinations of CNS symptoms, autonomic dysfunction, and peripheral nerve hyperexcitability. Furthermore, VGKC-complex antibodies are tightly associated with chronic idiopathic pain. Hyperexcitability of nociceptive pathways has also been implicated. These antibodies may be detected in sera of some patients with neurodegenerative diseases (for example, amyotrophic lateral sclerosis and Creutzfeldt-Jakob disease).

  9. Heavy chain only antibodies

    DEFF Research Database (Denmark)

    Moghimi, Seyed Moein; Rahbarizadeh, Fatemeh; Ahmadvand, Davoud;

    2013-01-01

    Unlike conventional antibodies, heavy chain only antibodies derived from camel contain a single variable domain (VHH) and two constant domains (CH2 and CH3). Cloned and isolated VHHs possess unique properties that enable them to excel conventional therapeutic antibodies and their smaller antigen-...... for combating HER2+ breast cancer. © 2013 by Tabriz University of Medical Sciences.......Unlike conventional antibodies, heavy chain only antibodies derived from camel contain a single variable domain (VHH) and two constant domains (CH2 and CH3). Cloned and isolated VHHs possess unique properties that enable them to excel conventional therapeutic antibodies and their smaller antigen......-binding fragments in cancer targeting and therapy. VHHs express low immunogenicity, are highly robust and easy to manufacture and have the ability to recognize hidden or uncommon epitopes. We highlight the utility of VHH in design of new molecular, multifunctional particulate and immune cell-based systems...

  10. Engineering antibody therapeutics.

    Science.gov (United States)

    Chiu, Mark L; Gilliland, Gary L

    2016-06-01

    The successful introduction of antibody-based protein therapeutics into the arsenal of treatments for patients has within a few decades fostered intense innovation in the production and engineering of antibodies. Reviewed here are the methods currently used to produce antibodies along with how our knowledge of the structural and functional characterization of immunoglobulins has resulted in the engineering of antibodies to produce protein therapeutics with unique properties, both biological and biophysical, that are leading to novel therapeutic approaches. Antibody engineering includes the introduction of the antibody combining site (variable regions) into a host of architectures including bi and multi-specific formats that further impact the therapeutic properties leading to further advantages and successes in patient treatment. PMID:27525816

  11. Intracellular expression of a single domain antibody reduces cytotoxicity of 15-acetyldeoxynivalenol in yeast.

    Science.gov (United States)

    Doyle, Patrick J; Saeed, Hanaa; Hermans, Anne; Gleddie, Steve C; Hussack, Greg; Arbabi-Ghahroudi, Mehdi; Seguin, Charles; Savard, Marc E; Mackenzie, C Roger; Hall, J Christopher

    2009-12-11

    15-Acetyldeoxynivalenol (15-AcDON) is a low molecular weight sesquiterpenoid trichothecene mycotoxin associated with Fusarium ear rot of maize and Fusarium head blight of small grain cereals. The accumulation of mycotoxins such as deoxynivalenol (DON) and 15-AcDON within harvested grain is subject to stringent regulation as both toxins pose dietary health risks to humans and animals. These toxins inhibit peptidyltransferase activity, which in turn limits eukaryotic protein synthesis. To assess the ability of intracellular antibodies (intrabodies) to modulate mycotoxin-specific cytotoxocity, a gene encoding a camelid single domain antibody fragment (V(H)H) with specificity and affinity for 15-AcDON was expressed in the methylotropic yeast Pichia pastoris. Cytotoxicity and V(H)H immunomodulation were assessed by continuous measurement of cellular growth. At equivalent doses, 15-AcDON was significantly more toxic to wild-type P. pastoris than was DON. In turn, DON was orders of magnitude more toxic than 3-acetyldeoxynivalenol. Intracellular expression of a mycotoxin-specific V(H)H within P. pastoris conveyed significant (p = 0.01) resistance to 15-AcDON cytotoxicity at doses ranging from 20 to 100 mug.ml(-1). We also documented a biochemical transformation of DON to 15-AcDON to account for the attenuation of DON cytotoxicity at 100 and 200 mug.ml(-1). The proof of concept established within this eukaryotic system suggests that in planta V(H)H expression may lead to enhanced tolerance to mycotoxins and thereby limit Fusarium infection of commercial agricultural crops. PMID:19783651

  12. RBC Antibody Screen

    Science.gov (United States)

    ... the baby is Rh-positive and the mother's antibody status is negative for anti-D, the mother is given additional RhIG. This test also may be used to help diagnose autoimmune-related hemolytic anemia ... when a person produces antibodies against his or her own RBC antigens. This ...

  13. Selection of Recombinant Human Antibodies.

    Science.gov (United States)

    Tomszak, Florian; Weber, Susanne; Zantow, Jonas; Schirrmann, Thomas; Hust, Michael; Frenzel, André

    2016-01-01

    Since the development of therapeutic antibodies the demand of recombinant human antibodies is steadily increasing. Traditionally, therapeutic antibodies were generated by immunization of rat or mice, the generation of hybridoma clones, cloning of the antibody genes and subsequent humanization and engineering of the lead candidates. In the last few years, techniques were developed that use transgenic animals with a human antibody gene repertoire. Here, modern recombinant DNA technologies can be combined with well established immunization and hybridoma technologies to generate already affinity maturated human antibodies. An alternative are in vitro technologies which enabled the generation of fully human antibodies from antibody gene libraries that even exceed the human antibody repertoire. Specific antibodies can be isolated from these libraries in a very short time and therefore reduce the development time of an antibody drug at a very early stage.In this review, we describe different technologies that are currently used for the in vitro and in vivo generation of human antibodies. PMID:27236551

  14. Anti-insulin antibody test

    Science.gov (United States)

    Insulin antibodies - serum; Insulin Ab test; Insulin resistance - insulin antibodies; Diabetes - insulin antibodies ... You appear to have an allergic response to insulin Insulin no longer seems to control your diabetes

  15. HIV Antibody Test

    Science.gov (United States)

    ... be limited. Home Visit Global Sites Search Help? HIV Antibody and HIV Antigen (p24) Share this page: Was this page helpful? Also known as: HIV Screening Tests; AIDS Test; AIDS Screen; HIV Serology; ...

  16. Antinuclear antibody panel

    Science.gov (United States)

    ... blood may be due to: Chronic liver disease Collagen vascular disease Drug-induced lupus erythematosus Myositis (inflammatory muscle disease) ... Saunders; 2011:chap 51. Read More Antibody Arthritis Collagen vascular disease Drug-induced lupus erythematosus Liver disease Scleroderma Systemic ...

  17. Anti-cartilage antibody.

    Science.gov (United States)

    Greenbury, C L; Skingle, J

    1979-08-01

    Antibody to cartilage has been demonstrated by indirect immunofluorescence on rat trachea in the serum of about 3% of 1126 patients with rheumatoid arthritis. Titres ranged from 1:20 to 1:640. The antibody was not found in 284 patients with primary or secondary osteoarthritis or in 1825 blood donors, nor, with the exception of two weak reactors, in 1314 paraplegic patients. In most cases the antibody appears to be specific for native type II collagen. Using this as an antigen in a haemagglutination test 94% of anti-cartilage sera were positive, whereas among 100 rheumatoid control sera there were only three weak positives. More than 80% of patients with antibody had some erosion of articular cartilage, but there was no correlation with age, sex, duration of disease, nor any recognisable clinical event or change.

  18. Antibody tumor penetration

    Science.gov (United States)

    Thurber, Greg M.; Schmidt, Michael M.; Wittrup, K. Dane

    2009-01-01

    Antibodies have proven to be effective agents in cancer imaging and therapy. One of the major challenges still facing the field is the heterogeneous distribution of these agents in tumors when administered systemically. Large regions of untargeted cells can therefore escape therapy and potentially select for more resistant cells. We present here a summary of theoretical and experimental approaches to analyze and improve antibody penetration in tumor tissue. PMID:18541331

  19. Expression of Recombinant Antibodies

    OpenAIRE

    Frenzel, André; Hust, Michael; Schirrmann, Thomas

    2013-01-01

    Recombinant antibodies are highly specific detection probes in research, diagnostics, and have emerged over the last two decades as the fastest growing class of therapeutic proteins. Antibody generation has been dramatically accelerated by in vitro selection systems, particularly phage display. An increasing variety of recombinant production systems have been developed, ranging from Gram-negative and positive bacteria, yeasts and filamentous fungi, insect cell lines, mammalian cells to transg...

  20. Antibody informatics for drug discovery

    DEFF Research Database (Denmark)

    Shirai, Hiroki; Prades, Catherine; Vita, Randi;

    2014-01-01

    for antibody rational design using computational approaches to affinity and stability improvement, as well as ab-initio and homology-based antibody modeling; (ii) resources for antibody sequences, structures, and immune epitopes and open drug discovery resources for development of antibody drugs; and (iii...

  1. Engineering antibodies for cancer therapy.

    Science.gov (United States)

    Boder, Eric T; Jiang, Wei

    2011-01-01

    The advent of modern antibody engineering has led to numerous successes in the application of these proteins for cancer therapy in the 13 years since the first Food and Drug Administration approval, which has stimulated active interest in developing more and better drugs based on these molecules. A wide range of tools for discovering and engineering antibodies has been brought to bear on this challenge in the past two decades. Here, we summarize mechanisms of monoclonal antibody therapeutic activity, challenges to effective antibody-based treatment, existing technologies for antibody engineering, and current concepts for engineering new antibody formats and antibody alternatives as next generation biopharmaceuticals for cancer treatment.

  2. Construction of a biotinylated cameloid-like antibody for lable-free detection of apolipoprotein B-100.

    Science.gov (United States)

    Li, Henan; Yan, Junrong; Ou, Weijun; Liu, Hong; Liu, Songqin; Wan, Yakun

    2015-02-15

    Nanobodies (Nbs), also known as the variable domain of the heavy-chain-only antibody (VHH), are single-domain antigen-binding fragments derived from heavy-chain antibodies that occur naturally in sera of camelids. Due to their unique properties of small size (15 kD), intrinsic stability, high affinity and specificity, Nbs are suitable for detecting clinical relevant antigens. Apolipoprotein B-100 (ApoB-100) is a highly predictive marker for coronary artery disease (CAD), which is frequently detected in clinical diagnosis. Herein, we successfully obtained anti-ApoB-100 Nbs for the first time and further fabricated a label-free and sensitive immunosensor for ApoB-100 based on isolated anti-ApoB-100 nanobody (Nb) using the electrochemical impedance spectroscopy (EIS) technique. We have generated an immunized phage display library against ApoB-100 and isolated four anti-ApoB-100 Nbs with high affinity and stability. The Nb with the highest affinity was biotinylated based on in vivo BirA system. Further, we developed a label-free electrochemical impedance immunosensor for ApoB-100 using this anti-ApoB-100 Nb. The attachment of ApoB-100 onto the anti-ApoB-100 Nb-immobilized sensing layer led to the increased electron-transfer resistance, which was proportional to ApoB-100 concentration in the range from 0.05 to 5 ng mL(-1) with a detection limit of 0.03 ng mL(-1). This proposed immunosensor revealed high specificity to detect ApoB-100, acceptable intra-assay precision and good stability, functioning as a feasible technique for CAD diagnosis. PMID:25203942

  3. Antibody affinity maturation

    DEFF Research Database (Denmark)

    Skjødt, Mette Louise

    linker for yeast surface display of scFv and scFab fragments, we compared a series of different Gly-Ser-based linkers in display and antigen binding proficiency. We show that these formats of the model antibody can accommodate linkers of different lengths and that introduction of alanine or glutamate......-2. Based on the presented data we suggest that affinity maturation of the model antibody proceeds through multiple incremental steps of subtle improvements. We moreover conclude that the best affinity improved candidates are likely to be obtained through optimization of both the antigen...... fragments by in vivo homologous recombination large combinatorial antibody libraries can easily be generated. We have optimized ordered assembly of three CDR fragments into a gapped vector and observed increased transformation efficiency in a yeast strain carrying a deletion of the SGS1 helicase...

  4. Antithyroglobulin Antibodies and Antimicrosomal Antibodies in Various Thyroid Diseases

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Gwon Jun; Hong, Key Sak; Choi, Kang Won; Lee, Kyu; Koh, Chang Soon; Lee, Mun Ho; Park, Sung Hoe; Chi, Je Geun; Lee, Sang Kook [Seoul National University College of Medicine, Seoul (Korea, Republic of)

    1979-03-15

    The authors investigated the incidence of antithyroglobulin antibodies and antibodies and antimicrosomal antibodies measured by tanned red cell hemagglutination method in subjects suffering from various thyroid disorders. 1) In 15 normal patients, neither suffering from any thyroid diseases nor from any other autoimmune disorders, the antithyroglobulin antibodies were all negative, but the antimicrosomal antibody was positive only in one patient (6.7%). 2) The antithyroglobulin antibodies were positive in 31.5% (34 patients) of 108 patients with various thyroid diseases, and the antimicrosomal antibodies were positive in 37.0% (40 patients). 3) of the 25 patients with Graves' diseases, 7 patients (28.0%) showed positive for the antithyroglobulin antibodies, and 9 (36.0%) for the antimicrosomal antibodies. There was no definite differences in clinical and thyroid functions between the groups with positive and negative results. 4) Both antibodies were positive in 16 (88.9%) and 17 (94.4%) patients respectively among 18 patients with Hashimoto's thyroiditis, all of them were diagnosed histologically. 5) Three out of 33 patients with thyroid adenoma showed positive antibodies, and 3 of 16 patients with thyroid carcinoma revealed positive antibodies. 6) TRCH antibodies demonstrated negative results in 2 patients with subacute thyroiditis, but positive in one patient with idiopathic primary myxedema. 7) The number of patients with high titers(>l:802) was 16 for antithyroglobulin antibody, and 62.5% (10 patients) of which was Hashimoto's thyroiditis. Thirteen (65.0) of 20 patients with high titers (>l:802) for antimicrosomal antibody was Hashimoto's thyroiditis. TRCH test is a simple, sensitive method, and has high reliability and reproducibility. The incidences and titers of antithyroglobulin antibody and antimicrosomal antibody are especially high in Hashimoto's thyroiditis.

  5. Monoclonal antibodies in myeloma

    DEFF Research Database (Denmark)

    Sondergeld, P.; van de Donk, N. W. C. J.; Richardson, P. G.;

    2015-01-01

    The development of monoclonal antibodies (mAbs) for the treatment of disease goes back to the vision of Paul Ehrlich in the late 19th century; however, the first successful treatment with a mAb was not until 1982, in a lymphoma patient. In multiple myeloma, mAbs are a very recent and exciting add...

  6. Prediction of Antibody Epitopes

    DEFF Research Database (Denmark)

    Nielsen, Morten; Marcatili, Paolo

    2015-01-01

    self-proteins. Given the sequence or the structure of a protein of interest, several methods exploit such features to predict the residues that are more likely to be recognized by an immunoglobulin.Here, we present two methods (BepiPred and DiscoTope) to predict linear and discontinuous antibody...

  7. Lupus anticoagulants and antiphospholipid antibodies

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/000547.htm Lupus anticoagulants and antiphospholipid antibodies To use the sharing features on this page, please enable JavaScript. Lupus anticoagulants are antibodies against substances in the lining ...

  8. Recombinant antibodies and tumor targeting

    OpenAIRE

    Sheikholvaezin, Ali

    2006-01-01

    Different antibody derived constructs are rapidly advancing as putative tools for treatment of malignant diseases. Antibody engineering has added significant new technologies to modify size, affinities, solubility, stability and biodistribution properties for immunoconjugates. In the present thesis, the aim was to increase our knowledge on how new recombinant antibodies could be tailored to optimize localization to experimental tumors in mice. One hybridoma, producing the monoclonal antibody ...

  9. Antibody mimetics: promising complementary agents to animal-sourced antibodies.

    Science.gov (United States)

    Baloch, Abdul Rasheed; Baloch, Abdul Wahid; Sutton, Brian J; Zhang, Xiaoying

    2016-01-01

    Despite their wide use as therapeutic, diagnostic and detection agents, the limitations of polyclonal and monoclonal antibodies have inspired scientists to design the next generation biomedical agents, so-called antibody mimetics that offer many advantages over conventional antibodies. Antibody mimetics can be constructed by protein-directed evolution or fusion of complementarity-determining regions through intervening framework regions. Substantial progress in exploiting human, butterfly (Pieris brassicae) and bacterial systems to design and select mimetics using display technologies has been made in the past 10 years, and one of these mimetics [Kalbitor® (Dyax)] has made its way to market. Many challenges lie ahead to develop mimetics for various biomedical applications, especially those for which conventional antibodies are ineffective, and this review describes the current characteristics, construction and applications of antibody mimetics compared to animal-sourced antibodies. The possible limitations of mimetics and future perspectives are also discussed. PMID:25264572

  10. A monoclonal antibody against leptin.

    Science.gov (United States)

    Mahmoudian, Jafar; Jeddi-Tehrani, Mahmood; Bayat, Ali Ahmad; Mahmoudi, Ahmad Reza; Vojgani, Yasaman; Tavangar, Banafsheh; Hadavi, Reza; Zarei, Saeed

    2012-10-01

    Leptin is an important protein that regulates energy storage and homeostasis in humans and animals. Leptin deficiency results in various abnormalities such as diabetes, obesity, and infertility. Producing a high affinity monoclonal antibody against human leptin provides an important tool to monitor and trace leptin function in different biological fluids. In this study, recombinant human leptin was conjugated to KLH and injected into mice. After immunization, mouse myeloma SP2/0 cells were fused with murine splenocytes followed by selection of antibody-producing hybridoma cells. After screening of different hybridoma colonies by ELISA, a high affinity antibody was selected and purified by affinity chromatography. The affinity constant of the antibody was measured by ELISA. Western blot, immunocytochemistry, and flow cytometry experiments were used to characterize the antibody. The anti-leptin antibody had a high affinity (around 1.13 × 10(-9) M) for its antigen. The saturation of the antibody with leptin (20 moles leptin per 1 mole antibody) in Western blot analysis proved that the antibody had specific binding to its antigen. Immunocytochemistry and flow cytometry on JEG-3 (human placental choriocarcinoma cell) cells revealed that the anti-leptin antibody recognized intracellular leptin. In conclusion, we report here the production and characterization of a murine anti-leptin antibody with high affinity for human leptin. PMID:23098305

  11. Engineering antibodies by yeast display.

    Science.gov (United States)

    Boder, Eric T; Raeeszadeh-Sarmazdeh, Maryam; Price, J Vincent

    2012-10-15

    Since its first application to antibody engineering 15 years ago, yeast display technology has been developed into a highly potent tool for both affinity maturing lead molecules and isolating novel antibodies and antibody-like species. Robust approaches to the creation of diversity, construction of yeast libraries, and library screening or selection have been elaborated, improving the quality of engineered molecules and certainty of success in an antibody engineering campaign and positioning yeast display as one of the premier antibody engineering technologies currently in use. Here, we summarize the history of antibody engineering by yeast surface display, approaches used in its application, and a number of examples highlighting the utility of this method for antibody engineering.

  12. Antiphospholipid Antibody and Antiphospholipid Syndrome

    Institute of Scientific and Technical Information of China (English)

    吴竞生

    2008-01-01

    @@ Antiphospholipid antibodies (APA) APA is a big category for all kinds of negative charge phospholipid or lecithin - a protein complex autoantibodies or the same antibody, through its recognition of antigen (target protein) different, and phospholipids or lecithin - protein complex combination of various rely on the interference Phospholipid clotting and anti-coagulation factor, and promote endothelial cells, platelets, complement activation and play a role. APA including lupus anticoagulant(LA) and anticardiolipin antibody (ACA), In addition, there are anti-β2 glycoprotein-I (β2-GPI) antibody, anti-prothrombin (a- PT) antibody, anti-lysophosphatidic acid antibody and anti-phosphatidylserine antibody, and so on. APA as the main target of phospholipid-binding protein, including β2-GPI, prothrombin, annexin, protein C (PC) and protein S (PS), plasminogen, and so on.

  13. Site-directed immobilization of a genetically engineered anti-methotrexate antibody via an enzymatically introduced biotin label significantly increases the binding capacity of immunoaffinity columns.

    Science.gov (United States)

    Davenport, Kaitlynn R; Smith, Christopher A; Hofstetter, Heike; Horn, James R; Hofstetter, Oliver

    2016-05-15

    In this study, the effect of random vs. site-directed immobilization techniques on the performance of antibody-based HPLC columns was investigated using a single-domain camelid antibody (VHH) directed against methotrexate (MTX) as a model system. First, the high flow-through support material POROS-OH was activated with disuccinimidyl carbonate (DSC), and the VHH was bound in a random manner via amines located on the protein's surface. The resulting column was characterized by Frontal Affinity Chromatography (FAC). Then, two site-directed techniques were explored to increase column efficiency by immobilizing the antibody via its C-terminus, i.e., away from the antigen-binding site. In one approach, a tetra-lysine tail was added, and the antibody was immobilized onto DSC-activated POROS. In the second site-directed approach, the VHH was modified with the AviTag peptide, and a biotin-residue was enzymatically incorporated at the C-terminus using the biotin ligase BirA. The biotinylated antibody was subsequently immobilized onto NeutrAvidin-derivatized POROS. A comparison of the FAC analyses, which for all three columns showed excellent linearity (R(2)>0.999), revealed that both site-directed approaches yield better results than the random immobilization; the by far highest efficiency, however, was determined for the immunoaffinity column based on AviTag-biotinylated antibody. As proof of concept, all three columns were evaluated for quantification of MTX dissolved in phosphate buffered saline (PBS). Validation using UV-detection showed excellent linearity in the range of 0.04-12μM (R(2)>0.993). The lower limit of detection (LOD) and lower limit of quantification (LLOQ) were found to be independent of the immobilization strategy and were 40nM and 132nM, respectively. The intra- and inter-day precision was below 11.6%, and accuracy was between 90.7% and 112%. To the best of our knowledge, this is the first report of the AviTag-system in chromatography, and the first

  14. The antibody Hijikata Tatsumi

    Directory of Open Access Journals (Sweden)

    Éden Peretta

    2012-11-01

    Full Text Available Considered one of the most influential modern dance representatives in Japan, Tatsumi Hijikata’s work was a milestone in the Japanese post-war experimental artistic scene. Heretic son of his time, he staged a fertile mix of artistic and cultural influences, overlapping subversive elements of European arts and philosophy with radical references from pre-modern Japanese culture. In this way he built the foundations of its unstable antibody, its political-artistic project of dissolution of a organism, both physical and social.

  15. VIRAL ANTIBODIES IN PRESCHOOL CHILDREN

    Directory of Open Access Journals (Sweden)

    S. Saidi

    1974-08-01

    Full Text Available One hundred sera from children 1 - 6 years of age, representative of a large serum collection, were tested for the prevalence of antibodies against different viruses. Hemagglutination-inhibition (HI antibodies were found in 68% for measles; 61 % for rubella; 75'% for influenza A2/Hong Kong/68, 16% for influenza B/Md./59, 0% for group A arboviruses, 10% for group B arboviruses, 3% for phlebotomus fever group and 4% for Congo-Crimean hemorrhagic fever (C-CHF group of arboviruses Poliomyelitis-neutralizing antibodies for type 1, 2 and 3 were 90%; 85% and 84%~ respectively. Antibody to EH virus was detected in 84% of the sera by immuno-fluorescence. None of the sera were positive for hepatitis-B antigen or antibody by immuno-precipitation test. The prevalence of some viral antibodies found in this survey are compared with results obtained from surveys in other parts of the country.

  16. Metrics for antibody therapeutics development

    OpenAIRE

    Reichert, Janice M

    2010-01-01

    A wide variety of full-size monoclonal antibodies (mAbs) and therapeutics derived from alternative antibody formats can be produced through genetic and biological engineering techniques. These molecules are now filling the preclinical and clinical pipelines of every major pharmaceutical company and many biotechnology firms. Metrics for the development of antibody therapeutics, including averages for the number of candidates entering clinical study and development phase lengths for mAbs approv...

  17. Empowered Antibody Therapies - IBC conference.

    Science.gov (United States)

    Herold, Jens

    2010-10-01

    The Empowered Antibody Therapies conference, held in Burlingame, CA, USA, included topics covering new therapeutic developments in the field of multispecific antibodies. This conference report highlights selected presentations on DVD-Igs from Abbott Laboratories, ImmTACs from Immunocore, 'Dock-and-Lock' technology from Immunomedics, the bispecific BiTE antibody blinatumomab from Micromet, and Triomabs from TRION Pharma and Fresenius Biotech. PMID:20878591

  18. Monoclonal antibodies for treating cancer

    International Nuclear Information System (INIS)

    The purpose of this study is to assess the current status of in-vivo use of monoclonal antibodies for treating cancer. Publications appearing between 1980 and 1988 were identified by computer searches using MEDLINE and CANCERLIT, by reviewing the table of contents of recently published journals, and by searching bibliographies of identified books and articles. More than 700 articles, including peer-reviewed articles and book chapters, were identified and selected for analysis. The literature was reviewed and 235 articles were selected as relevant and representative of the current issues and future applications for in-vivo monoclonal antibodies for cancer therapy and of the toxicity and efficacy which has been associated with clinical trials. Approaches include using antibody alone (interacting with complement or effector cells or binding directly with certain cell receptors) and immunoconjugates (antibody coupled to radioisotopes, drugs, toxins, or other biologicals). Most experience has been with murine antibodies. Trials of antibody alone and radiolabeled antibodies have confirmed the feasibility of this approach and the in-vivo trafficking of antibodies to tumor cells. However, tumor cell heterogeneity, lack of cytotoxicity, and the development of human antimouse antibodies have limited clinical efficacy. Although the immunoconjugates are very promising, heterogeneity and the antimouse immune response have hampered this approach as has the additional challenge of chemically or genetically coupling antibody to cytotoxic agents. As a therapeutic modality, monoclonal antibodies are still promising but their general use will be delayed for several years. New approaches using human antibodies and reducing the human antiglobulin response should facilitate treatment. 235 references

  19. Antineutrophil cytoplasmic antibodies

    Directory of Open Access Journals (Sweden)

    G.D. Sebastiani

    2011-06-01

    Full Text Available Antineutrophil cytoplasmic antibodies (ANCA are predominantly IgG autoantibodies directed against constituents of primary granules of neutrophils and monocytes’ lysosomes. Although several antigenic targets have been identified, those ANCA directed to proteinase 3 or myeloperoxidase are clinically relevant, whereas the importance of other ANCA remains unknown. Both are strongly associated with small vessel vasculitides, the ANCA-associated vasculitides, which include Wegener’s granulomatosis, microscopic polyangiitis, and Churg-Strauss syndrome, and the localised forms of these diseases (eg, pauci-immune necrotising and crescentic glomerulonephritis. ANCA is a useful serological test to assist in diagnosis of small-vessel vasculitides. 85-95% of patients with Wegener’s granulomatosis, microscopic polyangiitis, and pauci-immune necrotising and crescentic glomerulonephritis have serum ANCA. ANCA directed to either proteinase 3 or myeloperoxidase are clinically relevant, yet the relevance of other ANCA remains unknown. Besides their diagnostic potential, ANCA might be valuable in disease monitoring. In addition, data seem to confirm the long-disputed pathogenic role of these antibodies. There is increasing evidence that myeloperoxidase- ANCA are directly involved in the pathogenesis of necrotizing vasculitis. This is less clear for proteinase 3-ANCA, markers for Wegener’s granulomatosis. With respect to proteinase 3-ANCA, complementary proteinase 3, a peptide translated from the antisense DNA strand of proteinase 3 and homologous to several microbial peptides, may be involved in induction of proteinase 3-antineutrophil cytoplasmic autoantibodies.

  20. Targeting of Antibodies using Aptamers

    OpenAIRE

    Missailidis, Sotiris

    2003-01-01

    The chapter presents a methodology for the rapid selection of aptamers against antibody targets. It is a detailed account of the various methodological steps that describe the selection of aptamers, including PCR steps, buffers to be used, target immobilisation, partitioning and amplification of aptamers, clonning and sequencing, to results in high affinity and specificity ligands for the chosen target antibody.

  1. Structural Characterization of Peptide Antibodies

    DEFF Research Database (Denmark)

    Chailyan, Anna; Marcatili, Paolo

    2015-01-01

    The role of proteins as very effective immunogens for the generation of antibodies is indisputable. Nevertheless, cases in which protein usage for antibody production is not feasible or convenient compelled the creation of a powerful alternative consisting of synthetic peptides. Synthetic peptide...

  2. Pathogenic role of antiphospholipid antibodies

    NARCIS (Netherlands)

    Salmon, J. E.; de Groot, P. G.

    2008-01-01

    The antiphospholipid antibody syndrome (APS) is characterized by recurrent arterial and venous thrombosis and/or pregnancy in association with antiphospholipid (aPL) antibodies. The pathogenic mechanisms in APS that lead to in vivo injury are incompletely understood. Recent evidence suggests that AP

  3. Metrics for antibody therapeutics development.

    Science.gov (United States)

    Reichert, Janice M

    2010-01-01

    A wide variety of full-size monoclonal antibodies (mAbs) and therapeutics derived from alternative antibody formats can be produced through genetic and biological engineering techniques. These molecules are now filling the preclinical and clinical pipelines of every major pharmaceutical company and many biotechnology firms. Metrics for the development of antibody therapeutics, including averages for the number of candidates entering clinical study and development phase lengths for mAbs approved in the United States, were derived from analysis of a dataset of over 600 therapeutic mAbs that entered clinical study sponsored, at least in part, by commercial firms. The results presented provide an overview of the field and context for the evaluation of on-going and prospective mAb development programs. The expansion of therapeutic antibody use through supplemental marketing approvals and the increase in the study of therapeutics derived from alternative antibody formats are discussed. PMID:20930555

  4. Metrics for antibody therapeutics development.

    Science.gov (United States)

    Reichert, Janice M

    2010-01-01

    A wide variety of full-size monoclonal antibodies (mAbs) and therapeutics derived from alternative antibody formats can be produced through genetic and biological engineering techniques. These molecules are now filling the preclinical and clinical pipelines of every major pharmaceutical company and many biotechnology firms. Metrics for the development of antibody therapeutics, including averages for the number of candidates entering clinical study and development phase lengths for mAbs approved in the United States, were derived from analysis of a dataset of over 600 therapeutic mAbs that entered clinical study sponsored, at least in part, by commercial firms. The results presented provide an overview of the field and context for the evaluation of on-going and prospective mAb development programs. The expansion of therapeutic antibody use through supplemental marketing approvals and the increase in the study of therapeutics derived from alternative antibody formats are discussed.

  5. The antineutrophil antibody in uveitis.

    OpenAIRE

    Young, D W

    1991-01-01

    Ninety eight patients with uveitis of various types were tested for the presence of the antineutrophil antibody or ANCA by an indirect immunofluorescence method. This antibody is found in patients with diseases associated with small vessel vasculitis, including Wegener's granulomatosis and microscopic polyarteritis. Eleven true positive cases were found. A positive test was not associated with the anatomical site of the uveitis but was related to the time course of the disease. In particular ...

  6. Functional effects of anticardiolipin antibodies.

    Science.gov (United States)

    Harris, E N; Pierangeli, S S

    1996-10-01

    The 'lupus anticoagulant' phenomenon is the best documented functional effect of antiphospholipid (aPL) antibodies, occurring either by inhibition of the prothrombinase and/or Factor X activation reactions. Understanding the mechanism by which aPL antibodies inhibit phospholipid dependent coagulation reactions may yield important clues about their 'thrombogenic effects' in vivo. We conducted a series of studies to determine the specificity, diversity, and mechanism by which aPL antibodies inhibit phospholipid dependent reactions. Results showed that purified immunoglobulins with lupus anticoagulant and anti-cardiolipin activities were absorbed by negatively charged phospholipids and both activities were recovered from the phospholipid-antibody precipitate. Purified aPL antibodies inhibited the prothrombinase reaction in a plasma free system in which beta 2-glycoprotein 1 (beta 2-GP1) was absent. Affinity purified aPL antibodies had 25-50 times the inhibitory activity of immunoglobulin preparations. The phospholipid binding proteins, beta 2-GPI and placental anticoagulant protein I (PAP I), independently inhibited the prothrombinase reaction, and when these proteins were combined with aPL, inhibition of the prothrombinase reaction was additive. Antibodies of syphilis had no inhibitory effect, partially accounted for by lack of specificity for phosphotidylserine (PS). Although aPL antibodies inhibited the protein C activation reaction, there was no correlation of these activities with inhibition of the prothrombinase reaction. Together, these results show that aPL exert their effects by interaction with negatively charged phospholipids, in particular phosphotidylserine, but lack of correlation between inhibition of the prothrombinase and protein C activation reactions, suggests that the nature of the coagulation protein is also important. PMID:8902763

  7. Antibodies to watch in 2014.

    Science.gov (United States)

    Reichert, Janice M

    2014-01-01

    Since 2010, mAbs has documented the biopharmaceutical industry's progress in transitioning antibody therapeutics to first Phase 3 clinical studies and regulatory review, and its success at gaining first marketing approvals for antibody-based products. This installment of the "Antibodies to watch" series outlines events anticipated to occur between December 2013 and the end of 2014, including first regulatory actions on marketing applications for vedolizumab, siltuximab, and ramucirumab, as well as the Fc fusion proteins Factor IX-Fc and Factor VIII-Fc; and the submission of first marketing applications for up to five therapeutics (secukinumab, ch14.18, onartuzumab, necitumumab, gevokizumab). Antibody therapeutics in Phase 3 studies are described, with an emphasis on those with study completion dates in 2014, including antibodies targeting interleukin-17a or the interleukin-17a receptor (secukinumab, ixekizumab, brodalumab), proprotein convertase subtilisin/kexin type 9 (alirocumab, evolocumab, bococizumab), and programmed death 1 receptor (lambrolizumab, nivolumab). Five antibodies with US Food and Drug Administration's Breakthrough Therapy designation (obinutuzumab, ofatumumab, lambrolizumab, bimagrumab, daratumumab) are also discussed. PMID:24284914

  8. Radiolabeled monoclonal antibodies: a review

    International Nuclear Information System (INIS)

    Since the description by Kohler and Milstein 1975 of their technique for producing monoclonal antibodies of predefined specificity, it has become a mainstay in most laboratories that utilize immunochemical techniques to study problems in basic, applied or clinical research. Paradoxically, the very success of monoclonal antibodies has generated a literature which is now so vast and scattered that it has become difficult to obtain a perspective. This brief review represents the distillation of many publications relating to the production and use of monoclonaal antibodies as radiopharmaceuticals. Significant advances were made possible in the last few years by combined developments in the fields of tumor-associated antigens and of monoclonal antibodies. In fact monoclonal antibodies against some well defined tumor-associated antigens, has led to significantly greater practical possibilities for producing highly specific radiolabeled antibodies as radiopharmaceuticals for diagnosis and therapy of human tumors. One of the main requirements of this methodology is the availability of stable radiopharmaceutical reagents which after labeling in vivo injection retain the capacity of specific interaction with the defined antigen and their molecular integrity. Since injection into human is the objetive of this kind of study all the specifications of radiopharmaceutical have to be fulfilled e.g. sterility, apirogenicity and absence of toxicity. (author)

  9. Radioimmunoguided surgery using monoclonal antibody

    International Nuclear Information System (INIS)

    The potential proficiency of radioimmunoguided surgery in the intraoperative detection of tumors was assessed using labeled monoclonal antibody B72.3 in 66 patients with tissue-proved tumor. Monoclonal antibody B72.3 was injected 5 to 42 days preoperatively, and the hand-held gamma-detecting probe was used intraoperatively to detect the presence of tumor. Intraoperative probe counts of less than 20 every 2 seconds, or tumor-to-adjacent normal tissue ratios less than 2:1 were considered negative (system failure). Positive probe counts were detected in 5 of 6 patients with primary colon cancer (83 percent), in 31 of 39 patients with recurrent colon cancer (79 percent), in 4 of 5 patients with gastric cancer (80 percent), in 3 of 8 patients with breast cancer (37.5 percent), and in 4 of 8 patients with ovarian cancer (50 percent) undergoing second-look procedures. Additional patients in each group were scored as borderline positive. Overall, radioimmunoguided surgery using B72.3 identified tumors in 47 patients (71.2 percent), bordered on positive in 6 patients (9.1 percent), and failed to identify tumor in 13 patients (19.7 percent). Improved selection of patients for antigen-positive tumors, the use of higher affinity second-generation antibodies, alternate routes of antibody administration, alternate radionuclides, and more sophisticatedly bioengineered antibodies and antibody combinations should all lead to improvements in radioimmunoguided surgery

  10. Tabhu: tools for antibody humanization.

    KAUST Repository

    Olimpieri, Pier Paolo

    2014-10-09

    SUMMARY: Antibodies are rapidly becoming essential tools in the clinical practice, given their ability to recognize their cognate antigens with high specificity and affinity, and a high yield at reasonable costs in model animals. Unfortunately, when administered to human patients, xenogeneic antibodies can elicit unwanted and dangerous immunogenic responses. Antibody humanization methods are designed to produce molecules with a better safety profile still maintaining their ability to bind the antigen. This can be accomplished by grafting the non-human regions determining the antigen specificity into a suitable human template. Unfortunately, this procedure may results in a partial or complete loss of affinity of the grafted molecule that can be restored by back-mutating some of the residues of human origin to the corresponding murine ones. This trial-and-error procedure is hard and involves expensive and time-consuming experiments. Here we present tools for antibody humanization (Tabhu) a web server for antibody humanization. Tabhu includes tools for human template selection, grafting, back-mutation evaluation, antibody modelling and structural analysis, helping the user in all the critical steps of the humanization experiment protocol. AVAILABILITY: http://www.biocomputing.it/tabhu CONTACT: anna.tramontano@uniroma1.it, pierpaolo.olimpieri@uniroma1.it SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

  11. Avian Diagnostic and Therapeutic Antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Bradley, David Sherman [UND SMHS

    2012-12-31

    A number of infectious agents have the potential of causing significant clinical symptomology and even death, but dispite this, the number of incidence remain below the level that supports producing a vaccine. Therapeutic antibodies provide a viable treatment option for many of these diseases. We proposed that antibodies derived from West Nile Virus (WNV) immunized geese would be able to treat WNV infection in mammals and potential humans. We demonstrated that WNV specific goose antibodies are indeed successful in treating WNV infection both prophylactically and therapeutically in a golden hamster model. We demonstrated that the goose derived antibodies are non-reactogenic, i.e. do not cause an inflammatory response with multiple exposures in mammals. We also developed both a specific pathogen free facility to house the geese during the antibody production phase and a patent-pending purification process to purify the antibodies to greater than 99% purity. Therefore, the success of these study will allow a cost effective rapidly producible therapeutic toward clinical testing with the necessary infrastructure and processes developed and in place.

  12. Production and characterization of peptide antibodies

    DEFF Research Database (Denmark)

    Trier, Nicole Hartwig; Hansen, Paul Robert; Houen, Gunnar

    2012-01-01

    Proteins are effective immunogens for generation of antibodies. However, occasionally the native protein is known but not available for antibody production. In such cases synthetic peptides derived from the native protein are good alternatives for antibody production. These peptide antibodies are...

  13. Production of recombinant antibodies using bacteriophages

    OpenAIRE

    Shukra, A. M.; Sridevi, N. V.; Dev Chandran,; Kapil Maithal,

    2014-01-01

    Recombinant antibody fragments such as Fab, scFv, diabodies, triabodies, single domain antibodies and minibodies have recently emerged as potential alternatives to monoclonal antibodies, which can be engineered using phage display technology. These antibodies match the strengths of conventionally produced monoclonal antibodies and offer advantages for the development of immunodiagnostic kits and assays. These fragments not only retain the specificity of the whole monoclonal ...

  14. Antibody-Directed Phototherapy (ADP

    Directory of Open Access Journals (Sweden)

    M. Adil Butt

    2013-04-01

    Full Text Available Photodynamic therapy (PDT is a clinically-approved but rather under-exploited treatment modality for cancer and pre-cancerous superficial lesions. It utilises a cold laser or LED to activate a photochemical reaction between a light activated drug (photosensitiser-drug and oxygen to generate cytotoxic oxygen species. These free radical species damage cellular components leading to cell death. Despite its benefits, the complexity, limited potency and side effects of PDT have led to poor general usage. However, the research area is very active with an increasing understanding of PDT-related cell biology, photophysics and significant progress in molecular targeting of disease. Monoclonal antibody therapy is maturing and the next wave of antibody therapies includes antibody-drug conjugates (ADCs, which promise to be more potent and curable. These developments could lift antibody-directed phototherapy (ADP to success. ADP promises to increase specificity and potency and improve drug pharmacokinetics, thus delivering better PDT drugs whilst retaining its other benefits. Whole antibody conjugates with first generation ADP-drugs displayed problems with aggregation, poor pharmacokinetics and loss of immuno-reactivity. However, these early ADP-drugs still showed improved selectivity and potency. Improved PS-drug chemistry and a variety of conjugation strategies have led to improved ADP-drugs with retained antibody and PS-drug function. More recently, recombinant antibody fragments have been used to deliver ADP-drugs with superior drug loading, more favourable pharmacokinetics, enhanced potency and target cell selectivity. These improvements offer a promise of better quality PDT drugs.

  15. Antibodies: an alternative for antibiotics?

    Science.gov (United States)

    Berghman, L R; Abi-Ghanem, D; Waghela, S D; Ricke, S C

    2005-04-01

    In 1967, the success of vaccination programs, combined with the seemingly unstoppable triumph of antibiotics, prompted the US Surgeon General to declare that "it was time to close the books on infectious diseases." We now know that the prediction was overly optimistic and that the fight against infectious diseases is here to stay. During the last 20 yr, infectious diseases have indeed made a staggering comeback for a variety of reasons, including resistance against existing antibiotics. As a consequence, several alternatives to antibiotics are currently being considered or reconsidered. Passive immunization (i.e., the administration of more or less pathogen-specific antibodies to the patient) prior to or after exposure to the disease-causing agent is one of those alternative strategies that was almost entirely abandoned with the introduction of chemical antibiotics but that is now gaining interest again. This review will discuss the early successes and limitations of passive immunization, formerly referred to as "serum therapy," the current use of antibody administration for prophylaxis or treatment of infectious diseases in agriculture, and, finally, recent developments in the field of antibody engineering and "molecular farming" of antibodies in various expression systems. Especially the potential of producing therapeutic antibodies in crops that are routine dietary components of farm animals, such as corn and soy beans, seems to hold promise for future application in the fight against infectious diseases. PMID:15844826

  16. Antibodies to watch in 2016.

    Science.gov (United States)

    Reichert, Janice M

    2016-01-01

    The number of novel antibody therapeutics that received first marketing approvals in 2015 met expectations, with 6 (alirocumab (Praluent®), evolocumab (Repatha®), daratumumab (Darzalex®), dinutuximab (Unituxin®), idarucizumab (Praxbind®), mepolizumab (Nucala®)) granted first approvals as of mid-November*. Seven novel antibody therapeutics (begelomab, brodalumab, elotuzumab, ixekizumab, necitumumab, obiltoxaximab, reslizumab) are in regulatory review, and thus a similar number, if not more, are projected to gain first approvals in 2016. Commercial late-stage antibody therapeutics development exceeded expectations by increasing from 39 candidates in Phase 3 studies as of late 2014 to 53 as of late 2015. Of the 53 candidates, transitions to regulatory review by the end of 2016 are projected for 8 (atezolizumab, benralizumab, bimagrumab, durvalumab, inotuzumab ozogamicin, lebrikizumab, ocrelizumab, tremelimumab). Other "antibodies to watch" include 15 candidates (bavituximab, bococizumab, dupilumab, fasinumab, fulranumab, gevokizumab, guselkumab, ibalizumab, LY2951742, onartuzumab, REGN2222, roledumab, romosozumab, sirukumab, Xilonix) undergoing evaluation in Phase 3 studies that have estimated primary completion dates in 2016. As evidenced by the antibody therapeutics discussed in this perspective, the biopharmaceutical industry has a highly active late-stage clinical pipeline that may deliver numerous new products to the global market in the near future. *See Note added in proof for updates through December 31, 2015. PMID:26651519

  17. Epigenetics of the antibody response.

    Science.gov (United States)

    Li, Guideng; Zan, Hong; Xu, Zhenming; Casali, Paolo

    2013-09-01

    Epigenetic marks, such as DNA methylation, histone post-translational modifications and miRNAs, are induced in B cells by the same stimuli that drive the antibody response. They play major roles in regulating somatic hypermutation (SHM), class switch DNA recombination (CSR), and differentiation to plasma cells or long-lived memory B cells. Histone modifications target the CSR and, possibly, SHM machinery to the immunoglobulin locus; they together with DNA methylation and miRNAs modulate the expression of critical elements of that machinery, such as activation-induced cytidine deaminase (AID), as well as factors central to plasma cell differentiation, such as B lymphocyte-induced maturation protein-1 (Blimp-1). These inducible B cell-intrinsic epigenetic marks instruct the maturation of antibody responses. Their dysregulation plays an important role in aberrant antibody responses to foreign antigens, such as those of microbial pathogens, and self-antigens, such as those targeted in autoimmunity, and B cell neoplasia.

  18. Molecular-specific urokinase antibodies

    Science.gov (United States)

    Atassi, M. Zouhair (Inventor); Morrison, Dennis R. (Inventor)

    2009-01-01

    Antibodies have been developed against the different molecular forms of urokinase using synthetic peptides as immunogens. The peptides were synthesized specifically to represent those regions of the urokinase molecules which are exposed in the three-dimensional configuration of the molecule and are uniquely homologous to urokinase. Antibodies are directed against the lysine 158-isoleucine 159 peptide bond which is cleaved during activation from the single-chain (ScuPA) form to the bioactive double chain (54 KDa and 33 KDa) forms of urokinase and against the lysine 135 lysine 136 bond that is cleaved in the process of removing the alpha-chain from the 54 KDa form to produce the 33 KDa form of urokinase. These antibodies enable the direct measurement of the different molecular forms of urokinase from small samples of conditioned medium harvested from cell cultures.

  19. Tabhu: tools for antibody humanization

    DEFF Research Database (Denmark)

    Olimpieri, Pier Paolo; Marcatili, Paolo; Tramontano, Anna

    2015-01-01

    into a suitable human template. Unfortunately, this procedure may results in a partial or complete loss of affinity of the grafted molecule that can be restored by back-mutating some of the residues of human origin to the corresponding murine ones. This trial-and-error procedure is hard and involves expensive......Antibodies are rapidly becoming essential tools in the clinical practice, given their ability to recognize their cognate antigens with high specificity and affinity, and a high yield at reasonable costs in model animals. Unfortunately, when administered to human patients, xenogeneic antibodies can...... elicit unwanted and dangerous immunogenic responses. Antibody humanization methods are designed to produce molecules with a better safety profile still maintaining their ability to bind the antigen. This can be accomplished by grafting the non-human regions determining the antigen specificity...

  20. Autologous antibodies that bind neuroblastoma cells.

    Science.gov (United States)

    Sun, Yujing; Sholler, Giselle S; Shukla, Girja S; Pero, Stephanie C; Carman, Chelsea L; Zhao, Ping; Krag, David N

    2015-11-01

    Antibody therapy of neuroblastoma is promising and our goal is to derive antibodies from patients with neuroblastoma for developing new therapeutic antibodies. The feasibility of using residual bone marrow obtained for clinical indications as a source of tumor cells and a source of antibodies was assessed. From marrow samples, neuroblastoma cells were recovered, grown in cell culture and also implanted into mice to create xenografts. Mononuclear cells from the marrow were used as a source to generate phage display antibody libraries and also hybridomas. Growth of neuroblastoma patient cells was possible both in vitro and as xenografts. Antibodies from the phage libraries and from the monoclonal hybridomas bound autologous neuroblastoma cells with some selectivity. It appears feasible to recover neuroblastoma cells from residual marrow specimens and to generate human antibodies that bind autologous neuroblastoma cells. Expansion of this approach is underway to collect more specimens, optimize methods to generate antibodies, and to evaluate the bioactivity of neuroblastoma-binding antibodies.

  1. Radioimmunotherapy with engineered antibody fragments

    International Nuclear Information System (INIS)

    Authors have developed and begun evaluating radiometal-chelated (213Bi) engineered antibody fragments as radioimmunotherapy agents that target the HER2/neu (c-erbB-2) antigen. The diabody format was found to have 40-fold greater affinity for HER2/neu and to be associated with significantly greater tumor localization than is achieved with scFv molecule. It is shown that short-lived isotopes like 213Bi would be most effective when used in conjunction with antibodies that targeted diffuse malignancies (leukemia or lymphoma) or when used for very rapid pretargeted radioimmunotherapy application in which the radioisotope is conjugated to a very small ligand

  2. Antibody profiling sensitivity through increased reporter antibody layering

    Energy Technology Data Exchange (ETDEWEB)

    Apel, William A.; Thompson, Vicki S.

    2013-02-26

    A method for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an embodiment of the invention, the analyte is a drug, such as marijuana, Cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method comprises attaching antigens to a surface of a solid support in a preselected pattern to form an array wherein locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to the antigens in the array to form immune complexes; washing away antibodies that do form immune complexes; and detecting the immune complexes, to form an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to the subject's identity.

  3. Antibody profiling sensitivity through increased reporter antibody layering

    Energy Technology Data Exchange (ETDEWEB)

    Apel, William A; Thompson, Vicki S

    2013-02-26

    A method for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an embodiment of the invention, the analyte is a drug, such as marijuana, Cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method comprises attaching antigens to a surface of a solid support in a preselected pattern to form an array wherein locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to the antigens in the array to form immune complexes; washing away antibodies that do form immune complexes; and detecting the immune complexes, to form an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to the subject's identity.

  4. Antibody profiling sensitivity through increased reporter antibody layering

    Energy Technology Data Exchange (ETDEWEB)

    Apel, William A.; Thompson, Vicki S

    2010-04-13

    A method for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an embodiment of the invention, the analyte is a drug, such as marijuana, Cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method comprises attaching antigens to a surface of a solid support in a preselected pattern to form an array wherein locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to the antigens in the array to form immune complexes; washing away antibodies that do form immune complexes; and detecting the immune complexes, to form an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to the subject's identity.

  5. Antibody profiling sensitivity through increased reporter antibody layering

    International Nuclear Information System (INIS)

    A method for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an embodiment of the invention, the analyte is a drug, such as marijuana, Cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method comprises attaching antigens to a surface of a solid support in a preselected pattern to form an array wherein locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to the antigens in the array to form immune complexes; washing away antibodies that do form immune complexes; and detecting the immune complexes, to form an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to the subject's identity.

  6. Pharmacokinetics interactions of monoclonal antibodies.

    Science.gov (United States)

    Ferri, Nicola; Bellosta, Stefano; Baldessin, Ludovico; Boccia, Donatella; Racagni, Giorgi; Corsini, Alberto

    2016-09-01

    The clearance of therapeutic monoclonal antibodies (mAbs) typically does not involve cytochrome P450 (CYP450)-mediated metabolism or interaction with cell membrane transporters, therefore the pharmacokinetics interactions of mAbs and small molecule drugs are limited. However, a drug may affect the clearance of mAbs through the modulation of immune response (e.g., methotrexate reduces the clearance of infliximab, adalimumab, and golimumab, possibly due to methotrexate's inhibitory effect on the formation of antibodies against the mAbs). In addition, mAbs that are cytokine modulators may modify the metabolism of drugs through their effects on P450 enzymes expression. For example, cytokine modulators such as tocilizumab (anti-IL-6 receptor antibody) may reverse the "inhibitory" effect of IL-6 on CYP substrates, resulting in a "normalization" of CYP activities. Finally, a drug may alter the clearance of mAbs by either increasing or reducing the levels of expression of targets of mAbs on the cell surface. For instance, statins and fibrates induce PCSK9 expression and therefore increase cellular uptake and clearance of alirocumab and evolocumab, anti-PCSK9 antibodies. In the present review, we will provide an overview on the pharmacokinetics properties of mAbs as related to the most relevant examples of mAbs-small molecule drug interaction.

  7. Alternative affinity tools: more attractive than antibodies?

    NARCIS (Netherlands)

    Ruigrok, V.J.B.; Levisson, M.; Eppink, M.H.M.; Smidt, H.; Oost, van der J.

    2011-01-01

    Antibodies are the most successful affinity tools used today, in both fundamental and applied research (diagnostics, purification and therapeutics). Nonetheless, antibodies do have their limitations, including high production costs and low stability. Alternative affinity tools based on nucleic acids

  8. Detection of Campylobacter species using monoclonal antibodies

    Science.gov (United States)

    Young, Colin R.; Lee, Alice; Stanker, Larry H.

    1999-01-01

    A panel of species specific monoclonal antibodies were raised to Campylobacter coli, Campylobacter jejuni and Campylobacter lari. The isotypes, and cross-reactivity profiles of each monoclonal antibody against an extensive panel of micro- organisms, were determined.

  9. [Neuroimmunological diseases associated with VGKC complex antibodies].

    Science.gov (United States)

    Watanabe, Osamu

    2013-05-01

    Antibodies to voltage-gated potassium channels(VGKC) were first identified by radioimmunoassay of radioisotope labeled alpha-dendrotoxin-VGKCs solubilized from rabbit brain. These antibodies were found only in a proportion of patients with acquired neuromyotonia (Isaacs' syndrome). VGKC antibodies were also detected in Morvan's syndrome and in a form of autoimmune limbic encephalitis. Recent studies indicated that the "VGKC" antibodies are mainly directed toward associated proteins(for example LGI-1, Caspr-2) that complex with the VGKCs themselves. The "VGKC" antibodies are now usually known as VGKC-complex antibodies. In general, LGI-1 antibodies are most common in limbic encephalitis with SIADH. Caspr-2 antibodies are present in the majority of patients with Morvan's syndrome. These patients develop combinations of CNS symptoms, autonomic dysfunction, and peripheral nerve hyperexcitability.

  10. Production and Screening of Monoclonal Peptide Antibodies.

    Science.gov (United States)

    Trier, Nicole Hartwig; Mortensen, Anne; Schiolborg, Annette; Friis, Tina

    2015-01-01

    Hybridoma technology is a remarkable and indispensable tool for generating high-quality monoclonal antibodies. Hybridoma-derived monoclonal antibodies not only serve as powerful research and diagnostic reagents, but have also emerged as the most rapidly expanding class of therapeutic biologicals. In this chapter, an overview of hybridoma technology and the laboratory procedures used routinely for hybridoma production and antibody screening are presented, including characterization of peptide antibodies.

  11. Virus Strain Discrimination Using Recombinant Antibodies

    OpenAIRE

    Boonham, N.; Barker, I.

    2002-01-01

    Most routine testing for plant viruses is currently carried out using monoclonal and polyclonal antibodies. Traditional methods of antibody production however can be time consuming and require the use of expensive cell culture facilities. Recombinant antibody technology however is starting to make an impact in this area, enabling the selection of antibody fragments in a few weeks compared with the many months associated with traditional methods and requires only basic microbiological faciliti...

  12. Sequence and structural analysis of antibodies

    OpenAIRE

    Raghavan, A. K.

    2009-01-01

    The work presented in this thesis focusses on the sequence and structural analysis of antibodies and has fallen into three main areas. First I developed a method to assess how typical an antibody sequence is of the expressed human antibody repertoire. My hypothesis was that the more \\humanlike" an antibody sequence is (in other words how typical it is of the expressed human repertoire), the less likely it is to elicit an immune response when used in vivo in humans. In practi...

  13. Immunoglobulin G4: an odd antibody

    NARCIS (Netherlands)

    R.C. Aalberse; S.O. Stapel; J. Schuurman; T. Rispens

    2009-01-01

    Despite its well-known association with IgE-mediated allergy, IgG4 antibodies still have several poorly understood characteristics. IgG4 is a very dynamic antibody: the antibody is involved in a continuous process of half-molecules (i.e. a heavy and attached light-chain) exchange. This process, also

  14. Anti-DNA antibodies in SLE

    Energy Technology Data Exchange (ETDEWEB)

    Voss, E.W.

    1988-01-01

    This book contains 8 chapters. Some of the titles are: Anti-DNA Antibodies in SLE: Historical Perspective; Specificity of Anti-DNA Antibodies in Systemic Lupus Erythematosus; Monoclonial Autoimmune Anti-DNA Antibodies; and Structure--Function Analyses of Anti-DNA Autoantibodies.

  15. The role of antibodies in myasthenia gravis.

    Science.gov (United States)

    De Baets, M; Stassen, M H W

    2002-10-15

    Myasthenia gravis is an autoimmune disease associated with antibodies directed to the postsynaptic acetylcholine receptor. These antibodies reduce the number of receptors. Autoantibodies against AChR and other muscle antigens can be used for the diagnosis of myasthenia gravis and related disorders. The origin and the role of these antibodies in the disease are discussed. Experimental autoimmune myasthenia gravis, an experimental model closely mimicking the disease, has provided answers to many questions about the role of antibodies, complement macrophages and AChR anchor proteins. Genetically modified anti-AChR antibodies may also be used in the future to treat myasthenia. PMID:12220686

  16. Production of Monoclonal Antibody against Human Nestin.

    Science.gov (United States)

    Hadavi, Reza; Zarnani, Amir Hassan; Ahmadvand, Negah; Mahmoudi, Ahmad Reza; Bayat, Ali Ahmad; Mahmoudian, Jafar; Sadeghi, Mohammad-Reza; Soltanghoraee, Haleh; Akhondi, Mohammad Mehdi; Tarahomi, Majid; Jeddi-Tehrani, Mahmood; Rabbani, Hodjattallah

    2010-04-01

    We have employed a peptide-based antibody generation protocol for producing antibody against human nestin. Using a 12-mer synthetic peptide from repetitive region of human nestin protein devoid of any N- or O-glyco-sylation sequences, we generated a mouse monoclonal antibody capable of recognizing human, mouse, bovine, and rat nestin. A wide variety of nestin proteins ranging from 140-250 kDa was detected by this antibody. This antibody is highly specific and functional in applications such as ELISA, flow cytometry, immunocytochemistry, and Western blot assays. PMID:23407796

  17. Production of Monoclonal Antibody against Human Nestin

    OpenAIRE

    Hadavi, Reza; Zarnani, Amir Hassan; Ahmadvand, Negah; Mahmoudi, Ahmad Reza; Bayat, Ali Ahmad; Mahmoudian, Jafar; Sadeghi, Mohammad-Reza; Soltanghoraee, Haleh; Akhondi, Mohammad mehdi; Tarahomi, Majid; Jeddi-Tehrani, Mahmood; Rabbani, Hodjattallah

    2010-01-01

    We have employed a peptide-based antibody generation protocol for producing antibody against human nestin. Using a 12-mer synthetic peptide from repetitive region of human nestin protein devoid of any N- or O-glyco-sylation sequences, we generated a mouse monoclonal antibody capable of recognizing human, mouse, bovine, and rat nestin. A wide variety of nestin proteins ranging from 140–250 kDa was detected by this antibody. This antibody is highly specific and functional in applications such a...

  18. Monoclonal Antibody Therapies against Anthrax

    OpenAIRE

    Zhaochun Chen; Mahtab Moayeri; Robert Purcell

    2011-01-01

    Anthrax is a highly lethal infectious disease caused by the spore-forming bacterium Bacillus anthracis. It not only causes natural infection in humans but also poses a great threat as an emerging bioterror agent. The lethality of anthrax is primarily attributed to the two major virulence factors: toxins and capsule. An extensive effort has been made to generate therapeutically useful monoclonal antibodies to each of the virulence components: protective antigen (PA), lethal factor (LF) and ede...

  19. Antibody Peptide Based Antifungal Immunotherapy

    OpenAIRE

    Magliani, Walter; Conti, Stefania; Giovati, Laura; Zanello, Pier Paolo; Sperindè, Martina; Ciociola, Tecla; Polonelli, Luciano

    2012-01-01

    Fungal infections still represent relevant human illnesses worldwide and some are accompanied by unacceptably high mortality rates. The limited current availability of effective and safe antifungal agents makes the development of new drugs and approaches of antifungal vaccination/immunotherapy every day more needed. Among them, small antibody(Ab)-derived peptides are arousing great expectations as new potential antifungal agents. In this topic, the search path from the study of the yeast kill...

  20. Construction of phage display VHH antibody library against avian H5N1 virus from alpaca%抗H5N1禽流感病毒VHH抗体库的构建

    Institute of Scientific and Technical Information of China (English)

    严安; 熊慧; 王颖; 孙冰玉; 夏立亮; 吴标; 包文静; 车小燕; 孙志伟; 金维荣; 赵国屏

    2011-01-01

    Objective: To construct phage display variable domain of heavy chain antibody library (VHH antibody library) from alpaca immunized with inactivated H5N1 vaccine for the future screening of VHH antibodies against avian H5N1 influenza virus. Methods: The camelid species (alpaca, Lama pacos ) was selected for immunization with inactivated H5N1 vaccine. Hemagglutinin inhibition (HI) assay of serum from immunized alpaca was performed against H5N1 avian influenza virus one week after the fourth inntmization. Peripheral lymphocytes were isolated for the amplification of VHH fragments by RT-PCR. PCR products were then purified and inserted into phagemid vector pCANTAB5E. The VHH antibody gene library was obtained by electroporating recombinant pCANTAB5E-VHH vectors into E. coli TG1 cells.The library capacity and diversity of VHH antibody gene library was determined by sequencing analysis. The HI assay was performed with the culture supernatant of primary phage display VHH antibody library. Results:After four rounds of immunization with inactivated H5N1 vaccine,HI antibody titer of the alpaca serum reached to 1: 2 560, which was higher than those fiom immunized mice. A first set of antibody gene library totalling 3 × l08 members were created after cloning VHH genes into a phagemid vector pCANTAB5E. The sequence of 14 members of the unselected library indicated that the camelid VHH antibody library we constructed possessed high diversity and good capacity. The supematant from the primary phage display library displayed effieient HI effect against avian H5N1 influenza virus. And the titration of our phage display VHH library reached 2.17 × l011. Conclusion: Taken together, phage display VHH antibody library from immunized alpaca is successfully constructed,which provides a platform for VHH antibody preparation against H5N1 virus. This will give light to future study on treatment and diagnosis of avian H5N1 influenza virus.%目的:构建抗H5N1禽流感病毒的小羊驼免

  1. Application of Monoclonal Antibodies in Veterinary Parasitology

    Directory of Open Access Journals (Sweden)

    Gupta A.

    2011-08-01

    Full Text Available The discovery of hybridoma technology by Kohler and Milstein in 1975, heralded a new era in antibody research. Mouse hybridomas were the first reliable source of monoclonal antibodies. The generation of monoclonal antibodies from species other than rats and mice, has developed slowly over the last 30 years. The advent of antibody engineering and realization of the advantages of non murine antibodies has increased their relevance recently. However, in the area of veterinary parasitology, monoclonal antibodies are just beginning to fulfill the promises inherent in their great specificity for recognizing and selectively binding to antigens. This review describes the recent advances in the application of monoclonal antibodies for immunodiagnosis / prophylaxis and immunotherapy of parasitic diseases. [Vet. World 2011; 4(4.000: 183-188

  2. Advances in monoclonal antibody application in myocarditis

    Institute of Scientific and Technical Information of China (English)

    Li-na HAN; Shuang HE; Yu-tang WANG; Li-ming YANG; Si-yu LIU; Ting ZHANG

    2013-01-01

    Monoclonal antibodies have become a part of daily preparation technologies in many laboratories.Attempts have been made to apply monoclonal antibodies to open a new train of thought for clinical treatments of autoimmune diseases,inflammatory diseases,cancer,and other immune-associated diseases.This paper is a prospective review to anticipate that monoclonal antibody application in the treatment of myocarditis,an inflammatory disease of the heart,could be a novel approach in the future.In order to better understand the current state of the art in monoclonal antibody techniques and advance applications in myocarditis,we,through a significant amount of literature research both domestic and abroad,developed a systematic elaboration of monoclonal antibodies,pathogenesis of myocarditis,and application of monoclonal antibodies in myocarditis.This paper presents review of the literature of some therapeutic aspects of monoclonal antibodies in myocarditis and dilated cardiomyopathy to demonstrate the advance of monoclonal antibody application in myocarditis and a strong anticipation that monoclonal antibody application may supply an effective therapeutic approach to relieve the severity of myocarditis in the future.Under conventional therapy,myocarditis is typically associated with congestive heart failure as a progressive outcome,indicating the need for alternative therapeutic strategies to improve long-term results.Reviewing some therapeutic aspects of monoclonal antibodies in myocarditis,we recently found that monoclonal antibodies with high purity and strong specificity can accurately act on target and achieve definite progress in the treatment of viral myocarditis in rat model and may meet the need above.However,several issues remain.The technology on howto make a higher homologous and weak immunogenic humanized or human source antibody and the treatment mechanism of monoclonal antibodies may provide solutions for these open issues.If we are to further stimulate

  3. Controlled delivery of antibodies from injectable hydrogels.

    Science.gov (United States)

    Fletcher, Nathan A; Babcock, Lyndsey R; Murray, Ellen A; Krebs, Melissa D

    2016-02-01

    Therapeutic antibodies are currently used for the treatment of various diseases, but large doses delivered systemically are typically required. Localized controlled delivery techniques would afford major benefits such as decreasing side effects and required doses. Injectable biopolymer systems are an attractive solution due to their minimally invasive potential for controlled release in a localized area. Here, alginate-chitosan hydrogels are demonstrated to provide controlled delivery of IgG model antibodies and also of Fab antibody fragments. Also, an alternate delivery system comprised of poly(lactic-co-glycolic acid) (PLGA) microspheres loaded with antibodies and encapsulated in alginate was shown to successfully provide another level of control over release. These biopolymer systems that offer controlled delivery for antibodies and antibody fragments will be promising for many applications in drug delivery and regenerative medicine.

  4. Factors determining antibody distribution in tumors

    Science.gov (United States)

    Thurber, Greg M.; Schmidt, Michael M.; Wittrup, K. Dane

    2009-01-01

    The development of antibody therapies for cancer is increasing rapidly, primarily owing to their specificity. Antibody distribution in tumors is often extremely uneven, however, leading to some malignant cells being exposed to saturating concentrations of antibody, whereas others are completely untargeted. This is detrimental because large regions of cells escape therapy, whereas other regions might be exposed to suboptimal concentrations that promote a selection of resistant mutants. The distribution of antibody depends on a variety of factors, including dose, affinity, antigens per cell and molecular size. Because these parameters are often known or easily estimated, a quick calculation based on simple modeling considerations can predict the uniformity of targeting within a tumor. Such analyses should enable experimental researchers to identify in a straightforward way the limitations in achieving evenly distributed antibody, and design and test improved antibody therapeutics more rationally. PMID:18179828

  5. Stabilization of antibody fragments in adverse environments.

    Science.gov (United States)

    Dooley, H; Grant, S D; Harris, W J; Porter, A J

    1998-08-01

    Antibody fragments have the potential to be used as sensitive and specific binding agents in a broad range of industrial applications. Genetic manipulation has been used to design a series of antibody fragment configurations with a flexible linker and/or a disulphide bond between the heavy chain and light chain of an antibody fragment against the herbicide atrazine. The thermostability and stability to a range of denaturants, polar and non-polar solvents, surfactants and proteases have been compared. It has been found that a novel antibody fragment construct (STAB: stabilized antibody) containing both a flexible linker and a disulphide bond can be effectively produced and shows greatly improved stability in these diverse environments. These STABs should be useful in environmental diagnostics and remediation, and may provide a generic approach for stabilizing antibody fragments in formulations containing detergents and penetrants for topical application in the pharmaceutical and cosmetic industries.

  6. Monoclonal antibodies as diagnostics; an appraisal

    Directory of Open Access Journals (Sweden)

    Siddiqui M

    2010-01-01

    Full Text Available Ever since the development of Hybridoma Technology in 1975 by Kohler and Milstein, our vision for antibodies as tools for research for prevention, detection and treatment of diseases, vaccine production, antigenic characterization of pathogens and in the study of genetic regulation of immune responses and disease susceptibility has been revolutionized. The monoclonal antibodies being directed against single epitopes are homogeneous, highly specific and can be produced in unlimited quantities. In animal disease diagnosis, they are very useful for identification and antigenic characterization of pathogens. Monoclonal antibodies have tremendous applications in the field of diagnostics, therapeutics and targeted drug delivery systems, not only for infectious diseases caused by bacteria, viruses and protozoa but also for cancer, metabolic and hormonal disorders. They are also used in the diagnosis of lymphoid and myeloid malignancies, tissue typing, enzyme linked immunosorbent assay, radio immunoassay, serotyping of microorganisms, immunological intervention with passive antibody, antiidiotype inhibition, or magic bullet therapy with cytotoxic agents coupled with anti mouse specific antibody. Recombinant deoxyribonucleic acid technology through genetic engineering has successfully led to the possibility of reconstruction of monoclonal antibodies viz. chimeric antibodies, humanized antibodies and complementarily determining region grafted antibodies and their enormous therapeutic use.

  7. Snake venom antibodies in Ecuadorian Indians.

    Science.gov (United States)

    Theakston, R D; Reid, H A; Larrick, J W; Kaplan, J; Yost, J A

    1981-10-01

    Serum samples from 223 Waorani Indians, a tribe in eastern Ecuador, were investigated by enzyme-linked immunosorbent assay for antibodies to snake venom. Seventy-eight per cent were positive, confirming the highest incidence and mortality from snake bite poisoning yet recorded in the world. Most samples were positive for more than one venom antibody. Antibodies were found to venoms of Bothrops viper in 60% of positive cases, of Micrurus coral snake in 21%, and of the bushmaster, Lachesis muta, in 18%. Further studies are needed to determine whether high venom-antibody levels afford protection against further snake envenoming. PMID:7299877

  8. Sequence and structual analysis of antibodies.

    OpenAIRE

    Raghavan, A. K.

    2008-01-01

    The work presented in this thesis focuses on the sequence and structural analysis of antibodies and has fallen into three main areas. First I developed a method to assess how typical an antibody sequence is of the expressed human antibody repertoire. My hypothesis was that the more "human like" an antibody sequence is (in other words how typical it is of the expressed human repertoire), the less likely it is to elicit an immune response when used in vivo in humans. In practice, I found that, ...

  9. Antiphospholipid Antibodies and Systemic Scleroderma

    Directory of Open Access Journals (Sweden)

    Awa Oumar Touré

    2013-03-01

    Full Text Available Objective: Antiphospholipid antibodies (APLs could be associated with an increased risk of vascular pathologies in systemic scleroderma. The aim of our study was to search for APLs in patients affected by systemic scleroderma and to evaluate their involvement in the clinical manifestations of this disease. Materials and Methods: We conducted a cross-sectional descriptive study, from January 2009 until August 2010, with patients received at the Department of Dermatology (Dakar, Senegal. Blood samples were taken at the hematology laboratory and were analyzed for the presence of APLs. Results: Forty patients were recruited. Various types of either isolated or associated APLs were found in 23 patients, i.e. 57.5% of the study population. The most frequently encountered antibody was IgG anti-β2 GPI (37.5% of the patients, followed by anticardiolipins (17.5% and lupus anticoagulants (5%. No statistically significant association of positive antiphospholipid-related tests to any of the scleroderma complications could be demonstrated. Conclusion: A high proportion of patients showing association of systemic scleroderma and APLs suggests the presence of a morbid correlation between these 2 pathologies. It would be useful to follow a cohort of patients affected by systemic scleroderma in order to monitor vascular complications following confirmation of the presence of antiphospholipid syndrome.

  10. Antibodies to human fetal erythroid cells from a nonimmune phage antibody library

    OpenAIRE

    Huie, Michael A.; Cheung, Mei-Chi; Muench, Marcus O.; Becerril, Baltazar; Kan, Yuet W.; Marks, James D.

    2001-01-01

    The ability to isolate fetal nucleated red blood cells (NRBCs) from the maternal circulation makes possible prenatal genetic analysis without the need for diagnostic procedures that are invasive for the fetus. Such isolation requires antibodies specific to fetal NRBCs. To generate a panel of antibodies to antigens present on fetal NRBCs, a new type of nonimmune phage antibody library was generated in which multiple copies of antibody fragments are displayed on each pha...

  11. High level transient production of recombinant antibodies and antibody fusion proteins in HEK293 cells

    OpenAIRE

    Jäger, Volker; Büssow, Konrad; Wagner, Andreas; Weber, Susanne; Hust, Michael; Frenzel, André; Schirrmann, Thomas

    2013-01-01

    Background The demand of monospecific high affinity binding reagents, particularly monoclonal antibodies, has been steadily increasing over the last years. Enhanced throughput of antibody generation has been addressed by optimizing in vitro selection using phage display which moved the major bottleneck to the production and purification of recombinant antibodies in an end-user friendly format. Single chain (sc)Fv antibody fragments require additional tags for detection and are not as suitable...

  12. The production of antibody fragments and antibody fusion proteins by yeasts and filamentous fungi

    NARCIS (Netherlands)

    Joosten, V.; Lokman, C.; Hondel, C.A.M.J.J. van den; Punt, P.J.

    2003-01-01

    In this review we will focus on the current status and views concerning the production of antibody fragments and antibody fusion proteins by yeasts and filamentous fungi. We will focus on single-chain antibody fragment production (scFv and VHH) by these lower eukaryotes and the possible applications

  13. Anti-influenza M2e antibody

    Energy Technology Data Exchange (ETDEWEB)

    Bradbury, Andrew M.

    2013-04-16

    Humanized recombinant and monoclonal antibodies specific for the ectodomain of the influenza virus M2 ion channel protein are disclosed. The antibodies of the invention have anti-viral activity and may be useful as anti-viral therapeutics and/or prophylactic/vaccine agents for inhibiting influenza virus replication and for treating individuals infected with influenza.

  14. Anti-influenza M2e antibody

    Energy Technology Data Exchange (ETDEWEB)

    Bradbury, Andrew M. (Santa Fe, NM)

    2011-12-20

    Humanized recombinant and monoclonal antibodies specific for the ectodomain of the influenza virus M2 ion channel protein are disclosed. The antibodies of the invention have anti-viral activity and may be useful as anti-viral therapeutics and/or prophylactic/vaccine agents for inhibiting influenza virus replication and for treating individuals infected with influenza.

  15. "Unconventional" Neutralizing Activity of Antibodies Against HIV

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Neutralizing antibodies are recognized to be one of the essential elements of the adaptive immune response that must be induced by an effective vaccine against HIV. However, only a limited number of antibodies have been identified to neutralize a broad range of primary isolates of HIV-1 and attempts to induce such antibodies by immunization were unsuccessful. The difficulties to generate such antibodies are mainly due to intrinsic properties of HIV-1 envelope spikes, such as high sequence diversity, heavy glycosylation, and inducible and transient nature of certain epitopes. In vitro neutralizing antibodies are identified using "conventional" neutralization assay which uses phytohemagglutinin (PHA)-stimulated human PBMCs as target cells. Thus, in essence the assay evaluates HIV-1 replication in CD4+ T cells. Recently, several laboratories including us demonstrated that some monoclonal antibodies and HIV-1-specific polyclonal IgG purified from patient sera, although they do not have neutralizing activity when tested by the "conventional" neutralization assay, do exhibit potent and broad neutralizing activity in "unconventional" ways. The neutralizing activity of these antibodies and IgG fractions is acquired through post-translational modifications, through opsonization of virus particles into macrophages and immature dendritic cells (iDCs), or through expression of antibodies on the surface of HIV-1-susceptible cells. This review will focus on recent findings of this area and point out their potential applications in the development of preventive strategies against HIV.

  16. Photonic crystal fiber based antibody detection

    DEFF Research Database (Denmark)

    Duval, A; Lhoutellier, M; Jensen, J B;

    2004-01-01

    An original approach for detecting labeled antibodies based on strong penetration photonic crystal fibers is introduced. The target antibody is immobilized inside the air-holes of a photonic crystal fiber and the detection is realized by the means of evanescent-wave fluorescence spectroscopy...... and the use of a transversal illumination setup....

  17. Monoclonal antibodies reactive with hairy cell leukemia

    NARCIS (Netherlands)

    Visser, L; Shaw, A; Slupsky, J; Vos, H; Poppema, S

    1989-01-01

    Monoclonal antibodies reactive with hairy cell leukemia were developed to aid in the diagnosis of this subtype of B cell chronic lymphocytic leukemia and to gain better insight into the origin of hairy cells. Three antibodies were found to be of value in the diagnosis of hairy cell leukemia. Antibod

  18. Synthetic Antibodies for Reversible Cell Recognition

    Science.gov (United States)

    Zhou, Jing Zhou

    2011-12-01

    Antibody-mediated cell recognition plays a critical role in various biological and biomedical applications. However, strong antibody-cell interactions can lead to the difficulty of separating antibodies from the bound cells in a simple and non-destructive manner, which is often necessary to numerous applications such as cell sorting or separation. Thus, this thesis research is aimed to create an antibody-like nanomaterial with the function of reversible cell recognition It was hypothesized that nucleic acid aptamer and dendrimer could be used as fundamental structural components to develop an antibody-like nanomaterial. The aptamer functions as the binding site of an antibody; the dendrimer is used as a robust, defined nano-scaffold to support the aptamer and to carry small molecules (e.g., fluorophores). To test this hypothesis, a novel method was first developed to discover the essential nucleotides of full-length aptamers to mimic the binding sites of antibodies. The essential nucleotides were further conjugated with a dendrimer to synthesize a monovalent aptamer-dendrimer nanomaterial. The results clearly showed that the essential nucleotides could maintain high affinity and specificity after tethered on dendrimer surface. To further test the hypothesis that antibody-like nanomaterials can be rationally designed to acquire the capability of reversible cell recognition, an aptamer that was selected at 0 °C was used as a model to synthesize a "Y-shaped" nanomaterial by conjugating two aptamers to the same dendrimer. The results showed that the nanomaterial-cell interaction could be affected by the distance between two binding aptamers. In addition, the "Y-shaped" antibody-like nanomaterial could bind target cells more strongly than its monovalent control. Importantly, the strong cell-nanomaterial interaction could be rapidly reversed when the temperature was shifted from 0 °C to 37 °C. In summary, we developed a synthetic antibody that can not only mimic the

  19. Trends in Malignant Glioma Monoclonal Antibody Therapy

    Science.gov (United States)

    Chekhonin, Ivan; Gurina, Olga

    2015-01-01

    Although new passive and active immunotherapy methods are emerging, unconjugated monoclonal antibodies remain the only kind of biological preparations approved for high-grade glioma therapy in clinical practice. In this review, we combine clinical and experimental data discussion. As antiangiogenic therapy is the standard of care for recurrent glioblastoma multiforme (GBM), we analyze major clinical trials and possible therapeutic combinations of bevacizumab, the most common monoclonal antibody to vascular endothelial growth factor (VEGF). Another humanized antibody to gain recognition in GBM is epidermal growth factor (EGFR) antagonist nimotuzumab. Other antigens (VEGF receptor, platelet-derived growth factor receptor, hepatocyte growth factor and c-Met system) showed significance in gliomas and were used to create monoclonal antibodies applied in different malignant tumors. We assess the role of genetic markers (isocitrate dehydrogenase, O6-methylguanine-DNA methyltransnsferase) in GBM treatment outcome prediction. Besides antibodies studied in clinical trials, we focus on perspective targets and briefly list other means of passive immunotherapy.

  20. Antiphospholipid antibody: laboratory, pathogenesis and clinical manifestations

    Directory of Open Access Journals (Sweden)

    T. Ziglioli

    2011-06-01

    Full Text Available Antiphospholipid antibodies (aPL represent a heterogeneous group of antibodies that recognize various antigenic targets including beta2 glycoprotein I (β2GPI, prothrombin (PT, activated protein C, tissue plasminogen activator, plasmin and annexin A2. The most commonly used tests to detect aPL are: lupus anticoagulant (LAC, a functional coagulation assay, anticardiolipin antibody (aCL and anti-β2GPI antibody (anti-β2GPI, which are enzyme-linked immunoassay (ELISA. Clinically aPL are associated with thrombosis and/or with pregnancy morbidity. Apparently aPL alone are unable to induce thrombotic manifestations, but they increase the risk of vascular events that can occur in the presence of another thrombophilic condition; on the other hand obstetrical manifestations were shown to be associated not only to thrombosis but mainly to a direct antibody effect on the trophoblast.

  1. Vector-Mediated In Vivo Antibody Expression.

    Science.gov (United States)

    Schnepp, Bruce C; Johnson, Philip R

    2014-08-01

    This article focuses on a novel vaccine strategy known as vector-mediated antibody gene transfer, with a particular focus on human immunodeficiency virus (HIV). This strategy provides a solution to the problem of current vaccines that fail to generate neutralizing antibodies to prevent HIV-1 infection and AIDS. Antibody gene transfer allows for predetermination of antibody affinity and specificity prior to "immunization" and avoids the need for an active humoral immune response against the HIV envelope protein. This approach uses recombinant adeno-associated viral (rAAV) vectors, which have been shown to transduce muscle with high efficiency and direct the long-term expression of a variety of transgenes, to deliver the gene encoding a broadly neutralizing antibody into the muscle. Following rAAV vector gene delivery, the broadly neutralizing antibodies are endogenously synthesized in myofibers and passively distributed to the circulatory system. This is an improvement over classical passive immunization strategies that administer antibody proteins to the host to provide protection from infection. Vector-mediated gene transfer studies in mice and monkeys with anti-HIV and simian immunodeficiency virus (SIV)-neutralizing antibodies demonstrated long-lasting neutralizing activity in serum with complete protection against intravenous challenge with virulent HIV and SIV. These results indicate that existing potent anti-HIV antibodies can be rapidly moved into the clinic. However, this methodology need not be confined to HIV. The general strategy of vector-mediated antibody gene transfer can be applied to other difficult vaccine targets such as hepatitis C virus, malaria, respiratory syncytial virus, and tuberculosis. PMID:26104192

  2. Immobilization of antibodies and enzyme-labeled antibodies by radiation polymerization

    International Nuclear Information System (INIS)

    Immobilization of antibodies and enzyme-labeled antibodies by radiation polymerization at low temperatures was studied. The antibody activity of antibody was not affected by irradiation at an irradiation dose of below 8 MR and low temperatures. Immobilization of peroxidase-labeled anti-rabbit IgG goat IgG, anti-peroxidase, peroxidase, and anti-alpha-fetoprotein was carried out with hydrophilic and hydrophobic monomers. The activity of the immobilized enzyme-labeled antibody membranes varied with the thickness of the membranes and increased with decreasing membrane thickness. The activity of the immobilized antibody particles was varied by particle size. Immobilized anti-alpha-fetoprotein particles and membranes can be used for the assay of alpha-fetoprotein by the antigen-antibody reaction, such as a solid-phase sandwich method with high sensitivity

  3. Radiohalogenated half-antibodies and maleimide intermediate therefor

    Science.gov (United States)

    Kassis, A.I.; Khawli, L.A.

    1991-02-19

    N-(m-radiohalophenyl) maleimide can be conjugated with a reduced antibody having a mercapto group to provide a radiolabeled half-antibody having immunological specific binding characteristics of whole antibody. No Drawings

  4. 抗H5N1禽流感病毒羊驼重链单域抗体的制备和功能鉴定%Preparation and characterization of recombinant VHH antibody against H5N1 hemagglutinin from avian influenza virus

    Institute of Scientific and Technical Information of China (English)

    夏立亮; 吴标; 程亚庭; 包文静; 赵国屏; 王颖

    2012-01-01

    目的:获得具有中和活性、高特异性和稳定性的抗H5N1禽流感病毒血凝素蛋白(HA)的羊驼重链单域(VHH)抗体.方法:利用pET-22b表达载体诱导表达抗H5N1禽流感病毒HA VHH抗体蛋白,以包涵体形式表达的VHH抗体蛋白采用最优复性方法进行复性后,获得高纯度的VHH抗体,分别采用ELISA法鉴定VHH抗体的亲和力和热稳定性,采用血凝抑制实验鉴定抗体的特异性和体外中和活性.结果:经复性的抗H5N1禽流感HA VHH抗体对H5N1禽流感病毒HA具有良好的特异性.通过对三种不同复性方法比较,利用柱上复性的VHH23抗体具有较好的热稳定性,亲和力为9.1×10-7 mol/L,同时对H5N1禽流感病毒HA具有良好的体外中和活性.结论:实验结果表明通过原核表达获得具有较好中和活性、特异性及稳定性的抗H5N1禽流感病毒VHH抗体,为进一步开展抗体的体内病毒中和试验奠定良好基础.%Objective; To prepare and characterize camelid VHH antibody against hemagglutinin from H5N1 avian influenza virus. Methods: Recombinant expression vector pET-22b-VHH23 was constructed and VHH antibody against hemagglutinin from H5N1 a-vian influenza virus was expressed in E. Coli BL2l( DE3 ) strain by IPTG induction. As VHH antibody protein was expressed in inclusion body,there different refolding methods were compared for the refolding of VHH antibody in which the optimal methods were adopted for preparation of VHH antibody. The activity and thermal stability of VHH antibodies were tested by ELISA while hemagglutination-inhibition assay was perfomed to determine the specificity and neutralizing activity. Results; By using on-column refolding procedure, VHH antibody has been obtained with highest yield and good quality. This antibody specifically recognized hemagglutinin derived from H5N1 avian influenza virus as well as in vitro inhibition against hemagglutinin-mediated red blood cell agglutination. The affinity constant of VHH

  5. Utilización de los test de Fluorescencia Polarizada (FP y Elisa de Competencia (C-Elisa en el diagnóstico de brucelosis de camélidos The use of polarized fluorescence assay (PF and competitive ELISA test (C-Elisa for the diagnosis of Brucellosis in South American camelids

    Directory of Open Access Journals (Sweden)

    X Rojas

    2004-01-01

    Full Text Available En Chile, como en muchos países de Latinoamérica, la brucelosis es una zoonosis presente, representando no sólo un factor de riesgo para las explotaciones pecuarias sino también para quienes laboran y hacen usufructo de ella, especialmente por el consumo de leche cruda y sus derivados. Como la Brucella afecta a diferentes especies de animales, el objetivo de este trabajo fue utilizar las técnicas de unión primaria, como son Fluorescencia Polarizada (FP y Elisa de Competencia (C-Elisa, para la detección de anticuerpos anti-Brucella en suero de camélidos y, comparar sus respuestas con las pruebas de Rosa de Bengala (RB, Aglutinación Lenta en Tubo (SAT y Fijación de Complemento (FC en atención a que estas especies representan un potencial pecuario de cierta trascendencia. Para ello se analizaron 336 sueros, de los cuales 315 pertenecían a llamas y 21 a alpacas, provenientes de un predio de la Novena Región clínica y serológicamente libre de Brucella. Los resultados obtenidos señalan que el umbral de positividad para FP, calculado según la mediana del último percentil, fue de 98.7 milipolarizaciones (mP y, el corte para C-Elisa con el mismo método fue de 32% de Inhibición (% I. Todas las muestras analizadas dieron resultados negativos a RB y SAT. Los resultados con FC no pudieron ser considerados, por cuanto un alto porcentaje de las muestras fueron anticomplementarias. Con las técnicas de C-Elisa y FP, dos muestras reaccionaron positivamente en cada uno de los test con valores en el nivel de corte o levemente superiores. Se discute la significación de ellas.Brucellosis is a zoonotic disease representing a risk contact with, or consuming, their raw milk and raw dairy several animal species, including camelids, the aim of this Polarization assay (PF and Competitive ELISA (C-ELISA, along with conventional serological tests such as the Rose Complement Fixation test (CF. A total of 336 serum samples herd of the IXth Region (Chile

  6. Antibody Engineering & Therapeutics, the annual meeting of The Antibody Society December 7-10, 2015, San Diego, CA, USA.

    Science.gov (United States)

    Pauthner, Matthias; Yeung, Jenny; Ullman, Chris; Bakker, Joost; Wurch, Thierry; Reichert, Janice M; Lund-Johansen, Fridtjof; Bradbury, Andrew R M; Carter, Paul J; Melis, Joost P M

    2016-01-01

    The 26th Antibody Engineering & Therapeutics meeting, the annual meeting of The Antibody Society united over 800 participants from all over the world in San Diego from 6-10 December 2015. The latest innovations and advances in antibody research and development were discussed, covering a myriad of antibody-related topics by more than 100 speakers, who were carefully selected by The Antibody Society. As a prelude, attendees could join the pre-conference training course focusing, among others, on the engineering and enhancement of antibodies and antibody-like scaffolds, bispecific antibody engineering and adaptation to generate chimeric antigen receptor constructs. The main event covered 4 d of scientific sessions that included antibody effector functions, reproducibility of research and diagnostic antibodies, new developments in antibody-drug conjugates (ADCs), preclinical and clinical ADC data, new technologies and applications for bispecific antibodies, antibody therapeutics for non-cancer and orphan indications, antibodies to harness the cellular immune system, building comprehensive IgVH-gene repertoires through discovering, confirming and cataloging new germline IgVH genes, and overcoming resistance to clinical immunotherapy. The Antibody Society's special session focused on "Antibodies to watch" in 2016. Another special session put the spotlight on the limitations of the new definitions for the assignment of antibody international nonproprietary names introduced by the World Health Organization. The convention concluded with workshops on computational antibody design and on the promise and challenges of using next-generation sequencing for antibody discovery and engineering from synthetic and in vivo libraries. PMID:26909869

  7. Utilisation of antibody microarrays for the selection of specific and informative antibodies from recombinant library binders of unknown quality

    DEFF Research Database (Denmark)

    Kibat, Janek; Schirrmann, Thomas; Knape, Matthias J;

    2016-01-01

    Many diagnostic and therapeutic concepts require antibodies of high specificity. Recombinant binder libraries and related selection approaches allow the efficient isolation of antibodies against almost every target of interest. Nevertheless, it cannot be guaranteed that selected antibodies perfor...

  8. Combinatorial antibody libraries: new advances, new immunological insights.

    Science.gov (United States)

    Lerner, Richard A

    2016-08-01

    Immunochemists have become quite proficient in engineering existing antibody molecules to control their pharmacological properties. However, in terms of generating new antibodies, the combinatorial antibody library has become a central feature of modern immunochemistry. These libraries are essentially an immune system in a test tube and enable the selection of antibodies without the constraints of whole animal or cell-based systems. This Review provides an overview of how antibody libraries are constructed and discusses what can be learnt from these synthetic systems. In particular, the Review focuses on new biological insights from antibody libraries - such as the concept of 'SOS antibodies' - and the growing use of intracellular antibodies to perturb cellular functions.

  9. Antiphospholipid Antibodies in Lupus Nephritis.

    Science.gov (United States)

    Parodis, Ioannis; Arnaud, Laurent; Gerhardsson, Jakob; Zickert, Agneta; Sundelin, Birgitta; Malmström, Vivianne; Svenungsson, Elisabet; Gunnarsson, Iva

    2016-01-01

    Lupus nephritis (LN) is a major manifestation of systemic lupus erythematosus (SLE). It remains unclear whether antiphospholipid antibodies (aPL) alter the course of LN. We thus investigated the impact of aPL on short-term and long-term renal outcomes in patients with LN. We assessed levels of aPL cross-sectionally in SLE patients diagnosed with (n = 204) or without (n = 294) LN, and prospectively in 64 patients with active biopsy-proven LN (52 proliferative, 12 membranous), before and after induction treatment (short-term outcomes). Long-term renal outcome in the prospective LN cohort was determined by the estimated glomerular filtration rate (eGFR) and the Chronic Kidney Disease (CKD) stage, after a median follow-up of 11.3 years (range: 3.3-18.8). Cross-sectional analysis revealed no association between LN and IgG/IgM anticardiolipin or anti-β2-glycoprotein I antibodies, or lupus anticoagulant. Both aPL positivity and levels were similar in patients with active LN and non-renal SLE. Following induction treatment for LN, serum IgG/IgM aPL levels decreased in responders (p<0.005 for all), but not in non-responders. Both at active LN and post-treatment, patients with IgG, but not IgM, aPL had higher creatinine levels compared with patients without IgG aPL. Neither aPL positivity nor levels were associated with changes in eGFR from either baseline or post-treatment through long-term follow-up. Moreover, aPL positivity and levels both at baseline and post-treatment were similar in patients with a CKD stage ≥3 versus 1-2 at the last follow-up. In conclusion, neither aPL positivity nor levels were found to be associated with the occurrence of LN in SLE patients. However, IgG aPL positivity in LN patients was associated with a short-term impairment of the renal function while no effect on long-term renal outcome was observed. Furthermore, IgG and IgM aPL levels decreased following induction treatment only in responders, indicating that aPL levels are affected by

  10. Antiphospholipid Antibodies in Lupus Nephritis.

    Directory of Open Access Journals (Sweden)

    Ioannis Parodis

    Full Text Available Lupus nephritis (LN is a major manifestation of systemic lupus erythematosus (SLE. It remains unclear whether antiphospholipid antibodies (aPL alter the course of LN. We thus investigated the impact of aPL on short-term and long-term renal outcomes in patients with LN. We assessed levels of aPL cross-sectionally in SLE patients diagnosed with (n = 204 or without (n = 294 LN, and prospectively in 64 patients with active biopsy-proven LN (52 proliferative, 12 membranous, before and after induction treatment (short-term outcomes. Long-term renal outcome in the prospective LN cohort was determined by the estimated glomerular filtration rate (eGFR and the Chronic Kidney Disease (CKD stage, after a median follow-up of 11.3 years (range: 3.3-18.8. Cross-sectional analysis revealed no association between LN and IgG/IgM anticardiolipin or anti-β2-glycoprotein I antibodies, or lupus anticoagulant. Both aPL positivity and levels were similar in patients with active LN and non-renal SLE. Following induction treatment for LN, serum IgG/IgM aPL levels decreased in responders (p<0.005 for all, but not in non-responders. Both at active LN and post-treatment, patients with IgG, but not IgM, aPL had higher creatinine levels compared with patients without IgG aPL. Neither aPL positivity nor levels were associated with changes in eGFR from either baseline or post-treatment through long-term follow-up. Moreover, aPL positivity and levels both at baseline and post-treatment were similar in patients with a CKD stage ≥3 versus 1-2 at the last follow-up. In conclusion, neither aPL positivity nor levels were found to be associated with the occurrence of LN in SLE patients. However, IgG aPL positivity in LN patients was associated with a short-term impairment of the renal function while no effect on long-term renal outcome was observed. Furthermore, IgG and IgM aPL levels decreased following induction treatment only in responders, indicating that aPL levels are

  11. Structure Based Antibody-Like Peptidomimetics

    Directory of Open Access Journals (Sweden)

    Mark I. Greene

    2012-02-01

    Full Text Available Biologics such as monoclonal antibodies (mAb and soluble receptors represent new classes of therapeutic agents for treatment of several diseases. High affinity and high specificity biologics can be utilized for variety of clinical purposes. Monoclonal antibodies have been used as diagnostic agents when coupled with radionuclide, immune modulatory agents or in the treatment of cancers. Among other limitations of using large molecules for therapy the actual cost of biologics has become an issue. There is an effort among chemists and biologists to reduce the size of biologics which includes monoclonal antibodies and receptors without a reduction of biological efficacy. Single chain antibody, camel antibodies, Fv fragments are examples of this type of deconstructive process. Small high-affinity peptides have been identified using phage screening. Our laboratory used a structure-based approach to develop small-size peptidomimetics from the three-dimensional structure of proteins with immunoglobulin folds as exemplified by CD4 and antibodies. Peptides derived either from the receptor or their cognate ligand mimics the functions of the parental macromolecule. These constrained peptides not only provide a platform for developing small molecule drugs, but also provide insight into the atomic features of protein-protein interactions. A general overview of the reduction of monoclonal antibodies to small exocyclic peptide and its prospects as a useful diagnostic and as a drug in the treatment of cancer are discussed.

  12. Next generation of antibody therapy for cancer

    Institute of Scientific and Technical Information of China (English)

    Zhenping Zhu; Li Yan

    2011-01-01

    Monoclonal antibodies (mAbs) have become a major class of therapeutic agents providing effective altematives to treating various human diseases. To date, 15 mAbs have been approved by regulatory agencies in the world for clinical use in oncology indications. The selectivity and specificity, the unique pharmacokinetics, and the ability to engage and activate the host immune system differentiate these biologics from traditional small molecule anticancer drugs. mAb-basod regimens have brought clinical benefits, including improvements in overall survival, to patients with a variety of cancers. Many challenges still remain, however, to fully realize the potential of these new medicines. With our further understanding of cancer biology, mechanism of antibody action, and advancement of antibody engineering technologies, many novel antibody formats or antibody-derived molecules are emerging as promising new generation therapeutics. Carefully designed and engineered, they retain the advantage of specificity and selectivity of original antibodies, but in the meantime acquire additional special features such as improved pharmacokinetics, increased selectivity, and enhanced anticancer efficacy. Promising clinical results are being generated with these newly improved antibody-based therapeutics.

  13. Baculovirus display of functional antibody Fab fragments.

    Science.gov (United States)

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

    2015-08-01

    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.

  14. Studies on Purification of Methamidophos Monoclonal Antibodies and Comoarative Immunoactivity of Purified Antibodies

    Institute of Scientific and Technical Information of China (English)

    SU-QING ZHAO; YUAN-MING SUN; CHUN-YAN ZHANG; XIAO-YU HUANG; HOU-RUI ZHANG; ZHEN-YU ZHU

    2003-01-01

    Objective To purify Methamidophos (Met) monoclonal antibodies with two methods andcompare immune activity of purified antibodies. Method Caprylic acid ammonium sulphateprecipition (CAASP) method and Sepharose protein-A (SPA) affinity chromatography method wereused to purify Met monoclonal antibodies, UV spectrum scanning was used to determine proteincontent and recovery of purified antibodies, sodium dodecylsulphate polyacrylamide gelelectrophoresis (SDS-PAGE) was used to analyze the purity of purified antibodies, and enzyme-linkedimmunosorbent assay (ELISA) was used to determine immune activity of purified antibodies.Results Antibody protein content and recovery rate with CAASP method were 7.62 mg/mL and8.05% respectively, antibody protein content and recovery rate with SPA method were 6.45 mg/mLand 5.52% respectively. Purity of antibodies purified by SPA method was higher than that by CAASPmethod. The half-maximal inhibition concentration (IC50) of antibodies purified by SPA to Met was181.26 μg/mL, and the linear working range and the limit of quantification (LOD) were 2.43-3896.01μg/mL and 1.03 μg/mL, respectively. The IC50 of antibodies purified by CAASP to Met was 352.82μg/mL, and the linear working range and LOD were 10.91-11412.29 ug/mL and 3.42 μg/mL,respectively. Conclusion Antibodies purified by SPA method are better than those by CAASPmethod, and Met monoclonal antibodies purified by SPA method can be used to prepare gold-labelledtesting paper for analyzing Met residue in vegetable and drink water.

  15. Immunocytochemical and Immunohistochemical Staining with Peptide Antibodies.

    Science.gov (United States)

    Friis, Tina; Pedersen, Klaus Boberg; Hougaard, David; Houen, Gunnar

    2015-01-01

    Peptide antibodies are particularly useful for immunocytochemistry (ICC) and immunohistochemistry (IHC), where antigens may denature due to fixation of tissues and cells. Peptide antibodies can be made to any defined sequence, including unknown putative proteins and posttranslationally modified sequences. Moreover, the availability of large amounts of the antigen (peptide) allows inhibition/adsorption controls, which are important in ICC/IHC, due to the many possibilities for false-positive reactions caused by immunoglobulin Fc receptors, nonspecific reactions, and cross-reactivity of primary and secondary antibodies with other antigens and endogenous immunoglobulins, respectively. Here, simple protocols for ICC and IHC are described together with recommendations for appropriate controls.

  16. The antibody approach of labeling blood cells

    Energy Technology Data Exchange (ETDEWEB)

    Srivastava, S.C.

    1992-12-31

    Although the science of blood cell labeling using monoclonal antibodies directed against specific cellular antigens is still in its early stages, considerable progress has recently been accomplished in this area. The monoclonal antibody approach offers the promise of greater selectivity and enhanced convenience since specific cell types can be labeled in vivo, thus eliminating the need for complex and damaging cell separation procedures. This article focuses on these developments with primary emphasis on antibody labeling of platelets and leukocytes. The advantages and the shortcomings of the recently reported techniques are critically assessed and evaluated.

  17. The antibody approach of labeling blood cells

    International Nuclear Information System (INIS)

    Although the science of blood cell labeling using monoclonal antibodies directed against specific cellular antigens is still in its early stages, considerable progress has recently been accomplished in this area. The monoclonal antibody approach offers the promise of greater selectivity and enhanced convenience since specific cell types can be labeled in vivo, thus eliminating the need for complex and damaging cell separation procedures. This article focuses on these developments with primary emphasis on antibody labeling of platelets and leukocytes. The advantages and the shortcomings of the recently reported techniques are critically assessed and evaluated

  18. Reshaping Human Antibodies: Grafting an Antilysozyme Activity

    Science.gov (United States)

    Verhoeyen, Martine; Milstein, Cesar; Winter, Greg

    1988-03-01

    The production of therapeutic human monoclonal antibodies by hybridoma technology has proved difficult, and this has prompted the ``humanizing'' of mouse monoclonal antibodies by recombinant DNA techniques. It was shown previously that the binding site for a small hapten could be grafted from the heavy-chain variable domain of a mouse antibody to that of a human myeloma protein by transplanting the hypervariable loops. It is now shown that a large binding site for a protein antigen (lysozyme) can also be transplanted from mouse to human heavy chain. The success of such constructions may be facilitated by an induced-fit mechanism.

  19. The antibody approach of labeling blood cells

    Energy Technology Data Exchange (ETDEWEB)

    Srivastava, S.C.

    1991-01-01

    Although the science of blood cell labeling using monoclonal antibodies directed against specific cellular antigens is still in its early stages, considerable progress has recently been accomplished in this area. The monoclonal antibody approach offers the promise of greater selectivity and enhanced convenience since specific cell types can be labeled in vivo, thus eliminating the need for complex and damaging cell separation procedures. This article focuses on these developments with primary emphasis on antibody labeling of platelets and leukocytes. The advantages and the shortcomings of the recently reported techniques are criticality assessed and evaluated.

  20. The antibody approach of labeling blood cells

    Energy Technology Data Exchange (ETDEWEB)

    Srivastava, S.C.

    1991-12-31

    Although the science of blood cell labeling using monoclonal antibodies directed against specific cellular antigens is still in its early stages, considerable progress has recently been accomplished in this area. The monoclonal antibody approach offers the promise of greater selectivity and enhanced convenience since specific cell types can be labeled in vivo, thus eliminating the need for complex and damaging cell separation procedures. This article focuses on these developments with primary emphasis on antibody labeling of platelets and leukocytes. The advantages and the shortcomings of the recently reported techniques are criticality assessed and evaluated.

  1. Radioimmunological proof of thyroglobulin antibodies in humans by the use of a double antibody method

    International Nuclear Information System (INIS)

    Thyroid antibodies, especially thyroglobulin antibodies, allow themselves to be proven with the double antibody method, in competitive radio binding assays and with the solid phase technique. These methods offer advantages relative to sensitivity and quantifiability. In this work a sensitive radioimmunoassay as a double antibody method was worked out whereby a 125 I-thyroglobulin/thyroglobulin antibody immune complex was precipitated out using anti-human immunoglobulin. The measured results from the radioimmunoassay show a good correlation with the results of the immune histological findings. A high to very high Tg antibody level occurs with autoimmune thyroiditis (80%), primary hypothyroidism (74%) and hyperthyroidism (70%). The control values with healthy people came to less than 5% specific binding. In correlation with the results of other authors this method is advantageous relative to test start and evaluation procedures. (orig.)

  2. Chemical biology: How to minimalize antibodies

    Science.gov (United States)

    Rader, Christoph

    2015-02-01

    The success of antibodies as pharmaceuticals has triggered interest in crafting much smaller mimics. A crucial step forward has been taken with the chemical synthesis of small molecules that recruit immune cells to attack cancer cells.

  3. Patient-Derived Antibody Targets Tumor Cells

    Science.gov (United States)

    An NCI Cancer Currents blog on an antibody derived from patients that killed tumor cells in cell lines of several cancer types and slowed tumor growth in mouse models of brain and lung cancer without evidence of side effects.

  4. Localization of tumors by radiolabelled antibodies

    International Nuclear Information System (INIS)

    A method of utilizing radiolabelled antibodies to carcinoembryonic antigens for determining the site of tumors which produce or are associated with carcinoembryonic antigen is disclosed. 3 claims, no drawings

  5. Quantitative Assessment of Antibody Internalization with Novel Monoclonal Antibodies against Alexa Fluorophores

    OpenAIRE

    Liao-Chan, Sindy; Daine-Matsuoka, Barbara; Heald, Nathan; Wong, Tiffany; Lin, Tracey; Cai, Allen G.; Lai, Michelle; D’Alessio, Joseph A.; Theunissen, Jan-Willem

    2015-01-01

    Antibodies against cell surface antigens may be internalized through their specific interactions with these proteins and in some cases may induce or perturb antigen internalization. The anti-cancer efficacy of antibody-drug conjugates is thought to rely on their uptake by cancer cells expressing the surface antigen. Numerous techniques, including microscopy and flow cytometry, have been used to identify antibodies with desired cellular uptake rates. To enable quantitative measurements of inte...

  6. Influenza-Specific Antibody-Dependent Phagocytosis

    OpenAIRE

    Ana-Sosa-Batiz, Fernanda; Vanderven, Hillary; Jegaskanda, Sinthujan; Johnston, Angus; Rockman, Steven; Laurie, Karen; Barr, Ian; Reading, Patrick; Lichtfuss, Marit; Stephen J Kent

    2016-01-01

    Background Immunity to human influenza A virus (IAV) infection is only partially understood. Broadly non-neutralizing antibodies may assist in reducing disease but have not been well characterized. Methods We measured internalization of opsonized, influenza protein-coated fluorescent beads and live IAV into a monocytic cell line to study antibody-dependent phagocytosis (ADP) against multiple influenza hemagglutinin (HA) subtypes. We analyzed influenza HA-specific ADP in healthy human donors, ...

  7. Single-domain antibodies for brain targeting

    OpenAIRE

    Lalatsa, Katerina; Moreira Leite, Diana

    2014-01-01

    Smaller recombinant antibody fragments as single-domain antibodies (sdAbs) are emerging as credible alternatives because of their target specificity, high affinity, and cost-effective recombinant production. sdAbs have been forged into multivalent and multispecif ic therapeutics, or targeting moieties, that are able to shuttle their linked therapeutic cargo (i.e., drugs, nanoparticles, toxins, enzymes, and radionuclides) to the receptor of interest. Their ability to permeate across the blood ...

  8. Therapeutic monoclonal antibody for Sporotrichosis

    Directory of Open Access Journals (Sweden)

    Sandro eAlmeida

    2012-11-01

    Full Text Available Sporotrichosis is a chronic subcutaneous mycosis that affects either humans or animals and occurs worldwide. This subcutaneous mycosis had been attributed to a single etiological agent, Sporothrix schenckii. S. schenckii exhibits a considerable genetic variability, where recently, was suggesting that this taxon consists of a complex of species. Sporotrichosis is caused by traumatic inoculation of the fungus, which is a ubiquitous environmental saprophyte that can be isolated from soil and plant debris. The infection is limited to the cutaneous forms but, recently, occurrences of more severe clinical forms of this mycosis were described, especially among immunocompromized individuals. The immunological mechanisms involved in prevention and control of sporotrichosis are still not very well understood. Some works suggest that cell-mediated immunity plays an important role in protecting the host against S. schenckii. In contrast, the role of the humoral immune response in protection against this fungus have not been studied in detail. In a previous study, we showed that antigens secreted by S. schenckii induce a specific humoral response in infected animals, mainly against the 70-kDa molecules, indicating a possible participation of specific antibodies to this molecule in infection control. In an other work of the our group, we produced a mAb against a 70-kDa glycoprotein of S. schenckii in order to better understand the effect of passive immunization of mice infected with S. schenckii. Results showed a significant reduction in the number of CFU in organs of mice when the mAb was injected before and during S. schenckii infection. Similar results were observed when T-cell deficient mice were used. Drugs of choice in the treatment of sporothrichosis require long periods and frequently relapses are observed, mainly in immunocompromized patients. The strong protection induced by mAb against a 70-kDa glycoprotein makes it a strong candidate for a

  9. IMPORTANCE OF RESEARCH HLA ANTIBODIES CLASS I AND II, AND MICA ANTIBODIES IN KIDNEY TRANSPLANTATION

    Directory of Open Access Journals (Sweden)

    M. Sh. Khubutia

    2011-01-01

    Full Text Available The purpose of this study was to investigate the occurrence of HLA and MICA antibodies in patients from the waiting list for kidney transplantation and their influence on the course of post-transplant period. Determination of HLA antibodies class I and II, and MICA antibodies was performed on a platform of Luminex (xMAP-tech- nology using sets LABScreen ONE LAMBDA (U.S.. A total of 156 patients from the waiting list for kidney transplantation. Revealed the presence of HLA and MICA antibodies in the serum of 31.4% of patients. Regraf- ted patients increased the content of antibodies to the antigens of HLA system was noted in 88.2% of cases, 47% met the combination of antibodies to the I, II classes and MICA. In patients awaiting first kidney transplantation, HLA and MICA antibodies were determined in 23.7% of cases. The presence of pretransplant HLA and MICA antibodies had a significant influence on the course of post-transplant period. Patients with the presence of HLA and MICA in 50% of cases delayed graft function. Sessions of plasmapheresis can reduce the concentration of HLA and MICA antibodies on average by 61.1%. 

  10. Antibody-Conjugated Nanoparticles for Biomedical Applications

    Directory of Open Access Journals (Sweden)

    Manuel Arruebo

    2009-01-01

    Full Text Available Nanoscience and Nanotechnology have found their way into the fields of Biotechnology and Medicine. Nanoparticles by themselves offer specific physicochemical properties that they do not exhibit in bulk form, where materials show constant physical properties regardless of size. Antibodies are nanosize biological products that are part of the specific immune system. In addition to their own properties as pathogens or toxin neutralizers, as well as in the recruitment of immune elements (complement, improving phagocytosis, cytotoxicity antibody dependent by natural killer cells, etc., they could carry several elements (toxins, drugs, fluorochroms, or even nanoparticles, etc. and be used in several diagnostic procedures, or even in therapy to destroy a specific target. The conjugation of antibodies to nanoparticles can generate a product that combines the properties of both. For example, they can combine the small size of nanoparticles and their special thermal, imaging, drug carrier, or magnetic characteristics with the abilities of antibodies, such as specific and selective recognition. The hybrid product will show versatility and specificity. In this review, we analyse both antibodies and nanoparticles, focusing especially on the recent developments for antibody-conjugated nanoparticles, offering the researcher an overview of the different applications and possibilities of these hybrid carriers.

  11. Standardization of anti-DNA antibody assays.

    Science.gov (United States)

    Pisetsky, David S

    2013-07-01

    Antibodies to DNA (anti-DNA) are the serological hallmark of systemic lupus erythematosus and represent important biomarkers for clinical and research purposes. These antibodies are part of a family of antibodies to nucleosomes and bind to conserved sites widely present on DNA. While the value of anti-DNA as a biomarker is well established, the assay for these antibodies has involved a variety of DNA sources and systems to detect DNA-anti-DNA interactions. The influence of these variations on antibody detection has complicated assay standardization. As an antigen, DNA has unique features since it is a highly charged polymer that has structural heterogeneity. This heterogeneity can affect antigenicity which can vary on the basis of DNA origin, size, conformation and mobility. In addition, as a polymer, DNA can promote patterns of antibody binding based on monogamous or bivalent interaction which require an extended polynucleotide structure. Understanding the nature of DNA as an antigen can facilitate interpretation of serological tests and underpin efforts at better standardization.

  12. Quality control of antibodies for assay development.

    Science.gov (United States)

    Schumacher, Sarah; Seitz, Harald

    2016-09-25

    Antibodies are used as powerful tools in basic research, for example, in biomarker identification, and in various forms for diagnostics, for example, identification of allergies or autoimmune diseases. Due to their robustness and ease of handling, immunoassays are favourite methods for investigation of various biological or medical questions. Nevertheless in many cases, additional analyses such as mass spectrometry are used to validate or confirm the results of immunoassays. To minimize the workload and to increase confidence in immunoassays, there are urgent needs for antibodies which are both highly specific and well validated. Unfortunately many commercially available antibodies are neither well characterized nor fully tested for cross-reactivities. Adequate quality control and validation of an antibody is time-consuming and can be frustrating. Such validation needs to be performed for every assay/application. However, where an antibody validation is successful, a highly specific and stable reagent will be on hand. This article describes the validation processes of antibodies, including some often neglected factors, as well as unspecific binding to other sample compounds in a multiparameter diagnostic assay. The validation consists of different immunological methods, with important assay controls, and is performed in relation to the development of a diagnostic test. PMID:26873787

  13. Avian Diagnostic and Therapeutic Antibodies to Viral Emerging Pathogens

    Energy Technology Data Exchange (ETDEWEB)

    David Bradley

    2011-03-31

    During the current period the following key objectives were achieved: demonstration of high titer antibody production by geese following immunization with inactived H1N1 virus; completion of the epitope mapping of West Nile Virus-specific goose antibodies and initiation of epitope mapping of H1N1 flu-specific goose antibodies; advancement in scalable purification of goose antibodies.

  14. Detección de anticuerpos para BVDV y BoHV-1 en llamas de la región de Tandil - Provincia de Buenos Aires Detection of antibodies to BVDV and BoHV-1 in llamas from Tandil - Province of Buenos Aires

    Directory of Open Access Journals (Sweden)

    P.E. Morán

    2010-12-01

    Full Text Available La llama (Lama glama es la especie predominante de camélidos sudamericanos de la República Argentina, con una población aproximada de 200.000 animales. El aumento de la actividad productiva ha incrementado el transporte y la distribución de animales hacia diferentes regiones del país. Como consecuencia, estas especies interactúan con el ganado doméstico posibilitando la diseminación de agentes infecciosos propios y comunes entre especies. Considerando que los camélidos sudamericanos son susceptibles a la infección con pestivirus y herpesvirus, se planteó realizar un relevamiento serológico de anticuerpos neutralizantes para el virus de la diarrea viral bovina (BVDV y el herpesvirus bovino 1 (BoHV-1. Se analizaron 49 animales de dos rebaños de llamas de la región de Tandil mediante la técnica de seroneutralización sobre cultivos celulares de la línea Madin Darby Bovine Kidney. El 22% (11/49 de los animales expresaron seropositividad a BoHV-1, mientras que el 2% (1/49 fue positivo a BVDV. Estos resultados sugieren circulación viral en la población analizada. Es necesario el monitoreo continuo en las poblaciones de estos rumiantes para identificar factores de riesgo involucrados en la epidemiología de estos agentes virales, considerando los potenciales efectos sobre los programas de control y erradicación.Llama (Lama glama is the predominant species of South American camelids in Argentine Republic, with an estimated population of 200.000 animals. The increasing of productive activity raised the animals transport and distribution to different regions of the country. As a result, the direct contact of these species with cattle increase opportunities for spreading own and common infectious agents among species. Considering that American camelids are susceptible to pestivirus and herpesvirus infections, it was proposed to carry out a serological searching of neutralizing antibodies to bovine viral diarrhea (BVDV and bovine

  15. A multi-Fc-species system for recombinant antibody production

    OpenAIRE

    Nizak Clément; Vielemeyer Ole; El Marjou Ahmed; Moutel Sandrine; Benaroch Philippe; Dübel Stefan; Perez Franck

    2009-01-01

    Abstract Background Genomic, transcriptomic and proteomic projects often suffer from a lack of functional validation creating a strong demand for specific and versatile antibodies. Antibody phage display represents an attractive approach to select rapidly in vitro the equivalent of monoclonal antibodies, like single chain Fv antibodies, in an inexpensive and animal free way. However, so far, recombinant antibodies have not managed to impose themselves as efficient alternatives to natural anti...

  16. A recombinant, fully human monoclonal antibody with antitumor activity constructed from phage-displayed antibody fragments

    NARCIS (Netherlands)

    Huls, GA; Heijnen, IAFM; Cuomo, ME; Koningsberger, JC; Boel, E; de Vries, ARV; Loyson, SAJ; Helfrich, W; Henegouwen, GPV; van Meijer, M; de Kruif, J; Logtenberg, T

    1999-01-01

    A single-chain Fv antibody fragment specific for the tumor-associated Ep-CAM molecule was isolated from a semisynthetic phage display library and converted into an intact, fully human IgG1 monoclonal antibody (huMab), The purified huMab had an affinity of 5 nM and effectively mediated tumor cell kil

  17. Thermodynamics of antibody-antigen interaction revealed by mutation analysis of antibody variable regions.

    Science.gov (United States)

    Akiba, Hiroki; Tsumoto, Kouhei

    2015-07-01

    Antibodies (immunoglobulins) bind specific molecules (i.e. antigens) with high affinity and specificity. In order to understand their mechanisms of recognition, interaction analysis based on thermodynamic and kinetic parameters, as well as structure determination is crucial. In this review, we focus on mutational analysis which gives information about the role of each amino acid residue in antibody-antigen interaction. Taking anti-hen egg lysozyme antibodies and several anti-small molecule antibodies, the energetic contribution of hot-spot and non-hot-spot residues is discussed in terms of thermodynamics. Here, thermodynamics of the contribution from aromatic, charged and hydrogen bond-forming amino acids are discussed, and their different characteristics have been elucidated. The information gives fundamental understanding of the antibody-antigen interaction. Furthermore, the consequences of antibody engineering are analysed from thermodynamic viewpoints: humanization to reduce immunogenicity and rational design to improve affinity. Amino acid residues outside hot-spots in the interface play important roles in these cases, and thus thermodynamic and kinetic parameters give much information about the antigen recognition. Thermodynamic analysis of mutant antibodies thus should lead to advanced strategies to design and select antibodies with high affinity.

  18. The antibody mining toolbox: an open source tool for the rapid analysis of antibody repertoires.

    Science.gov (United States)

    D'Angelo, Sara; Glanville, Jacob; Ferrara, Fortunato; Naranjo, Leslie; Gleasner, Cheryl D; Shen, Xiaohong; Bradbury, Andrew R M; Kiss, Csaba

    2014-01-01

    In vitro selection has been an essential tool in the development of recombinant antibodies against various antigen targets. Deep sequencing has recently been gaining ground as an alternative and valuable method to analyze such antibody selections. The analysis provides a novel and extremely detailed view of selected antibody populations, and allows the identification of specific antibodies using only sequencing data, potentially eliminating the need for expensive and laborious low-throughput screening methods such as enzyme-linked immunosorbant assay. The high cost and the need for bioinformatics experts and powerful computer clusters, however, have limited the general use of deep sequencing in antibody selections. Here, we describe the AbMining ToolBox, an open source software package for the straightforward analysis of antibody libraries sequenced by the three main next generation sequencing platforms (454, Ion Torrent, MiSeq). The ToolBox is able to identify heavy chain CDR3s as effectively as more computationally intense software, and can be easily adapted to analyze other portions of antibody variable genes, as well as the selection outputs of libraries based on different scaffolds. The software runs on all common operating systems (Microsoft Windows, Mac OS X, Linux), on standard personal computers, and sequence analysis of 1-2 million reads can be accomplished in 10-15 min, a fraction of the time of competing software. Use of the ToolBox will allow the average researcher to incorporate deep sequence analysis into routine selections from antibody display libraries. PMID:24423623

  19. Passive antibody transfer in chickens to model maternal antibody after avian influenza vaccination.

    Science.gov (United States)

    Faulkner, Olivia B; Estevez, Carlos; Yu, Qingzhong; Suarez, David L

    2013-04-15

    Birds transfer maternal antibodies (MAb) to their offspring through the egg yolk where the antibody is absorbed and enters the circulatory system. Maternal antibodies provide early protection from disease, but may interfere with the vaccination efficacy in the chick. MAb are thought to interfere with vaccine antigen processing that reduces the subsequent immune response. Once MAb titers are depleted, the chick will respond to vaccination, but they are also susceptible to viral infection. This study examines the effect of MAb on seroconversion to different viral-vectored avian influenza virus (AIV) vaccines. Chicks were given passively transferred antibodies (PTA) using AIV hyperimmunized serum, and subsequently vaccinated with a fowlpox-AIV recombinant vaccine (FPr) or a Newcastle disease virus-AIV recombinant vaccine (NDVr). Our results indicate that passively transferred antibodies led to significant reduction of seroconversion and clinical protection from virulent challenge in recombinant virus vaccinated chicks thus demonstrating maternal antibody interference to vaccination. The passive antibody transfer model system provides an important tool to evaluate maternal antibody interference to vaccination. PMID:23398721

  20. Antibodies Produced in Response to Cryptococcus neoformans Pulmonary Infection in Mice Have Characteristics of Nonprotective Antibodies

    OpenAIRE

    Zaragoza, Oscar; Casadevall, Arturo

    2004-01-01

    Murine cryptocococcal pulmonary infection elicited serum immunoglobulin M (IgM) and IgG to the capsular polysaccharide, but only IgG stained yeast cells in alveoli. Both isotypes produced punctuate immunofluorescence patterns on yeast cells like those of nonprotective antibodies. The difficulties involved in associating humoral immunity with protection in murine cryptocococcal infection could reflect nonprotective antibody responses.

  1. A study on associations between antiprothrombin antibodies, antiplasminogen antibodies and thrombosis

    NARCIS (Netherlands)

    Simmelink, MJA; De Groot, PG; Derksen, RHWM

    2003-01-01

    Anti-prothrombin antibodies area frequent cause of lupus anticoagulant (LAC), a thrombotic risk factor. Prothrombin shares structural homology with plasminogen, a kringle protein with an important role in fibrinolysis. Cross-reactivity between antiprothrombin antibodies and plasminogen has been desc

  2. Antibody-Mediated Internalization of Infectious HIV-1 Virions Differs among Antibody Isotypes and Subclasses.

    Science.gov (United States)

    Tay, Matthew Zirui; Liu, Pinghuang; Williams, LaTonya D; McRaven, Michael D; Sawant, Sheetal; Gurley, Thaddeus C; Xu, Thomas T; Dennison, S Moses; Liao, Hua-Xin; Chenine, Agnès-Laurence; Alam, S Munir; Moody, M Anthony; Hope, Thomas J; Haynes, Barton F; Tomaras, Georgia D

    2016-08-01

    Emerging data support a role for antibody Fc-mediated antiviral activity in vaccine efficacy and in the control of HIV-1 replication by broadly neutralizing antibodies. Antibody-mediated virus internalization is an Fc-mediated function that may act at the portal of entry whereby effector cells may be triggered by pre-existing antibodies to prevent HIV-1 acquisition. Understanding the capacity of HIV-1 antibodies in mediating internalization of HIV-1 virions by primary monocytes is critical to understanding their full antiviral potency. Antibody isotypes/subclasses differ in functional profile, with consequences for their antiviral activity. For instance, in the RV144 vaccine trial that achieved partial efficacy, Env IgA correlated with increased risk of HIV-1 infection (i.e. decreased vaccine efficacy), whereas V1-V2 IgG3 correlated with decreased risk of HIV-1 infection (i.e. increased vaccine efficacy). Thus, understanding the different functional attributes of HIV-1 specific IgG1, IgG3 and IgA antibodies will help define the mechanisms of immune protection. Here, we utilized an in vitro flow cytometric method utilizing primary monocytes as phagocytes and infectious HIV-1 virions as targets to determine the capacity of Env IgA (IgA1, IgA2), IgG1 and IgG3 antibodies to mediate HIV-1 infectious virion internalization. Importantly, both broadly neutralizing antibodies (i.e. PG9, 2G12, CH31, VRC01 IgG) and non-broadly neutralizing antibodies (i.e. 7B2 mAb, mucosal HIV-1+ IgG) mediated internalization of HIV-1 virions. Furthermore, we found that Env IgG3 of multiple specificities (i.e. CD4bs, V1-V2 and gp41) mediated increased infectious virion internalization over Env IgG1 of the same specificity, while Env IgA mediated decreased infectious virion internalization compared to IgG1. These data demonstrate that antibody-mediated internalization of HIV-1 virions depends on antibody specificity and isotype. Evaluation of the phagocytic potency of vaccine

  3. Platform for high-throughput antibody selection using synthetically-designed antibody libraries.

    Science.gov (United States)

    Batonick, Melissa; Holland, Erika G; Busygina, Valeria; Alderman, Dawn; Kay, Brian K; Weiner, Michael P; Kiss, Margaret M

    2016-09-25

    Synthetic humanized antibody libraries are frequently generated by random incorporation of changes at multiple positions in the antibody hypervariable regions. Although these libraries have very large theoretical diversities (>10(20)), the practical diversity that can be achieved by transformation of Escherichia coli is limited to about 10(10). To constrain the practical diversity to sequences that more closely mimic the diversity of natural human antibodies, we generated a scFv phage library using entirely pre-defined complementarity determining regions (CDR). We have used this library to select for novel antibodies against four human protein targets and demonstrate that identification of enriched sequences at each of the six CDRs in early selection rounds can be used to reconstruct a consensus antibody with selectivity for the target. PMID:26607994

  4. Antibody engineering using phage display with a coiled-coil heterodimeric Fv antibody fragment.

    Directory of Open Access Journals (Sweden)

    Xinwei Wang

    Full Text Available A Fab-like antibody binding unit, ccFv, in which a pair of heterodimeric coiled-coil domains was fused to V(H and V(L for Fv stabilization, was constructed for an anti-VEGF antibody. The anti-VEGF ccFv showed the same binding affinity as scFv but significantly improved stability and phage display level. Furthermore, phage display libraries in the ccFv format were constructed for humanization and affinity maturation of the anti-VEGF antibody. A panel of V(H frameworks and V(H-CDR3 variants, with a significant improvement in affinity and expressibility in both E. coli and yeast systems, was isolated from the ccFv phage libraries. These results demonstrate the potential application of the ccFv antibody format in antibody engineering.

  5. Adaptive responses to antibody based therapy.

    Science.gov (United States)

    Rodems, Tamara S; Iida, Mari; Brand, Toni M; Pearson, Hannah E; Orbuch, Rachel A; Flanigan, Bailey G; Wheeler, Deric L

    2016-02-01

    Receptor tyrosine kinases (RTKs) represent a large class of protein kinases that span the cellular membrane. There are 58 human RTKs identified which are grouped into 20 distinct families based upon their ligand binding, sequence homology and structure. They are controlled by ligand binding which activates intrinsic tyrosine-kinase activity. This activity leads to the phosphorylation of distinct tyrosines on the cytoplasmic tail, leading to the activation of cell signaling cascades. These signaling cascades ultimately regulate cellular proliferation, apoptosis, migration, survival and homeostasis of the cell. The vast majority of RTKs have been directly tied to the etiology and progression of cancer. Thus, using antibodies to target RTKs as a cancer therapeutic strategy has been intensely pursued. Although antibodies against the epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) have shown promise in the clinical arena, the development of both intrinsic and acquired resistance to antibody-based therapies is now well appreciated. In this review we provide an overview of the RTK family, the biology of EGFR and HER2, as well as an in-depth review of the adaptive responses undertaken by cells in response to antibody based therapies directed against these receptors. A greater understanding of these mechanisms and their relevance in human models will lead to molecular insights in overcoming and circumventing resistance to antibody based therapy. PMID:26808665

  6. Cytolytic antibodies to melanocytes in vitiligo.

    Science.gov (United States)

    Cui, J; Arita, Y; Bystryn, J C

    1993-06-01

    Patients with vitiligo have been found to have circulating antibodies to pigment cells. To evaluate the functional activity of these antibodies, a highly sensitive europium release assay was used to compare complement-mediated cytolysis of human melanocytes by sera of 56 patients with vitiligo (20 with active disease, 25 with inactive disease, 11 with unidentified disease activity) and 47 control individuals. Significant melanocyte lysis was mediated by 32 (57%) of the patients with vitiligo but by only three (6%) of the control sera (p < 0.001), and by 17 (85%) of 20 patients with active vitiligo versus 11 (44%) of 25 patients with inactive disease (p < 0.025). Mean melanocyte lysis by vitiligo sera was 24% versus 6% by control sera (p < 0.0001). A subset of 12 vitiligo sera with high titers of cytolytic antibodies to melanocytes (34% mean cytolysis) reacted minimally (< 2% mean cytolysis) to a panel of control cells that included human and murine melanomas, human fibroblasts, lung carcinoma, and rhabdomyosarcoma. These findings indicate that antibodies present in patients with vitiligo have the functional ability to selectively kill melanocytes and are more common in active disease. These observations support, but do not prove, the hypothesis that vitiligo is an autoimmune disease and that anti-pigment cell antibodies have a role in inducing the disease. PMID:8496621

  7. Antigen-Antibody Interaction Database (AgAbDb): a compendium of antigen-antibody interactions.

    Science.gov (United States)

    Kulkarni-Kale, Urmila; Raskar-Renuse, Snehal; Natekar-Kalantre, Girija; Saxena, Smita A

    2014-01-01

    Antigen-Antibody Interaction Database (AgAbDb) is an immunoinformatics resource developed at the Bioinformatics Centre, University of Pune, and is available online at http://bioinfo.net.in/AgAbDb.htm. Antigen-antibody interactions are a special class of protein-protein interactions that are characterized by high affinity and strict specificity of antibodies towards their antigens. Several co-crystal structures of antigen-antibody complexes have been solved and are available in the Protein Data Bank (PDB). AgAbDb is a derived knowledgebase developed with an objective to compile, curate, and analyze determinants of interactions between the respective antigen-antibody molecules. AgAbDb lists not only the residues of binding sites of antigens and antibodies, but also interacting residue pairs. It also helps in the identification of interacting residues and buried residues that constitute antibody-binding sites of protein and peptide antigens. The Antigen-Antibody Interaction Finder (AAIF), a program developed in-house, is used to compile the molecular interactions, viz. van der Waals interactions, salt bridges, and hydrogen bonds. A module for curating water-mediated interactions has also been developed. In addition, various residue-level features, viz. accessible surface area, data on epitope segment, and secondary structural state of binding site residues, are also compiled. Apart from the PDB numbering, Wu-Kabat numbering and explicit definitions of complementarity-determining regions are provided for residues of antibodies. The molecular interactions can be visualized using the program Jmol. AgAbDb can be used as a benchmark dataset to validate algorithms for prediction of B-cell epitopes. It can as well be used to improve accuracy of existing algorithms and to design new algorithms. AgAbDb can also be used to design mimotopes representing antigens as well as aid in designing processes leading to humanization of antibodies. PMID:25048123

  8. Antibody-based biological toxin detection

    Energy Technology Data Exchange (ETDEWEB)

    Menking, D.E.; Goode, M.T. [Army Edgewood Research, Development and Engineering Center, Aberdeen Proving Ground, MD (United States)

    1995-12-01

    Fiber optic evanescent fluorosensors are under investigation in our laboratory for the study of drug-receptor interactions for detection of threat agents and antibody-antigen interactions for detection of biological toxins. In a direct competition assay, antibodies against Cholera toxin, Staphylococcus Enterotoxin B or ricin were noncovalently immobilized on quartz fibers and probed with fluorescein isothiocyanate (FITC) - labeled toxins. In the indirect competition assay, Cholera toxin or Botulinum toxoid A was immobilized onto the fiber, followed by incubation in an antiserum or partially purified anti-toxin IgG. These were then probed with FITC-anti-IgG antibodies. Unlabeled toxins competed with labeled toxins or anti-toxin IgG in a dose dependent manner and the detection of the toxins was in the nanomolar range.

  9. Origin and pathogenesis of antiphospholipid antibodies

    Directory of Open Access Journals (Sweden)

    C.M. Celli

    1998-06-01

    Full Text Available Antiphospholipid antibodies (aPL are a heterogeneous group of antibodies that are detected in the serum of patients with a variety of conditions, including autoimmune (systemic lupus erythematosus, infectious (syphilis, AIDS and lymphoproliferative disorders (paraproteinemia, myeloma, lymphocytic leukemias. Thrombosis, thrombocytopenia, recurrent fetal loss and other clinical complications are currently associated with a subgroup of aPL designating the antiphospholipid syndrome. In contrast, aPL from patients with infectious disorders are not associated with any clinical manifestation. These findings led to increased interest in the origin and pathogenesis of aPL. Here we present the clinical features of the antiphospholipid syndrome and review the origin of aPL, the characteristics of experimentally induced aPL and their historical background. Within this context, we discuss the most probable pathogenic mechanisms induced by these antibodies.

  10. Imaging spectrum of primary antiphospholipid antibody syndrome

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Kwon Ha; Won, Jong Jin [Wonkwang University Hospital, Iksan (Korea, Republic of); Ha, Hyun Kwon; Kim, Jung Hoon; Kim, Jeong Gon; Ki, Won Woo; Kim, Pyo Nyun; Lee, Moon Gyu; Auh, Yong Ho [Asan Medical Center, Seoul (Korea, Republic of)

    1998-04-01

    Antiphospholipid antibody syndrome is recognized as one of the most important causes of hypercoagulability. It can be clinically diagnosed if patients have experienced unexplained recurrent venous or arterial thrombosis, recurrent fetal loss, or thrombocytopenia in the presence of circulating autoantibodies to phospholipids, such as anticardiolipin antibody or lupus anticoagulant. Approximately half of all patients with this syndrome do not have associated systemic disease, and their condition is described as primary antiphospholipid antibody syndrome (PAPS). In the remainder, the syndrome is accompanied by systemic lupus erythematosus or other connective tissue diseases, and is known as secondary antiphospholipid syndrome (1). The purpose of this paper is to illustrate the systemic manifestation of PAPS, focusing on the radiological findings of CT, MR and angiography in clinically proven patients. (author). 8 refs., 10 figs.

  11. Back to the future: recombinant polyclonal antibody therapeutics.

    Science.gov (United States)

    Wang, Xian-Zhe; Coljee, Vincent W; Maynard, Jennifer A

    2013-11-01

    Antibody therapeutics are one of the fastest growing classes of pharmaceuticals, with an annual US market over $20 billion, developed to treat a variety of diseases including cancer, auto-immune and infectious diseases. Most are currently administered as a single molecule to treat a single disease, however there is mounting evidence that cocktails of multiple antibodies, each with a unique binding specificity and protective mechanism, may improve clinical efficacy. Here, we review progress in the development of oligoclonal combinations of antibodies to treat disease, focusing on identification of synergistic antibodies. We then discuss the application of modern antibody engineering technologies to produce highly potent antibody preparations, including oligoclonal antibody cocktails and truly recombinant polyclonal antibodies. Specific examples illustrating the synergy conferred by multiple antibodies will be provided for diseases caused by botulinum toxin, cancer and immune thrombocytopenia. The bioprocessing and regulatory options for these preparations will be discussed. PMID:24443710

  12. Characterization of monoclonal antibodies directed against human thyroid stimulating hormone

    International Nuclear Information System (INIS)

    Monoclonal antibodies directed against human thyroid stimulating hormone (TSH) were obtained from hybrid myelomas, following fusion of mouse NSI myeloma cells with mouse spleen cells. Ten different antibodies were obtained from 4 separate fusions. Eight antibodies were of the IgG1 subclass. Affinities of antibodies for TSH were in the range 2 x 108-5 x 1010 M-1. Five of the antibodies were specific for TSH and did not react with LH, FSH or hCG. The remaining antibodies reacted with all these hormones and were assumed to recognise their common (α) subunit. The 5 specific antibodies fell into 3 subgroups recognising distinct antigenic determinants, whereas the 5 non-specific antibodies recognised a single determinant or closely related set of sites. It is concluded that these antibodies should be valuable reagents for use in sensitive and specific two-site immunoradiometric assays. (Auth.)

  13. Anti-DNA antibodies--quintessential biomarkers of SLE.

    Science.gov (United States)

    Pisetsky, David S

    2016-02-01

    Antibodies that recognize and bind to DNA (anti-DNA antibodies) are serological hallmarks of systemic lupus erythematosus (SLE) and key markers for diagnosis and disease activity. In addition to common use in the clinic, anti-DNA antibody testing now also determines eligibility for clinical trials, raising important questions about the nature of the antibody-antigen interaction. At present, no 'gold standard' for serological assessment exists, and anti-DNA antibody binding can be measured with a variety of assay formats, which differ in the nature of the DNA substrates and in the conditions for binding and detection of antibodies. A mechanism called monogamous bivalency--in which high avidity results from simultaneous interaction of IgG Fab sites with a single polynucleotide chain--determines anti-DNA antibody binding; this mechanism might affect antibody detection in different assay formats. Although anti-DNA antibodies can promote pathogenesis by depositing in the kidney or driving cytokine production, they are not all alike, pathologically, and anti-DNA antibody expression does not necessarily correlate with active disease. Levels of anti-DNA antibodies in patients with SLE can vary over time, distinguishing anti-DNA antibodies from other pathogenic antinuclear antibodies. Elucidation of the binding specificities and the pathogenic roles of anti-DNA antibodies in SLE should enable improvements in the design of informative assays for both clinical and research purposes.

  14. The maturation of antibody technology for the HIV epidemic.

    Science.gov (United States)

    Winnall, Wendy R; Beasley, Matthew D; Center, Rob J; Parsons, Matthew S; Kiefel, Ben R; Kent, Stephen J

    2014-08-01

    Antibodies are one of our most useful biological tools. Indeed, improvements in antibody-based technologies have ushered in a new era of antibody-based therapeutics, research and diagnostic tools. Although improved technologies have led to the development of therapeutic antibodies for treatment of malignancies and inflammatory conditions, the use of advanced antibody technology in the therapy of viral infections is in its infancy. Non-human primate studies have demonstrated that antibodies against the HIV envelope can both prevent viral infection and control viremia. Despite the obvious potential of antibody therapies against HIV, there remain limitations in production and purification capacity that require further research. Recent advances in recombinant antibody technology have led to the development of a range of novel antibody fragments, such as single-domain nanobodies and bispecific antibodies, that are capable of targeting cancer cells to cytotoxic T cells. Novel antibody production techniques have also been designed, allowing antibodies to be obtained from non-mammalian cells, bovine colostrum and the periplasm and cytoplasm of bacteria. These advances may allow large-scale production of HIV antibodies that are capable of protecting against HIV infection or serving as therapeutics that reduce the need for life-long antiretroviral treatment. This review summarises recent advances in antibody-based technologies and discusses the possibilities and challenges of using these advances to design prophylactics and therapeutics against HIV. PMID:24797582

  15. Antibody engineering & therapeutics, the annual meeting of the antibody society December 7–10, 2015, San Diego, CA, USA

    Science.gov (United States)

    Pauthner, Matthias; Yeung, Jenny; Ullman, Chris; Bakker, Joost; Wurch, Thierry; Reichert, Janice M.; Lund-Johansen, Fridtjof; Bradbury, Andrew R.M.; Carter, Paul J.; Melis, Joost P.M.

    2016-01-01

    ABSTRACT The 26th Antibody Engineering & Therapeutics meeting, the annual meeting of The Antibody Society united over 800 participants from all over the world in San Diego from 6–10 December 2015. The latest innovations and advances in antibody research and development were discussed, covering a myriad of antibody-related topics by more than 100 speakers, who were carefully selected by The Antibody Society. As a prelude, attendees could join the pre-conference training course focusing, among others, on the engineering and enhancement of antibodies and antibody-like scaffolds, bispecific antibody engineering and adaptation to generate chimeric antigen receptor constructs. The main event covered 4 d of scientific sessions that included antibody effector functions, reproducibility of research and diagnostic antibodies, new developments in antibody-drug conjugates (ADCs), preclinical and clinical ADC data, new technologies and applications for bispecific antibodies, antibody therapeutics for non-cancer and orphan indications, antibodies to harness the cellular immune system, building comprehensive IgVH-gene repertoires through discovering, confirming and cataloging new germline IgVH genes, and overcoming resistance to clinical immunotherapy. The Antibody Society's special session focused on “Antibodies to watch” in 2016. Another special session put the spotlight on the limitations of the new definitions for the assignment of antibody international nonproprietary names introduced by the World Health Organization. The convention concluded with workshops on computational antibody design and on the promise and challenges of using next-generation sequencing for antibody discovery and engineering from synthetic and in vivo libraries. PMID:26909869

  16. Utility of feline coronavirus antibody tests.

    Science.gov (United States)

    Addie, Diane D; le Poder, Sophie; Burr, Paul; Decaro, Nicola; Graham, Elizabeth; Hofmann-Lehmann, Regina; Jarrett, Oswald; McDonald, Michael; Meli, Marina L

    2015-02-01

    Eight different tests for antibodies to feline coronavirus (FCoV) were evaluated for attributes that are important in situations in veterinary practice. We compared four indirect immunofluorescent antibody tests (IFAT), one enzyme-linked immunosorbent assay (ELISA) (FCoV Immunocomb; Biogal) and three rapid immunochromatographic (RIM) tests against a panel of samples designated by consensus as positive or negative. Specificity was 100% for all but the two IFATs based on transmissible gastroenteritis virus (TGEV), at 83.3% and 97.5%. The IFAT and ELISA tests were best for obtaining an antibody titre and for working in the presence of virus. The RIM tests were the best for obtaining a result quickly (10-15 mins); of these, the Speed F-Corona was the most sensitive, at 92.4%, followed by FASTest feline infectious peritonitis (FIP; 84.6%) and Anigen Rapid FCoV antibody test (64.1%). Sensitivity was 100% for the ELISA, one FCoV IFAT and one TGEV IFAT; and 98.2% for a second TGEV IFA and 96.1% for a second FCoV IFAT. All tests worked with effusions, even when only blood products were stipulated in the instruction manual. The ELISA and Anigen RIM tests were best for small quantities of sample. The most appropriate FCoV antibody test to use depends on the reason for testing: in excluding a diagnosis of FIP, sensitivity, specificity, small sample quantity, rapidity and ability to work in the presence of virus all matter. For FCoV screening, speed and sensitivity are important, and for FCoV elimination antibody titre is essential. PMID:24966245

  17. Diagnostic Value of Animal-Side Antibody Assays for Rapid Detection of Mycobacterium bovis or Mycobacterium microti Infection in South American Camelids▿

    OpenAIRE

    Lyashchenko, Konstantin P; Greenwald, Rena; Esfandiari, Javan; Rhodes, Shelley; Dean, Gillian; de la Rua-Domenech, Ricardo; Meylan, Mireille; Vordermeier, HMartin; Zanolari, Patrik

    2011-01-01

    Tuberculosis (TB) in South American camelids (SAC) is caused by Mycobacterium bovis or Mycobacterium microti. Two serological methods, rapid testing (RT) and the dual-path platform (DPP) assay, were evaluated using naturally infected SAC. The study population included 156 alpacas and 175 llamas in Great Britain, Switzerland, and the United States. TB due to M. bovis (n = 44) or M. microti (n = 8) in 35 alpacas and 17 llamas was diagnosed by gross pathology examination and culture. Control ani...

  18. Antibodies for detecting and quantifying anticoagulant agents

    OpenAIRE

    Salvador, Juan Pablo; Marco, María Pilar

    2012-01-01

    [EN] The present invention relates to the design of haptens that are structurally related to coumarin oral anticoagulant compounds (COAC), to be used for the production of specific antibodies against said type of substances and the subsequent use thereof for the development of diagnosis tools for use in laboratories or in point-of-care (PoC) devices. In particular, the produced antibodies have been used to develop a diagnosis tool that enables the plasma levels of COAC to be quantified in pat...

  19. Basic immunology of antibody targeted radiotherapy

    International Nuclear Information System (INIS)

    Antibody targeted radiotherapy brings an important new treatment modality to Radiation oncology clinic. Radiation dose to tumor and normal tissues are determined by a complex interplay of antibody, antigen, tumor, radionuclide, and host-related factors. A basic understanding of these immunologic and physiologic factors is important to optimally utilize this therapy in the clinic. Preclinical and clinical studies need to be continued to broaden our understanding and to develop new strategies to further improve the efficacy of this promising form of targeted therapy

  20. Radiolabelling of monoclonal antibodies for radiotherapy. Thailand

    International Nuclear Information System (INIS)

    Nuclear medicine is now playing a great role not only in diagnostic application but also in therapy of cancer patients. Under the concept of targeted radiotherapy, a number of radiopharmaceuticals based on radiolabelled biomolecules had been evaluated for treatment of cancer by many investigators. Of these, monoclonal antibodies and some small specific peptides labelled with beta emitting radiometals such as Sm-153, Re-186, Re-188 or Y-90, are being introduced into clinical trials. The objective of this project is to develop laboratory procedures to label monoclonal antibodies, peptide or other proteins with beta emitting radionuclides to prepare radiopharmaceuticals for therapeutic purpose

  1. Radiolabelling of monoclonal antibodies for radiotherapy

    International Nuclear Information System (INIS)

    Nuclear medicine is now playing a great role not only in diagnostic application but also in therapy of cancer patients. Under the concept of targeted radiotherapy, a number of radiopharmaceuticals based on radiolabelled biomolecules had been evaluated for treatment of cancer by many investigators. Of these, monoclonal antibodies and some small specific peptides labelled with beta emitting radiometals such as Sm-153, Re-186, Re-188 or Y-90, are being introduced into clinical trials. The objective of this project is to develop laboratory procedures to label monoclonal antibodies, peptide or other proteins with beta emitting radionuclides to prepare radiopharmaceuticals for therapeutic purpose

  2. Modeling single cell antibody excretion on a biosensor.

    Science.gov (United States)

    Stojanović, Ivan; Baumgartner, Wolfgang; van der Velden, Thomas J G; Terstappen, Leon W M M; Schasfoort, Richard B M

    2016-07-01

    We simulated, using Comsol Multiphysics, the excretion of antibodies by single hybridoma cells and their subsequent binding on a surface plasmon resonance imaging (SPRi) sensor. The purpose was to confirm that SPRi is suitable to accurately quantify antibody (anti-EpCAM) excretion. The model showed that antibody loss by diffusion away from the sensor was less than 1%. Unexpectedly, more than 99% of the excreted antibodies were captured on the sensor. These data prove the remarkable phenomenon that the SPRi output of cellular antibody excretion and its subsequent binding, performed under the conditions described here, is directly usable for quantification of single cell antibody production rates. PMID:27040182

  3. Antibody-mediated immune suppression is improved when blends of anti-RBC monoclonal antibodies are used in mice.

    Science.gov (United States)

    Bernardo, Lidice; Amash, Alaa; Marjoram, Danielle; Lazarus, Alan H

    2016-08-25

    Although the prevention of hemolytic disease of the fetus and newborn is highly effective using polyclonal anti-D, a recombinant alternative is long overdue. Unfortunately, anti-D monoclonal antibodies have been, at best, disappointing. To determine the primary attribute defining an optimal antibody, we assessed suppression of murine red blood cell (RBC) immunization by single-monoclonal antibodies vs defined blends of subtype-matched antibodies. Allogeneic RBCs expressing the HOD antigen (hen egg lysozyme [HEL]-ovalbumin-human transmembrane Duffy(b)) were transfused into naïve mice alone or together with selected combinations of HEL-specific antibodies, and the resulting suppressive effect was assessed by evaluating the antibody response. Polyclonal HEL antibodies dramatically inhibited the antibody response to the HOD antigen, whereas single-monoclonal HEL antibodies were less effective despite the use of saturating doses. A blend of monoclonal HEL-specific antibodies reactive with different HEL epitopes significantly increased the suppressive effect, whereas a blend of monoclonal antibodies that block each other's binding to the HEL protein did not increase suppression. In conclusion, these data show that polyclonal antibodies are superior to monoclonal antibodies at suppressing the immune response to the HOD cells, a feature that can be completely recapitulated using monoclonal antibodies to different epitopes. PMID:27330002

  4. Treatment of leukemia with radiolabeled monoclonal antibodies.

    Science.gov (United States)

    Sgouros, G; Scheinberg, D A

    1993-01-01

    In contrast to radioimmunotherapy of solid disease, wherein the primary obstacle to success is access of radiolabeled antibody to antigen-positive cells, in the treatment of leukemia delivering a lethal absorbed dose to the isolated cell appears to be the primary obstacle. The isolated cell is defined as one that is exposed only to self-irradiation (from internalized or surface-bound radiolabeled antibody) and to irradiation from free antibody in the blood. It is isolated in the sense that the particulate (beta, electron, alpha) emissions from its nearest neighboring antigen-positive cell do not contribute to its absorbed dose. Disease in the bone marrow and other tissues, since it is confined to a smaller volume, is more easily eradicated because the absorbed dose to a given cell nucleus is enhanced by emissions from adjacent cells (a smaller fraction of the emission energy is 'wasted'). The optimization simulations presented above for the M195 antibody suggest that the optimum dose of antibody that should be administered is that required to yield a concentration within the distribution volume of the antibody that is approximately equal to the concentration of antigen sites as determined by the tumor burden. Although not specifically considered in the modeling example presented above, antibody internalization and catabolism may be expected to play an important role in radioimmunotherapy treatment planning of leukemia. Depending upon the kinetics of internalization and catabolism, the absorbed dose to the red marrow and to antigen-positive cells may be reduced considerably, since catabolism, assuming that it is followed by rapid extrusion of the radioactive label, would decrease the cells' exposure time considerably. The recently demonstrated effectiveness of radioimmunotherapy in certain cases of B-cell lymphoma and in reducing tumor burden in acute myelogenous leukemia suggests that radioimmunotherapy is beginning to fulfill the promise held when it was initially

  5. Anti Rh Hemolytic Disease due to Anti C Antibody: Is Testing for Anti D Antibodies Enough?

    OpenAIRE

    Negi, Gita; Singh, Gaur Dushyant

    2011-01-01

    Rh blood group system is a complex blood group system. Rh antibodies are produced in Rh negative individuals following exposure to foreign RBCs after transfusion or pregnancy. Anti C is a rare cause of hemolytic disease of newborn and is very scarcely reported in the literature. The aim of the present case report of Hemolytic disease caused by Anti C antibody is to bring out the fact that antibodies other than anti D should be considered in cases that give a suggestive history but no evidence...

  6. Phase Transitions in Antibody Solutions: from Pharmaceuticals to Human Disease

    Science.gov (United States)

    Wang, Ying; Lomakin, Aleksey; Benedek, George; Dana Farber Cancer Institute Collaboration; Amgen Inc. Collaboration

    2014-03-01

    Antibodies are very important proteins. Natural antibodies play essential role in the immune system of human body. Pharmaceutical antibodies are used as drugs. Antibodies are also indispensable tools in biomedical research and diagnostics. Recently, a number of observations of phase transitions of pharmaceutical antibodies have been reported. These phase transitions are undesirable from the perspective of colloid stability of drug solutions in processing and storage, but can be used for protein purification, X-ray crystallography, and improving pharmokinetics of drugs. Phase transitions of antibodies can also take place in human body, particularly in multiple myeloma patients who overproduce monoclonal antibodies. These antibodies, in some cases, crystallize at body temperature and cause severe complications called cryoglobulinemia. I will present the results of our current studies on phase transitions of both pharmaceutical antibodies and cryoglobulinemia-associated antibodies. These studies have shown that different antibodies have different propensity to undergo phase transitions, but their phase behavior has universal features which are remarkably different from those of spherical proteins. I will discuss how studies of phase behavior can be useful in assessing colloid stability of pharmaceutical antibodies and in early diagnostics of cryoglobulinemia, as well as general implications of the fact that some antibodies can precipitate at physiological conditions.

  7. IgA Antibodies in Rett Syndrome

    Science.gov (United States)

    Reichelt, K. L.; Skjeldal, O.

    2006-01-01

    The level of IgA antibodies to gluten and gliadin proteins found in grains and to casein found in milk, as well as the level of IgG to gluten and gliadin, have been examined in 23 girls with Rett syndrome and 53 controls. Highly statistically significant increases were found for the Rett population compared to the controls. The reason for this…

  8. Antibody-based imaging strategies for cancer

    NARCIS (Netherlands)

    Warram, Jason M.; de Boer, Esther; Sorace, Anna G.; Chung, Thomas K.; Kim, Hyunki; Pleijhuis, Rick G.; van Dam, Gooitzen M.; Rosenthal, Eben L.

    2014-01-01

    Although mainly developed for preclinical research and therapeutic use, antibodies have high antigen specificity, which can be used as a courier to selectively deliver a diagnostic probe or therapeutic agent to cancer. It is generally accepted that the optimal antigen for imaging will depend on both

  9. Bone marrow dosimetry for monoclonal antibody therapy

    International Nuclear Information System (INIS)

    Immunoglobulins must permeate through the basement membrane of capillaries in order to enter the extracellular space (ECS) of tissue. Since the process is quite slow, the blood plasma activity in various organs contributes considerably to the radiation dose of the dose-limiting tissues. In bone marrow the basement membrane is absent and the blood circulation is functionally open. Therefore, blood plasma and marrow ECS maintain equal concentrations of labeled immunoglobulins. A combination of factors including intravenous administration, slow absorption into most tissues, slow breakdown and elimination of labeled immunoglobulin, and rapid entry into bone marrow ECS as well as known radiosensitivity of marrow led the authors to expect this tissue would prove to be the primary tissue at risk for systemic monoclonal antibody therapy. They have developed and applied in a Phase I clinical study of 131I labeled CEA antibody a procedure for estimation of radiation dose to red bone marrow. Serieal measurements of blood plasma and total body retention are carried out. Binding of labeled antibody to the cellular components of blood is verified to be very low. They have observed bone marrow depression at doses greater than 400 rad. If no special procedures are used to reconstitute marrow after radiation treatment, this level represents a much greater than generally recognized limitation to radiolabeled monoclonal antibody therapy. 25 references, 4 tables

  10. JDIP Genomics, Antibodies, and Proteomics Core Update

    Science.gov (United States)

    The JDIP Genomics, Proteomics, and Antibodies Core has developed several resources that are available for use by JDIP researchers. Five tasks have been completed or are in progress: Task 1 – Transposon mutants: Nearly 24,000 gene disruption M. paratuberculosis mutants are now available for JDIP re...

  11. New Antibody Conjugates in Cancer Therapy

    Directory of Open Access Journals (Sweden)

    Serengulam V. Govindan

    2010-01-01

    Full Text Available Targeting of radiation, drugs, and protein toxins to cancers selectively with monoclonal antibodies (MAbs has been a topic of considerable interest and an area of continued development. Radioimmunotherapy (RAIT of lymphoma using directly labeled MAbs is of current interest after approval of two radiolabeled anti-CD20 MAbs, as illustrated with the near 100% overall response rate obtained in a recent clinical trial using an investigational radiolabeled anti-CD22 MAb, 90Y-epratuzumab. The advantage of pretargeted RAIT over directly labeled MAbs is continuing to be validated in preclinical models of lymphoma and solid tumors. Importantly, the advantages of combining RAIT with radiation sensitizers, with immunotherapy, or a drug conjugate targeting a different antigen are being studied clinically and preclinically. The area of drug-conjugated antibodies is progressing with encouraging data published for the trastuzumab-DM1 conjugate in a phase I clinical trial in HER2-positive breast cancer. The Dock-and-Lock platform technology has contributed to the design and the evaluation of complex antibody-cytokine and antibody-toxin conjugates. This review describes the advances made in these areas, with illustrations taken from advances made in the authors' institutions.

  12. Developing recombinant antibodies for biomarker detection

    Energy Technology Data Exchange (ETDEWEB)

    Baird, Cheryl L.; Fischer, Christopher J.; Pefaur, Noah B.; Miller, Keith D.; Kagen, Jacob; Srivastava, Sudhir; Rodland, Karin D.

    2010-10-01

    Monoclonal antibodies (mAbs) have an essential role in biomarker validation and diagnostic assays. A barrier to pursuing these applications is the reliance on immunization and hybridomas to produce mAbs, which is time-consuming and may not yield the desired mAb. We recommend a process flow for affinity reagent production that utilizes combinatorial protein display systems (eg, yeast surface display or phage display) rather than hybridomas. These systems link a selectable phenotype-binding conferred by an antibody fragment-with a means for recovering the encoding gene. Recombinant libraries obtained from immunizations can produce high-affinity antibodies (<10 nM) more quickly than other methods. Non-immune libraries provide an alternate route when immunizations are not possible, or when suitable mAbs are not recovered from an immune library. Directed molecular evolution (DME) is an integral part of optimizing mAbs obtained from combinatorial protein display, but can also be used on hybridoma-derived mAbs. Variants can easily be obtained and screened to increase the affinity of the parent mAb (affinity maturation). We discuss examples where DME has been used to tailor affinity reagents to specific applications. Combinatorial protein display also provides an accessible method for identifying antibody pairs, which are necessary for sandwich-type diagnostic assays.

  13. Ebola Virus Antibodies in Fruit Bats, Bangladesh

    OpenAIRE

    Kevin J Olival; Islam, Ariful; Yu, Meng; Anthony, Simon J.; Jonathan H Epstein; Khan, Shahneaz Ali; Khan, Salah Uddin; Crameri, Gary; Wang, Lin-Fa; Lipkin, W. Ian; Luby, Stephen P.; Daszak, Peter

    2013-01-01

    To determine geographic range for Ebola virus, we tested 276 bats in Bangladesh. Five (3.5%) bats were positive for antibodies against Ebola Zaire and Reston viruses; no virus was detected by PCR. These bats might be a reservoir for Ebola or Ebola-like viruses, and extend the range of filoviruses to mainland Asia.

  14. Neutralizing antibodies in hepatitis C virus infection

    Institute of Scientific and Technical Information of China (English)

    Mirjam B Zeisel; Samira Fafi-Kremer; Isabel Fofana; Heidi Barth; Fran(c)oise Stoll-Keller; Michel Doffo(e)l; Thomas F Baumert

    2007-01-01

    Hepatitis C virus (HCV) is a major cause of hepatitis world-wide. The majority of infected individuals develop chronic hepatitis which can then progress to liver cirrhosis and hepatocellular carcinoma. Spontaneous viral clearance occurs in about 20%-30% of acutely infected individuals and results in resolution of infection without sequaelae. Both viral and host factors appear to play an important role for resolution of acute infection. A large body of evidence suggests that a strong, multispecific and long-lasting cellular immune response appears to be important for control of viral infection in acute hepatitis C. Due too the lack of convenient neutralization assays,the impact of neutralizing responses for control of viral infection had been less defined. In recent years, the development of robust tissue culture model systems for HCV entry and infection has finally allowed study of antibody-mediated neutralization and to gain further insights into viral targets of host neutralizing responses.In addition, detailed analysis of antibody-mediated neutralization in individual patients as well as cohorts with well defined viral isolates has enabled the study of neutralizing responses in the course of HCV infection and characterization of the impact of neutralizing antibodies for control of viral infection. This review will summarize recent progress in the understanding of the molecular mechanisms of antibody-mediated neutralization and its impact for HCV pathogenesis.(C) 2007 The WJG Press. All rights reserved.

  15. Greasing the SCIDs for Universal Flu Antibodies

    Science.gov (United States)

    Yewdell, Jonathan W.; Ince, William L.

    2013-01-01

    Previews In this issue, Nakamura et al. describe a robust SCID mouse-based method for isolating human monoclonal antibodies of desired specificity from adoptively transferred human B cells. As proof-of principle, they isolate human mAbs that could potentially be used to treat or prevent human infection with any influenza A virus strain. PMID:23870308

  16. Burkholderia pseudomallei Antibodies in Children, Cambodia

    OpenAIRE

    Wuthiekanun, Vanaporn; Pheaktra, Ngoun; Putchhat, Hor; Sin, Lina; Sen, Bun; Kumar, Varun; Langla, Sayan; Peacock, Sharon J.; Nicholas P. Day

    2008-01-01

    Antibodies to Burkholderia pseudomallei were detected in 16% of children in Siem Reap, Cambodia. This organism was isolated from 30% of rice paddies in the surrounding vicinity. Despite the lack of reported indigenous cases, melioidosis is likely to occur in Cambodia.

  17. IgA as therapeutic antibody

    NARCIS (Netherlands)

    Leusen, Jeanette H W

    2015-01-01

    This review is focused on the promises of IgA as a new therapeutic antibody. For more than 30 years IgG molecules have been used in the clinic in the fields of oncology, hematology, auto immune diseases and infections. However, IgA might be a good alternative, since it recruits different effector ce

  18. Antiphospholipid Antibody Syndrome Presenting with Hemichorea

    Directory of Open Access Journals (Sweden)

    Yezenash Ayalew

    2012-01-01

    Full Text Available A 25-year-old Bangladeshi lady presented to neurology with a three-month history of involuntary movements of her right arm, associated with loss of power. There was progression to the right leg, and she subsequently developed episodes of slurred speech and blurred vision. At the time of presentation, she was 12 weeks pregnant and the symptoms were reported to have started at conception. Past medical history was unremarkable apart from one first trimester miscarriage and there was no significant family history suggestive of a hereditary neurological condition. MRI of the head revealed no abnormalities but serology showed positive antinuclear antibodies (ANAs at a titre of 1/400. Further investigations revealed strongly positive anticardiolipin antibodies (>120 and positive lupus anticoagulant antibodies. The patient had a second miscarriage at 19 weeks gestation strengthening the possibility that the chorea was related to antiphospholipid antibody syndrome and she was started on a reducing dose of Prednisolone 40 mg daily and aspirin 300 mg daily. Six months later, she had complete resolution of neurological symptoms. There are several reports of chorea as a feature of antiphospholipid syndrome, but no clear consensus on underlying pathophysiology.

  19. PRODUCTION OF MONOCLONAL ANTIBODY AGAINST HUMAN IMMUNOGLOBULIN

    Directory of Open Access Journals (Sweden)

    J. Majidi

    2000-04-01

    Full Text Available Immunoglobulin E is one of the five classes of immonoglobulins that plays an important role in allergic diseases. Production of monoclonal antibodies by a single clonotype against different epitopes of immunoglobulin E has high priority in development of diagnostic kits.In this study, an attempt was made to produce monoclonal antibodies against human immunoglobulin E. Balb/c mice were immunized with semipurified immunoglobulin E and spleen cells fused with SP2.0 mouse myeloma eel! line in the presence of polyethylene glycol. Supernatant of hybridoma cells was screened for detection of antibody by enzyme linked immonosorbent assay method. Cloning of selective high absorbance wells were done with limiting dilution method. The suitable clone (monoclone was selected by enzyme linked immunosorbent assay and confirmed by immunoblot. The subclass of the chosen monoclonal antibodies was determined and the clones freezed and kept in liquid nitrogen.During this study three successful fusions were carried out, which resulted in development of 156 clones with high production of anti-IgE. Fourteen clones with the highest titres were selected for cloning. After limiting dilution more than 100 monoclonal antibodies were produced and the suitable (me (GJ0F7, i.e.; the clone which displayed the high absorbance in reaction with purified immunoglobulin E and the lowest cross-reactivity with immunoglobulin M, immunoglobulin G and immoglobulin A was chosen. In immunoblotting, presence of high density band in reaction with immunoglobulin E was confirmed. The suitable mab was shown to be IgG 1 subclass with kappa light chain. It seems that, this mab could be successfully used in diagnostic kits.

  20. The antiphospholipid antibody syndrome: a case report

    Directory of Open Access Journals (Sweden)

    Luma HN

    2012-10-01

    Full Text Available Henry Namme Luma,1,2 Marie-Solange Doualla,1,2 Elvis Temfack,1 Servais Albert Fiacre Eloumou Bagnaka,1 Emmanuella Wankie Mankaa,3 Dobgima Fofung41Department of Internal Medicine, Douala General Hospital, Douala, Cameroon; 2Faculty of Medicine and Biomedical Sciences, University of Yaoundé I, Yaoundé, Cameroon; 3Department of Radiology, Douala General Hospital Douala, Cameroon; 4Department of Abdominal Surgery, Daniel Muna Memorial Clinic, Douala, CameroonAbstract: Antiphospholipid antibody syndrome is defined by the presence of thromboembolic complications and/or pregnancy morbidity in the presence of persistently increased titers of antiphospholipid antibodies. Its clinical presentation can be diverse and any organ can be involved, with a current impact in most surgical and medical specialties. The authors present the case of a 43-year-old man who, over a 13-year period of follow-up, presented with thrombosis of the mesenteric vein, inferior vena cava, and axillary and subclavian veins in a setting where diagnostic and therapeutic options are limited and costly. Through this case report, the authors aim to describe the evolution of this complex pathology, which to date has not been described in the authors' milieu – probably because of its challenging diagnosis and the limited treatment options available. The authors conclude that clinicians need to have a high index of suspicion of APS in patients who present with a thrombotic episode – clinicians should investigate for the presence of antiphospholipid antibodies, as early diagnosis may influence the course of the disease. Furthermore, resources for the detection of antiphospholipid antibodies should be made readily available in resource-limited settings. Finally, patient education on the importance of drug compliance, periodic monitoring, and prevention of thrombosis is indispensable, especially as mortality could be associated with the effects of vascular thrombosis and/or the effects

  1. B cells contribute to MS pathogenesis through antibody-dependent and antibody-independent mechanisms

    Directory of Open Access Journals (Sweden)

    Wilson HL

    2012-05-01

    Full Text Available Heather L Wilson1,21Vaccine and Infectious Disease Organization-International Vaccine Center, 2Department of Biochemistry, University of Saskatchewan, Saskatoon, Saskatchewan, CanadaAbstract: For many years, central dogma defined multiple sclerosis (MS as a T cell-driven autoimmune disorder; however, over the past decade there has been a burgeoning recognition that B cells contribute to the pathogenesis of certain MS disease subtypes. B cells may contribute to MS pathogenesis through production of autoantibodies (or antibodies directed at foreign bodies, which unfortunately cross-react with self-antigens, through promotion of T cell activation via antigen presentation, or through production of cytokines. This review highlights evidence for antibody-dependent and antibody-independent B cell involvement in MS pathogenesis.Keywords: autoantibodies, antibody targets, clinically isolated MS, primary progressive MS, secondary progressive MS, relapsing and remitting MS, T cells, T regulatory cells

  2. Antibodies to poliovirus detected by immunoradiometric assay with a monoclonal antibody

    Energy Technology Data Exchange (ETDEWEB)

    Spitz, M.; Fossati, C.A.; Schild, G.C.; Spitz, L.; Brasher, M. (National Inst. for Biological Standards and Control, London (UK))

    1982-10-01

    An immunoradiometric assay (IRMA) for the assay of antibodies to poliovirus antigens is described. Dilutions of the test sera or whole (finger prick) blood samples were incubated with the poliovirus antigen bound to a solid phase and the specific antibody was detected by the addition of a mouse anti-human IgG monoclonal antibody (McAb), which was itself revealed by iodinated sheep IgG antimouse F(ab). The authors have shown that this technique is suitable for the estimation of IgG anti-poliovirus antibodies induced in children following polio vaccine. The present study shows that SPRIA provides a simple and inexpensive method for serological studies with poliovirus particularly for use in large-scale surveys.

  3. High-Throughput Tools for Characterization of Antibody Epitopes

    DEFF Research Database (Denmark)

    Christiansen, Anders

    Antibodies are molecules of tremendous importance. In their primary role, they protect our bodies against disease. However, in recent decades, scientists have harnessed the binding capabilities of antibodies and have applied them widely in research, diagnostics and therapeutics. Consequently, it ...

  4. Antibody avidity in swine lymphocyte antigen-defined miniature pigs.

    Science.gov (United States)

    Appleyard, G D; Mallard, B A; Kennedy, B W; Wilkie, B N

    1992-01-01

    Antibody avidity to hen egg white lysozyme (HEWL) was measured by thiocyanate ion elution enzyme-linked immunosorbent assay (ELISA) in swine lymphocyte antigen (SLA) defined miniature pigs. Serum antibody avidity was evaluated on day 14 and 30 after primary (day 0) and secondary (day 14) immunizations in eight to ten week old miniature pigs previously typed for swine lymphocyte antigen genotype. The effect of SLA genotype, litter, and gender on anti-HEWL antibody avidity was determined by least squares. Antibody avidity varied amongst individuals. Antibody avidity maturation was observed as a mean rise in antibody avidity from primary response (0.89 +/- 0.64) to secondary response (1.23 +/- 0.54) (p < 0.0005). Overall, SLA genotype did not significantly influence antibody avidity or avidity maturation, but pigs of dd genotype had greater avidity maturation between primary and secondary responses than other genotypes. Litter effects significantly affected antibody avidity and maturation. PMID:1477799

  5. Tetanus Neurotoxin Neutralizing Antibodies Screened from a Human Immune scFv Antibody Phage Display Library.

    Science.gov (United States)

    Wang, Han; Yu, Rui; Fang, Ting; Yu, Ting; Chi, Xiangyang; Zhang, Xiaopeng; Liu, Shuling; Fu, Ling; Yu, Changming; Chen, Wei

    2016-01-01

    Tetanus neurotoxin (TeNT) produced by Clostridium tetani is one of the most poisonous protein substances. Neutralizing antibodies against TeNT can effectively prevent and cure toxicosis. Using purified Hc fragments of TeNT (TeNT-Hc) as an antigen, three specific neutralizing antibody clones recognizing different epitopes were selected from a human immune scFv antibody phage display library. The three antibodies (2-7G, 2-2D, and S-4-7H) can effectively inhibit the binding between TeNT-Hc and differentiated PC-12 cells in vitro. Moreover, 2-7G inhibited TeNT-Hc binding to the receptor via carbohydrate-binding sites of the W pocket while 2-2D and S-4-7H inhibited binding of the R pocket. Although no single mAb completely protected mice from the toxin, they could both prolong survival when challenged with 20 LD50s (50% of the lethal dose) of TeNT. When used together, the mAbs completely neutralized 1000 LD50s/mg Ab, indicating their high neutralizing potency in vivo. Antibodies recognizing different carbohydrate-binding pockets could have higher synergistic toxin neutralization activities than those that recognize the same pockets. These results could lead to further production of neutralizing antibody drugs against TeNT and indicate that using TeNT-Hc as an antigen for screening human antibodies for TeNT intoxication therapy from human immune antibody library was convenient and effective. PMID:27626445

  6. Quality of histone modification antibodies undermines chromatin biology research

    OpenAIRE

    Goran Kungulovski; Albert Jeltsch

    2015-01-01

    Histone post-translational modification (PTM) antibodies are essential research reagents in chromatin biology. However, they suffer from variable properties and insufficient documentation of quality. Antibody manufacturers and vendors should provide detailed lot-specific documentation of quality, rendering further quality checks by end-customers unnecessary. A shift from polyclonal antibodies towards sustainable reagents like monoclonal or recombinant antibodies or histone binding domains wou...

  7. Anti-Cardiolipin Antibody in Acute Myocardial Infarction

    OpenAIRE

    Abdolreza S. Jahromi; Mohammad Shojaie; Samira Dana; Abdoulhossain Madani

    2010-01-01

    Problem statement: Myocardial infarction is the combined result of environmental and personal factors. Data concerning the relation between anti-Phospholipid (aPL) antibodies and myocardial infarction in subjects without evidence of overt autoimmune disease are conflicting. Anticardiolipin antibody is detected in various diseases like rheumatoid arthritis, systemic lupus erythematosus and anti-phospholipid antibody syndrome. The study of Anticardiolipin antibody in Acute Myocardial Infarction...

  8. ANTI-PHOSPHATIDYLSERINE ANTIBODIES IN ACUTE MYOCARDIAL INFARCTION

    OpenAIRE

    Abdolreza Sotoodeh Jahromi; Mohammad Shojaei; Mohammad Reza Farjam; Abdolhossien Madani

    2013-01-01

    Acute Myocardial Infarction (AMI) is the combined result of environmental factors and personal predispositions. Many factors play a role in AMI including anti-Phospholipid (aPL) antibodies, that may act in the induction of immunological response leading to the development of AMI. Anti-Phosphatidylserine (PS) antibody is detected in various diseases like rheumatoid arthritis, systemic lupus erythematosus and anti-phospholipid antibody syndrome. The study of anti-PS antibody in AMI might shed l...

  9. Evaluation of Salivary Antibodies to Detect Infection with Helicobacter pylori

    OpenAIRE

    1997-01-01

    Helicobacter pylori infection is an important cause of peptic ulcer disease and chronic gastritis. Infection with this bacterium stimulates the production of immunoglobulin (Ig) G antibody. Salivary IgG antibody tests to detect H pylori infection offer a convenient and noninvasive method of diagnosis. To evaluate an IgG salivary antibody kit, saliva was collected from 157 out-patients with dyspepsia referred for endoscopy to a tertiary centre. A salivary IgG ELISA antibody assay was performed...

  10. Cell-Free Synthesis Meets Antibody Production: A Review

    OpenAIRE

    Marlitt Stech; Stefan Kubick

    2015-01-01

    Engineered antibodies are key players in therapy, diagnostics and research. In addition to full size immunoglobulin gamma (IgG) molecules, smaller formats of recombinant antibodies, such as single-chain variable fragments (scFv) and antigen binding fragments (Fab), have emerged as promising alternatives since they possess different advantageous properties. Cell-based production technologies of antibodies and antibody fragments are well-established, allowing researchers to design and manufactu...

  11. Production and characterization of antibody against aflatoxin Q1.

    OpenAIRE

    Fan, T. S.; Zhang, G S; Chu, F. S.

    1984-01-01

    Antibodies against aflatoxin Q1 (AFQ1) were obtained from rabbits after immunization of either AFQ1-hemisuccinate or AFQ2a conjugated to bovine serum albumin. Both radioimmunoassay and enzyme-linked immunosorbent assaY (ELISA) were used for the determination of antibody titers and specificities. Antibodies obtained from rabbits after immunization with AFQ1-hemisuccinate-bovine serum albumin had the highest affinity to aflatoxin B1, whereas antibodies obtained from rabbits after immunization w...

  12. Development of syngeneic monoclonal anti-idiotype antibodies to mouse monoclonal anti-asialoglycoprotein receptor antibody.

    Directory of Open Access Journals (Sweden)

    Hirai M

    2002-06-01

    Full Text Available Anti-idiotype antibodies (Ab2 play an important role in the homeostasis of immune responses and are related to the development and the disease activity of certain autoimmune diseases. The asialoglycoprotein receptor (ASGPR is considered one of the target antigens in the pathogenesis of autoimmune chronic active hepatitis (AIH. We previously developed a mouse monoclonal antibody (clone 8D7 which recognizes rat and human ASGPR. In this study, to help investigate the anti-ASGPR antibody-anti-idiotype antibody network in patients with AIH, we developed a syngeneic mouse monoclonal Ab2 to the 8D7 anti-ASGPR antibody (Ab1. One clone, designated as 3C8, tested positive for specific reactivity to 8D7-Ab1 and did not bind to other irrelevant immunoglobulins. By competitive inhibition assays, the binding of 8D7-Ab1 to liver membrane extracts, i.e., the crude antigen preparation, was inhibited by 3C8-Ab2 in a dose-dependent manner, and the binding of 8D7-Ab1 to 3C8-Ab2 was inhibited by the liver membrane extracts. In the immunohistochemical analysis, 3C8-Ab2 blocked the specific staining of sinusoidal margins of rat hepatocytes by 8D7-Ab1. These results suggest that 3C8 anti-idiotype antibody recognizes the specific idiotypic determinants within the antigen-binding site of 8D7-Ab1.

  13. Development of monoclonal antibodies suitable for rabies virus antibody and antigen detection.

    Science.gov (United States)

    Chander, Vishal; Singh, R P; Verma, P C

    2012-12-01

    The control of an infectious viral disease as rabies is made easier by rapid and accurate diagnosis. Successful rabies prophylaxis is dependent upon the active immunization with vaccine along with passive administration of rabies virus neutralizing antibodies which together clear the virus before widespread infection of central nervous system occurs. The present study aimed at the development of monoclonal antibodies (MAbs) suitable for rabies virus antibody and antigen detection. For the production of rabies specific MAbs, immunization of Swiss albino mice with a commercially available vaccine was done and Polyethylene glycol mediated fusion of spleenocytes with myeloma cells was performed. The positive clones were selected on the basis of distinct reactivity by cell Enzyme linked immunosorbent assay and fluorescence in Indirect Fluorescent antibody test. The positive clones obtained were subjected to single cell cloning by limiting dilution method. The reactive clones were further titrated and employed for virus titration and virus neutralization. The neutralizing activity was evaluated using Fluorescence Activated Cell Sorter technique. Three MAb clones showed a distinct percent inhibition in the presence of positive serum. One of the MAb clone No. 5C3 was relatively more specific in detecting rabies antibodies and also found suitable for competitive ELISA to assess the antibody level in vaccinated subjects.

  14. Development of monoclonal antibodies suitable for rabies virus antibody and antigen detection.

    Science.gov (United States)

    Chander, Vishal; Singh, R P; Verma, P C

    2012-12-01

    The control of an infectious viral disease as rabies is made easier by rapid and accurate diagnosis. Successful rabies prophylaxis is dependent upon the active immunization with vaccine along with passive administration of rabies virus neutralizing antibodies which together clear the virus before widespread infection of central nervous system occurs. The present study aimed at the development of monoclonal antibodies (MAbs) suitable for rabies virus antibody and antigen detection. For the production of rabies specific MAbs, immunization of Swiss albino mice with a commercially available vaccine was done and Polyethylene glycol mediated fusion of spleenocytes with myeloma cells was performed. The positive clones were selected on the basis of distinct reactivity by cell Enzyme linked immunosorbent assay and fluorescence in Indirect Fluorescent antibody test. The positive clones obtained were subjected to single cell cloning by limiting dilution method. The reactive clones were further titrated and employed for virus titration and virus neutralization. The neutralizing activity was evaluated using Fluorescence Activated Cell Sorter technique. Three MAb clones showed a distinct percent inhibition in the presence of positive serum. One of the MAb clone No. 5C3 was relatively more specific in detecting rabies antibodies and also found suitable for competitive ELISA to assess the antibody level in vaccinated subjects. PMID:24293819

  15. Detection of antitrophoblast antibodies in the sera of patients with anticardiolipin antibodies and fetal loss.

    Science.gov (United States)

    McCrae, K R; DeMichele, A M; Pandhi, P; Balsai, M J; Samuels, P; Graham, C; Lala, P K; Cines, D B

    1993-11-01

    Women with anticardiolipin antibodies (ACLA) are at increased risk for fetal loss. One potential explanation for this outcome is that sera from these individuals contain antibodies reactive with trophoblast cells, which are involved in the establishment of the uteroplacental vasculature and maintenance of placental blood fluidity. To examine this hypothesis, we compared the incidence of trophoblast-reactive antibodies in 27 patients with ACLA and a history of fetal loss with that in 29 normal pregnant women. Sera from 20 patients, but only one control, contained trophoblast-reactive antibodies (P trophoblast cells. In most cases, sera from which ACLA were adsorbed by cardiolipin-containing liposomes maintained reactivity against cells. In addition, patient Ig fractions immunoprecipitated an approximately 62-kD protein from the trophoblast cell surface, stimulated the release of arachidonic acid and thromboxane A2 by trophoblasts, and inhibited the binding of prourokinase to trophoblast urokinase receptors. These observations show that sera from women with ACLA and a history of fetal loss contain antitrophoblast antibodies. These antibodies may be serologically distinct from ACLA, and may contribute to the pathogenesis of fetal demise. PMID:7693045

  16. Behavioral and Psychological Responses to HIV Antibody Testing.

    Science.gov (United States)

    Jacobsen, Paul B.; And Others

    1990-01-01

    Considers effects of informing individuals of their antibody status as determined by human immunodeficiency virus (HIV) antibody testing. Reviews research examining changes in psychological distress and in behaviors associated with HIV infections among individuals who have undergone antibody testing. Identifies methodological issues in studying…

  17. Donor non-specific MICA antibodies in renal transplant recipients.

    Science.gov (United States)

    Sapák, Michal; Chreňová, Silvia; Tirpáková, Jana; Žilinská, Zuzana; Ďurmanová, Vladimíra; Shawkatová, Ivana; Jakuš, Vladimír; Kuba, Daniel; Buc, Milan

    2014-02-01

    Despite recent advances in solid organ transplantations, an antibody mediated rejection caused by donor specific antibodies is still a major problem in kidney graft survival. Besides HLA-induced humoral response, antibodies against MICA antigens have recently attracted attention because of their possible role in graft rejection. The aim of our study was to establish whether renal recipients produce antibodies against MICA molecules due to the transplantation and if they are specific for MICA antigens of the donors. MICA antibody screening was performed in 124 kidney recipient sera. 22 sera, that were found to be MICA antibody positive, were further examined for MICA antibody profiles and compared with donor MICA alleles. The analysis of MICA antibody positive sera showed mostly more complex reactivity patterns. A significant fraction of patient sera (59%) reacted not only with the donor MICA antigens, but also with other MICA patterns. A match between antibody specificities and MICA antigens was observed in 41% of renal recipients only. On the other hand, as much as in 36% of recipient sera were detected antibodies against their own MICA molecules. We did not prove a complete correlation between the recipient MICA antibody specificities and MICA antigens of the donor. We assume that MICA antibody induction occurs not only due to the allogeneic stimulation itself but also due to other factors that need to be elucidated.

  18. Production and characterization of monoclonal antibodies against mink leukocytes

    DEFF Research Database (Denmark)

    Chen, W.S.; Pedersen, Mikael; Gram-Nielsen, S.;

    1997-01-01

    Three monoclonal antibodies (mAbs) were generated against mink leukocytes. One antibody reacted with all T lymphocytes, one with all monocytes and one had platelet reactivity. Under reducing conditions, the T lymphocyte reactive antibody immunoprecipitated 18 kDa, 23 kDa, 25 kDa and 32-40 kDa pol...

  19. Necrotizing and Crescentic Lupus Nephritis with Antineutrophil Cytoplasmic Antibody Seropositivity

    OpenAIRE

    Nasr, Samih H.; D'Agati, Vivette D; Park, Hye-Ran; Sterman, Paul L.; Goyzueta, Juan D.; Dressler, Robert M.; Hazlett, Shawn M.; Pursell, Robert N.; Caputo, Christopher; Markowitz, Glen S.

    2008-01-01

    Background and objectives: Lupus nephritis is a classic immune complex glomerulonephritis. In contrast, antineutrophil cytoplasmic antibodies are associated with necrotizing and crescentic glomerulonephritis, in the absence of significant immune deposits. Antineutrophil cytoplasmic antibodies are detected by indirect immunofluorescence in 20% of patients with systemic lupus erythematosus. We report 10 cases of necrotizing and crescentic lupus nephritis with antineutrophil cytoplasmic antibody...

  20. Antibody Based Surgical Imaging and Photodynamic Therapy for Cancer

    NARCIS (Netherlands)

    de Boer, Esther

    2016-01-01

    In 1944 Albert Coons was the first to show that a fluorescent molecule could be conjugated directly to an antibody made against a target site of interest. This binding does not affect antibody specificity so that labeled antibodies can be used to visualize the location and distribution of the target

  1. Serum Antibody Response to Clostridium botulinum Toxin in Infant Botulism

    OpenAIRE

    Rubin, Lorry G.; Dezfulian, Manuchehr; Yolken, Robert H.

    1982-01-01

    A serum antibody response has not been previously demonstrated after infection with Clostridium botulinum. We developed an enzyme immunoassay for measuring serum antibody to C. botulinum toxins A, B, and E. This assay system detected a specific immunoglobulin G and immunoglobulin M antibody response to C. botulinum toxin in two patients with infant botulism.

  2. Assay for the specificity of monoclonal antibodies in crossed immunoelectrophoresis

    DEFF Research Database (Denmark)

    Skjødt, K; Schou, C; Koch, C

    1984-01-01

    A method is described based on crossed immunoelectrophoresis of a complex antigen mixture in agarose gel followed by incubation of the gel with the monoclonal antibody. The bound monoclonal antibody is detected by the use of a secondary enzyme-labelled antibody. Using this technique we have been ...

  3. [Reactivity of antibodies to collagen types I to IV and antibodies to chondroitin sulfate in the spleen].

    Science.gov (United States)

    Galbavý, S; Ruzicková, M; Surmíková, E; Danihel, L; Porubský, J; Papincák, J; Holesa, S; Trnka, J

    1996-02-01

    Antibodies to collagen type I and III reacted negatively, antibodies to collagen type IV positively with reticulin, trabeculae and circumferent reticulum of lymphatic sheaths, poorly positively with capsula, strongly positively with subcapsular zone. Antibodies to collagen type II reacted positively with capsula, poorly with subcapsular zone, strongly with sinus wall and poorly with trabeculae. They did not react with circumferent reticulum of periarterial lymphoid sheaths. Antibodies to collagen type II and IV reacted positively with central arteries. Antibodies to chondroitinsulphate C reacted poorly and antibodies to chondroitinsulphate B strongly positively with sinus walls and oval cells spread in the white and red pulpa. Antibodies to chondroitin sulphate A reacted similarly as antibodies to chondroitinsulphate B. PMID:9560890

  4. Preparation and testing of a Haemophilus influenzae Type b/Hepatitis B surface antigen conjugate vaccine.

    Science.gov (United States)

    An, So Jung; Woo, Joo Sung; Chae, Myung Hwa; Kothari, Sudeep; Carbis, Rodney

    2015-03-24

    The majority of conjugate vaccines focus on inducing an antibody response to the polysaccharide antigen and the carrier protein is present primarily to induce a T-cell dependent response. In this study conjugates consisting of poly(ribosylribitolphosphate) (PRP) purified from Haemophilus influenzae Type b bound to Hepatitis B virus surface antigen (HBsAg) virus like particles were prepared with the aim of inducing an antibody response to not only the PRP but also the HBsAg. A conjugate consisting of PRP bound to HBsAg via an adipic acid dihydrazide (ADH) spacer induced strong IgG antibodies to both the PRP and HBsAg. When conjugation was performed without the ADH spacer the induction of an anti-PRP response was equivalent to that seen by conjugate with the ADH spacer, however, a negligible anti-HBsAg response was induced. For comparison, PRP was conjugated to diphtheria toxoid (DT) and Vi polysaccharide purified from Salmonella Typhi conjugated to HBsAg both using an ADH spacer. The PRPAH-DT conjugate induced strong anti-PRP and anti-DT responses, the Vi-AHHBsAg conjugate induced a good anti-HBsAg response but not as strong as that induced by the PRPAH-HBsAg conjugate. This study demonstrated that in mice it was possible to induce robust antibody responses to both polysaccharide and carrier protein provided the conjugate has certain physico-chemical properties. A PRPAH-HBsAg conjugate with the capacity to induce anti-PRP and anti-HBsAg responses could be incorporated into a multivalent pediatric vaccine and simplify formulation of such a vaccine.

  5. A monoclonal antibody toolkit for C. elegans.

    Directory of Open Access Journals (Sweden)

    Gayla Hadwiger

    Full Text Available BACKGROUND: Antibodies are critical tools in many avenues of biological research. Though antibodies can be produced in the research laboratory setting, most research labs working with vertebrates avail themselves of the wide array of commercially available reagents. By contrast, few such reagents are available for work with model organisms. METHODOLOGY/PRINCIPAL FINDINGS: We report the production of monoclonal antibodies directed against a wide range of proteins that label specific subcellular and cellular components, and macromolecular complexes. Antibodies were made to synaptobrevin (SNB-1, a component of synaptic vesicles; to Rim (UNC-10, a protein localized to synaptic active zones; to transforming acidic coiled-coil protein (TAC-1, a component of centrosomes; to CENP-C (HCP-4, which in worms labels the entire length of their holocentric chromosomes; to ORC2 (ORC-2, a subunit of the DNA origin replication complex; to the nucleolar phosphoprotein NOPP140 (DAO-5; to the nuclear envelope protein lamin (LMN-1; to EHD1 (RME-1 a marker for recycling endosomes; to caveolin (CAV-1, a marker for caveolae; to the cytochrome P450 (CYP-33E1, a resident of the endoplasmic reticulum; to beta-1,3-glucuronyltransferase (SQV-8 that labels the Golgi; to a chaperonin (HSP-60 targeted to mitochondria; to LAMP (LMP-1, a resident protein of lysosomes; to the alpha subunit of the 20S subcomplex (PAS-7 of the 26S proteasome; to dynamin (DYN-1 and to the alpha-subunit of the adaptor complex 2 (APA-2 as markers for sites of clathrin-mediated endocytosis; to the MAGUK, protein disks large (DLG-1 and cadherin (HMR-1, both of which label adherens junctions; to a cytoskeletal linker of the ezrin-radixin-moesin family (ERM-1, which localized to apical membranes; to an ERBIN family protein (LET-413 which localizes to the basolateral membrane of epithelial cells and to an adhesion molecule (SAX-7 which localizes to the plasma membrane at cell-cell contacts. In addition to

  6. Comparisons of the effect of naturally acquired maternal pertussis antibodies and antenatal vaccination induced maternal tetanus antibodies on infant's antibody secreting lymphocyte responses and circulating plasma antibody levels.

    Science.gov (United States)

    Ahmad, Shaikh Meshbahuddin; Alam, Jahangir; Afsar, Nure Alam; Huda, Nazmul; Kabir, Yearul; Qadri, Firdausi; Raqib, Rubhana; Stephensen, Charles B

    2016-04-01

    The goal of this study was to explore the effects of trans-placental tetanus toxoid (TT) and pertussis (PT) antibodies on an infant's response to vaccination in the context of antenatal immunization with tetanus but not with pertussis. 38 mothers received a single dose of TT vaccine during pregnancy. Infants received tetanus and pertussis vaccines at 6, 10 and 14 wk of age. TT and PT anti-IgG secretion by infant lymphocytes was measured at 15 wk. Plasma antibodies were measured at 6 wk (pre-vaccination), 15 wk and 1 y of age. Prior to vaccination, TT and PT antibody were detected in 94.6% and 15.2% of infants. At 15 wk anti-TT-IgG and anti-PT-IgG in plasma was increased by 7-9 fold over pre-vaccination levels, while at 1 y plasma anti-TT-IgG was decreased by approximately 5-fold from the peak and had returned to near the pre-vaccination level. At 1 y plasma anti-PT-IgG was decreased by 2-fold 1 yfrom the 15 wk level. However, 89.5% and 82.3% of infants at 1 y had protective levels of anti-TT and anti-PT IgG, respectively. Pre-vaccination plasma IgG levels were associated with lower vaccine-specific IgG secretion by infant lymphocytes at 15 wk (p < 0.10). This apparent inhibition was seen for anti-TT-IgG at both 15 wk (p < 0.05) and t 1 y (p < 0.10) of age. In summary, we report an apparent inhibitory effect of passively derived maternal antibody on an infants' own antibody response to the same vaccine. However, since the cut-off values for protective titers are low, infants had protective antibody levels throughout infancy. PMID:27176823

  7. Hybridization-based antibody cDNA recovery for the production of recombinant antibodies identified by repertoire sequencing

    OpenAIRE

    Valdés-Alemán, Javier; Téllez-Sosa, Juan; Ovilla-Muñoz, Marbella; Godoy-Lozano, Elizabeth; Velázquez-Ramírez, Daniel; Valdovinos-Torres, Humberto; Gómez-Barreto, Rosa E; Martinez-Barnetche, Jesús

    2013-01-01

    High-throughput sequencing of the antibody repertoire is enabling a thorough analysis of B cell diversity and clonal selection, which may improve the novel antibody discovery process. Theoretically, an adequate bioinformatic analysis could allow identification of candidate antigen-specific antibodies, requiring their recombinant production for experimental validation of their specificity. Gene synthesis is commonly used for the generation of recombinant antibodies identified in silico. Novel ...

  8. Transfusion management of patients with red blood cell antibodies

    Directory of Open Access Journals (Sweden)

    Bujandrić Nevenka B.

    2013-01-01

    Full Text Available Introduction. Red blood cell antibodies may cause a positive result of pre-transfusion blood compatibility testing (crossmatch test. It can be a problem to provide suitable blood units for patients with clinically significant antibodies to high-frequency antigens as well as for those with multiple alloantibody specificities. This study was aimed at identifying transfused patients in the population of South-Backa who had developed clinically significant red blood cell alloantibodies. Material and methods. We analyzed the records of crossmatch results and antibody screening performed at the Blood Transfusion Institute of Vojvodina during 2012. Results. Antibodies were found in 103 patients: A 63 patients with single antibodies: 1 16 with antibodies of unknown specificity (3 autoantibodies, 13 alloantibodies; 2 39 with clinically significant antibodies (23 from Rh system (2 anti-C, 2 anti-D, 12 anti-E, 7 anti-c, 4 anti-K, 3 anti-Fya, 7 anti-Jka, 2 anti-S; 3 8 with usually not significant antibodies (6 anti-M, 1 anti-A1, 1 anti- Cw; B 40 patients developed multiple antibodies: 1 all patients had at least one clinically significant antibody from various blood group system (44 Rh, 13 Kell, 7 Kidd, 7 MNSs (S, s; 2 3 patients had usually not significant antibodies (1 Lewis, 2 Lutheran; 3 3 patients occasionally had clinically significant antibody (3 anti- Yta; 4 3 patients had antibodies of unknown specificity (2 autoantibodies, 1alloantibody. Antibodies detected in the majority of patients (65-63.1% had a specificity of Rh and/or the Kell system. Conclusions. The main goal of pre-transfusion blood compatibility testing is to detect clinically significant antibodies. The provision of antigen negative blood units for those patients is a special challenge for blood establishments. Database with a sufficient number of typed blood donors can help to resolve this problem.

  9. A MONOCLONAL-ANTIBODY AGAINST HUMAN BETA-GLUCURONIDASE FOR APPLICATION IN ANTIBODY-DIRECTED ENZYME PRODRUG THERAPY

    NARCIS (Netherlands)

    Haisma, Hidde; VANMUIJEN, M; SCHEFFER, G; SCHEPER, RJ; PINEDO, HM; BOVEN, E

    1995-01-01

    The selectivity of anticancer agents may be improved by antibody-directed enzyme prodrug therapy (ADEPT), The immunogenicity of antibody-enzyme conjugates and the low tumor to normal tissue ratio calls for the use of a human enzyme and the development of a monoclonal antibody (MAb) against that enzy

  10. High throughput screening for antibody induced complement-dependent cytotoxicity in early antibody discovery using homogeneous macroconfocal fluorescence imaging

    NARCIS (Netherlands)

    Gerritsen, Arnout F.; Bosch, Martijn; de Weers, Michel; van de Winkel, Jan G. J.; Parren, Paul W. H. I.

    2010-01-01

    Complement-dependent cytotoxicity (CDC) represents an important Fc-mediated effector function of antibodies and is a quality often sought in candidates for therapeutic antibody development in cancer. Antibodies inducing potent CDC are relatively rare as the ability to induce CDC is strongly dependen

  11. Molecular aspects of antibody-antigen interactions : size reduction of a herpes simplex virus neutralizing antibody and its antigen

    NARCIS (Netherlands)

    Schellekens, Gerardus Antonius

    1996-01-01

    Antibody molecules, produced as a response against foreign substances, interact with their antigen in a very specific manner. Antibodies with a predetermined specificity (monoclonal antibodies) can be produced and are widely used in medicine and science as indicator molecules. Genetic engineering of

  12. Monoclonal antibodies against rabbit mammary prolactin receptors. Specific antibodies to the hormone binding domain

    International Nuclear Information System (INIS)

    Three monoclonal antibodies (M110, A82, and A917) were obtained by fusing myeloma cells and spleen cells from mice immunized with partially purified rabbit mammary gland prolactin (PRL) receptors. All 3 antibodies were capable of complete inhibition of 125I-ovine prolactin (oPRL) binding to rabbit mammary PRL receptors in either particulate or soluble form. M110 showed slightly greater potency than oPRL in competing for 125I-oPRL binding. These antibodies also inhibited PRL binding to microsomal fractions from rabbit liver, kidney, adrenal, ovary, and pig mammary gland, although A82 showed poor inhibition in pig mammary gland. There was no cross-reaction of any of the 3 monoclonal antibodies (mAbs) for the other species tested: human (T-47D breast cancer cells) and rat (liver, ovary). In order to confirm that these antibodies are specific to the binding domain, antibodies were purified, iodinated, and binding characteristics were investigated. 125I-M110 and 125I-A82 binding was completely inhibited by lactogenic hormones, whereas nonlactogenic hormones did not cross-react. Competition of 125I-M110 by oPRL was comparable to that of 125I-oPRL by unlabeled oPRL, while 125I-A917 binding was only partially competed (30-60%) by lactogenic hormones. Tissue and species specificity of labeled antibody binding paralleled results of binding inhibition experiments using 125I-oPRL. In addition, A82 and A917 completely inhibited 125I-M110 binding. In contrast, 125I-A82 binding was stimulated by A917 and 125I-A917 binding was stimulated by A82

  13. ON THE NOTION OF SYNERGY OF MONOCLONAL ANTIBODIES AS DRUGS

    Directory of Open Access Journals (Sweden)

    Michael Sela

    2013-08-01

    Full Text Available History of developing synergy between monoclonal antibodies, anti-tumor activity of monoclonal antibodies against tyrosine-kinases receptors EGFR/ErbB-1 and HER2/ErbB-2 as well as growth factor VEGF in various combinations are considered in the article. There were proposed hypotheses about potential molecular mechanisms underlay synergy between monoclonal antibodies (for homo- and hetero combinations of antibodies appropriately specific for antigenic determinants on the same or different receptors. Future trends in researches necessary to deeper understanding causes of this phenomenon and perspectives for practical application of monoclonal antibodies acted synergistically as immunotherapeutic drugs for human tumors treatment are reviewed.

  14. Antissaliva Antibodies of Lutzomyia Longipalpis in area of Visceral Leishmaniasis.

    Science.gov (United States)

    Fraga, Thiago Leite; Fernandes, Magda Freitas; Pontes, Elenir Rose Jardim Cury; Levay, Ana Paula Silva; Almeida da Cunha, Elenice Brandão; França, Adriana de Oliveira; Dorval, Maria Elizabeth Cavalheiros

    2016-07-01

    The aim of the present study was to assess the presence of antissaliva antibodies of Lutzomyia longipalpis in human hosts living in area of visceral leishmaniasis, located in the Center-West region of Brazil. The presence of antissaliva antibodies of L. longipalpis exhibited a strong correlation with the protection and development of antibodies against Leishmania sp. Of the 492 children studied, elevated antissaliva antibodies of L. longipalpis were detected in 38.4% of the participants. There was a higher percentage of positivity (64.7%) among children who exhibited anti-Leishmania sp. antibodies and among those who were positive in the delayed hypersensitivity test (34.8%). PMID:27093167

  15. Antissaliva Antibodies of Lutzomyia Longipalpis in area of Visceral Leishmaniasis.

    Science.gov (United States)

    Fraga, Thiago Leite; Fernandes, Magda Freitas; Pontes, Elenir Rose Jardim Cury; Levay, Ana Paula Silva; Almeida da Cunha, Elenice Brandão; França, Adriana de Oliveira; Dorval, Maria Elizabeth Cavalheiros

    2016-07-01

    The aim of the present study was to assess the presence of antissaliva antibodies of Lutzomyia longipalpis in human hosts living in area of visceral leishmaniasis, located in the Center-West region of Brazil. The presence of antissaliva antibodies of L. longipalpis exhibited a strong correlation with the protection and development of antibodies against Leishmania sp. Of the 492 children studied, elevated antissaliva antibodies of L. longipalpis were detected in 38.4% of the participants. There was a higher percentage of positivity (64.7%) among children who exhibited anti-Leishmania sp. antibodies and among those who were positive in the delayed hypersensitivity test (34.8%).

  16. Antiphospholipid antibody syndrome presenting as transverse myelitis

    Directory of Open Access Journals (Sweden)

    Javvid M Dandroo

    2015-01-01

    Full Text Available The antiphospholipid syndrome (APS is characterized by arterial and/or venous thrombosis and pregnancy morbidity in the presence of anticardiolipin antibodies and/or lupus anticoagulant. APS can occur either as a primary disorder or secondary to a connective tissue disease, most frequently systemic lupus erythematosus. Central nervous system involvement is one of the most prominent clinical manifestations of APS, and includes arterial and venous thrombotic events, psychiatric features, and a variety of other nonthrombotic neurological syndromes. Although the mechanism of neurological involvement in patients with APS is thought to be thrombotic in origin and endothelial dysfunction associated with antiphospholipid antibodies. APS presenting as acute transverse myelitis is very rarely seen with a prevalence rate of 1%. We are describing a foreigner female presenting as acute transverse myelitis which on evaluation proved to be APS induced. So far, very few cases have been reported in literature with APS as etiology.

  17. Monoclonal Antibodies to Plant Growth Regulators

    Science.gov (United States)

    Eberle, Joachim; Arnscheidt, Angelika; Klix, Dieter; Weiler, Elmar W.

    1986-01-01

    Four high affinity monoclonal antibodies, which recognize two plant growth regulators from the cytokinin group, namely trans-zeatin riboside and dihydrozeatin riboside and their derivatives are reported. Six hybridomas were produced from three independent fusions of Balb/c spleen cells with P3-NS1-Ag 4-1 (abbreviated NS1) or X63-Ag 8.653 (X63) myeloma cells. The mice had been hyperimmunized with zeatin riboside-bovine serum albumin conjugate or dihydrozeatin riboside-bovine serum albumin conjugate for 3 months. The hybridomas secrete antibodies of the IgG 1 or IgG 2b subclass and allow the detection of femtomole amounts of the free cytokinins, their ribosides, and ribotides in plant extracts. The use of these monoclonals in radio- and enzyme-linked immunosorbent assay is also discussed. PMID:16664848

  18. Poliarterite nodosa due to anti elastase antibody

    Directory of Open Access Journals (Sweden)

    Caterina Defendenti

    2008-09-01

    Full Text Available The Authors related one case of polyarteritis nodosa occurred to a men forty eight years old.The clinical was characterized by mesenteric and femoral arteries occlusion and chronic cutaneous ulcers to legs. There were bioptical aspects of systemic vasculitis with necrotizing inflammation and a paucity of immune deposit. It was effective oral cyclophosphamide plus steroids. This disease was closely associated with antibodies anti elastase (HLE.The patient had not a history of cocaine abuse or LES disease but the nucleolar pattern ANA was positive >1:640 (anti-nDNA negative. Similar case ANA positive associated with the anti-elastase antibodies, was described by Nassberger (Lancet 1989 for 6/104 patients with LES, anti-nDNA negative. The patient with the highest anti-elastase concentration subsequentely died after very rapid development of severe brain and kidney involvement.

  19. Hepatitis C Virus Antibodies and Vitiligo Disease

    Directory of Open Access Journals (Sweden)

    Z Jadali

    2005-06-01

    Full Text Available Vitiligo is a common skin disorder, characterized by depigmented patches due to selective destruction of melanocytes. The etiology of this disease is unknown. A number of hypotheses including viral theory have been proposed to explain the etiology. To determine the prevalence of antibody to hepatitis C virus infection in vitiligo patients, the present study was performed. Third generation ELISA test was used for detection of antibodies to HCV in human sera. All normal controls were anti-HCV negative whereas only one patient was positive for anti-HCV and there was no significant difference in the prevalence of anti-HCV between patients and controls. These results indicate that hepatitis C virus has not a direct causal role in the pathogenesis of vitiligo, however, this does not rul out a "hit and run" virus induced disease.

  20. Recent developments in monoclonal antibody radiolabeling techniques

    Energy Technology Data Exchange (ETDEWEB)

    Srivastava, S.C.; Mease, R.C.

    1989-01-01

    Monoclonal antibodies (MAbs) have shown the potential to serve as selective carriers of radionuclides to specific in vivo antigens. Accordingly, there has been an intense surge of research activity in an effort to develop and evaluate MAb-based radiopharmaceuticals for tumor imaging (radioimmunoscintigraphy) and therapy (radioimmunotherapy), as well as for diagnosing nonmalignant diseases. A number of problems have recently been identified, related to the MAbs themselves and to radiolabeling techniques, that comprise both the selectivity and the specificity of the in vivo distribution of radiolabeled MAbs. This paper will address some of these issues and primarily discuss recent developments in the techniques for radiolabeling monoclonal antibodies that may help resolve problems related to the poor in vivo stability of the radiolabel and may thus produce improved biodistribution. Even though many issues are identical with therapeutic radionuclides, the discussion will focus mainly on radioimmunoscintigraphic labels. 78 refs., 6 tabs.

  1. [Biophysical Characterization of Biopharmaceuticals, Including Antibody Drugs].

    Science.gov (United States)

    Uchiyama, Susumu

    2016-01-01

    Biopharmaceuticals, including antibody drugs, are now popular because of their high specificity with low adverse effects, especially in the treatment of cancer and autoimmune diseases. However, because the active pharmaceutical ingredients of biopharmaceuticals are proteins, biophysical characterization of these therapeutic proteins should be required. In this manuscript, methods of chemical and physical characterization of therapeutic proteins are described. In terms of chemical characterization, analysis of chemical modifications of the constituent amino acids is explained. Physical characterization includes higher order structural analysis and assessment of protein aggregates. Quantification methods of aggregates with different sizes, recently encouraged by the U.S. Food and Drug Administration (FDA), are introduced. As for the stability of therapeutic proteins, the importance of chemical and physical stability is explained. Finally, the contribution of colloidal and structural stability to the production of an antibody drug less prone to aggregation is introduced.

  2. Lung injury mediated by antibodies to endothelium. II. Study of the effect of repeated antigen-antibody interactions in rabbits tolerant to heterologous antibody.

    OpenAIRE

    Camussi, G.; Caldwell, P. R.; Andres, G.; Brentjens, J. R.

    1987-01-01

    The effect of repeated interactions of antibodies with cell surface antigens have been examined in in vitro, but not in in vivo systems. In this study are described the results of multiple antibody-cell surface antigen interactions in vivo. Rabbits were given repeated intravenous injections of goat antibodies to angiotensin converting enzyme (ACE), an antigen expressed on the surface of lung endothelial cells. For prevention of anaphylactic reactions, which would have been induced by multiple...

  3. Technological progresses in monoclonal antibody production systems

    OpenAIRE

    Rodrigues, E.; Costa, A R; Henriques, Mariana; Azeredo, Joana; Oliveira, Rosário

    2009-01-01

    Monoclonal antibodies (mAbs) have become vitally important to modern medicine and are currently one of the major biopharmaceutical products in development. However, the high clinical dose requirements of mAbs demand a greater biomanufacturing capacity, leading to the development of new technologies for their large-scale production, with mammalian cell culture dominating the scenario. Although some companies have tried to meet these demands by creating bioreactors of increased capacity, the op...

  4. Antibody conjugate radioimmunotherapy of superficial bladder cancer

    Directory of Open Access Journals (Sweden)

    Alan Perkins

    2002-09-01

    Full Text Available The administration of antibody conjugates for cancer therapy is now proving to be of clinical value. We are currently undertaking a programme of clinical studies using the monoclonal antibody C595 (IgG3 which reacts with the MUC1 glycoprotein antigen that is aberrantly expressed in a high proportion of bladder tumours. Radioimmunoconjugates of the C595 antibody have been produced with high radiolabelling efficiency and immunoreactivity using Tc-99m and In-111 for diagnostic imaging, and disease staging and the cytotoxic radionuclides Cu-67 and Re-188 for therapy of superficial bladder cancer. A Phase I/II therapeutic trail involving the intravesical administration of antibody directly into the bladder has now begun.A administração de anticorpos conjugados para o tratamento do câncer está agora provando ser de valor clínico. Nós estamos atualmente realizando um programa de estudos clínicos usando o anticorpo monoclonal C595 (IgG3 que reage com a glicoproteína MUC1 que está aberrantemente expressa numa alta proporção de tumores de bexiga. Tem sido produzidos radioimunoconjugados do anticorpo C595, com alta eficiência de radiomarcação e a imunoreatividade, usando-se o Tc-99m e In-111, para o diagnóstico por imagem e estagiamento de doenças. Tem sido produzidos, também, radionuclídeos citotóxicos (Cu-67 e Re-188 para o tratamento de cânceres superficiais de bexiga. A fase terapêutica I/II já se iniciou, envolvendo a administração intravesical do anticorpo diretamente na bexiga.

  5. Feature Selection Approaches In Antibody Display

    OpenAIRE

    Polaka, Inese

    2015-01-01

    Molecular diagnostics tools provide specific data that have high dimensionality due to many factors analyzed in one experiment and few records due to high costs of the experiments. This study addresses the problem of dimensionality in melanoma patient antibody display data by applying data mining feature selection techniques. The article describes feature selection ranking and subset selection approaches and analyzes the performance of various methods evaluating selected feature subsets using...

  6. Antibody induction therapy for lung transplant recipients

    DEFF Research Database (Denmark)

    Penninga, Luit; Møller, Christian H; Penninga, Ida Elisabeth Irene;

    2013-01-01

    Lung transplantation has become a valuable and well-accepted treatment option for most end-stage lung diseases. Lung transplant recipients are at risk of transplanted organ rejection, and life-long immunosuppression is necessary. Clear evidence is essential to identify an optimal, safe...... and effective immunosuppressive treatment strategy for lung transplant recipients. Consensus has not yet been achieved concerning use of immunosuppressive antibodies against T-cells for induction following lung transplantation....

  7. Antibody-Catalyzed Degradation of Cocaine

    Science.gov (United States)

    Landry, Donald W.; Zhao, Kang; Yang, Ginger X.-Q.; Glickman, Michael; Georgiadis, Taxiarchis M.

    1993-03-01

    Immunization with a phosphonate monoester transition-state analog of cocaine provided monoclonal antibodies capable of catalyzing the hydrolysis of the cocaine benzoyl ester group. An assay for the degradation of radiolabeled cocaine identified active enzymes. Benzoyl esterolysis yields ecgonine methyl ester and benzoic acid, fragments devoid of cocaine's stimulant activity. Passive immunization with such an artificial enzyme could provide a treatment for dependence by blunting reinforcement.

  8. Myopathy with anti-HMGCR antibodies

    OpenAIRE

    Alshehri, Ali; Choksi, Rati; Bucelli, Robert; Pestronk, Alan

    2015-01-01

    Objective: To analyze clinical features and myopathology changes in muscle fibers, connective tissue, and vessels in 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) antibody–associated myopathies. Methods: Retrospective review of records and myopathologic features of 49 consecutive patients with myopathies and serum HMGCR antibodies. Results: Clinical features included onset age from 12 to 83 years, female predominance (67%), proximal, symmetric weakness (84%), muscle discomfort (78%)...

  9. Sequencing antibody repertoires: The next generation

    OpenAIRE

    Fischer, Nicolas

    2011-01-01

    Genomic studies have been revolutionized by the use of next generation sequencing (NGS), which delivers huge amounts of sequence information in a short span of time. The number of applications for NGS is rapidly expanding and significantly transforming many areas of life sciences. The field of antibody research and discovery is no exception. Several recent studies have harnessed the power of NGS for analyzing natural or synthetic immunoglobulin repertoires with unprecedented resolution and ex...

  10. Antibody Discovery via Multiplexed Single Cell Characterization

    OpenAIRE

    Harriman, William D.; Collarini, Ellen J.; Sperinde, Gizette V.; Strandh, Magnus; Fatholahi, Marjan M.; Dutta, April; Lee, Yunji; Mettler, Shelley E.; Keyt, Bruce A.; Ellsworth, Stote L.; Kauvar, Lawrence M.

    2008-01-01

    The secreted immunoglobulin footprint of single hybridoma cells, containing ~10 fg of antibody purified in situ, has been probed for 9 properties concurrently by use of detection labels comprising 280 nm combinatorially colored fluorescent latex beads functionalized with proteins. Specificity of each individual hybridoma cell’s product has thereby been assessed in a primary screen. Varying the density of antigen on beads to modulate the avidity of the interaction between bead and secreted ant...

  11. Seropositivity of Dengue Antibodies during Pregnancy

    OpenAIRE

    Nor Azlin Mohamed Ismail; Wan Elly Rushima Wan Abd Rahim; Sharifah Azura Salleh; Hui-Min Neoh; Rahman Jamal; Muhammad Abdul Jamil

    2014-01-01

    Purpose. Malaysia a dengue endemic country with dengue infections in pregnancy on the rise. The present study was aimed at determining dengue seroprevalence (IgG or IgM) during pregnancy and its neonatal transmission in dengue seropositive women. Methods. Maternal with paired cord blood samples were tested for dengue antibodies (IgG and IgM) using an enzyme-linked immunosorbent assay (ELISA). Maternal age, parity, occupation, ethnic group, and gestational age were recorded. Data on neonatal A...

  12. Rare blood donors with irregular antibodies

    Directory of Open Access Journals (Sweden)

    Krga-Milanović Mirjana

    2013-01-01

    Full Text Available Introduction. Blood groups are inherited biological characteristics that do not change throughout life in healthy people. Blood groups represent antigens found on the surface of red blood cells. Kell blood group system consists of 31 antigens. Kell antigen (K is present in 0.2% of the population (the rare blood group. Cellano antigen is present in more than 99% (the high-frequency antigen. These antigens have a distinct ability to cause an immune response in the people after blood transfusion or pregnancy who, otherwise, did not have them before. Case Report. This paper presents a blood donor with a rare blood group, who was found to have an irregular antibody against red blood cells by indirect antiglobulin test. Further testing determined the specificity of antibody to be anti-Cellano. The detected antibody was found in high titers (1024 with erythrocyte phenotype Kell-Cellano+. The blood donor was found to have a rare blood group KellKell. This donor was excluded from further blood donation. It is difficult to find compatible blood for a person who has developed an antibody to the high-frequency antigen. The donor’s family members were tested and Cellano antigen was detected in her husband and child. A potential blood donor was not found among the family members. There was only one blood donor in the Register of blood donors who was compatible in the ABO and Kell blood group system. Conclusion. For the successful management of blood transfusion it is necessary to establish a unified national register of donors of rare blood groups and cooperate with the International Blood Group Reference Laboratory in Bristol with the database that registers donors of rare blood groups from around the world.

  13. Solid-phase fluoroimmunoassay for treponemal antibody.

    OpenAIRE

    Stevens, R W; Schell, R F

    1982-01-01

    An objective, solid-phase fluoroimmunoassay for treponemal antibody was developed with a lysate of virulent Treponema pallidum (Nichols strain) adsorbed on cellulose acetate disks. A probe containing both the antigen and control disks is inserted successively into a serum specimen dilution, a buffer rinse, fluoroscein isothiocyanate-conjugated goat anti-human immunoglobulin G, and a second buffer rinse. Fluorescence signal units are measured with a fluorometer. To establish test calibration c...

  14. DETECTION OF THE BOVINE VIRAL DIARRHEA ANTIBODIES

    Directory of Open Access Journals (Sweden)

    I. V. Goraichuk

    2013-06-01

    Full Text Available Bovine viral diarrhea is a widespread infection of cattle that has a wide range of clinical symptoms in domestic and wild ruminants. It is a major problem in cattle and causes significant economic losses in the cattle industry. The virus infects bovines of all ages and causes both immunosuppression and reproductive, respiratory and digestive disorders. Persistently infected cattle are the main factor in transmission of the disease between and among herds. Comparative results of antibodies presence received by two methods of enzymoimmunoassay and virus neutralization test are given in the paper. During the work, 1010 samples of blood serum of cattle from three farms in the Kharkiv region were selected and analyzed. Bovine viral diarrhea virus concerning antibodies were found by enzymoimmunoassay in 704 samples (69.7% using commercial kit and in 690 samples (68.3% using in house method. After results clarification by virus neutralization test, bovine viral diarrhea antibodies were found in 712 samples (70.5%. Immunoenzyme analysis is recommended for mass screening of cattle for viral diarrhea occurrence. The results confirm that the sensitivity immunoenzyme analysis satisfies the requirements of the diagnostic methods. Using the neutralization reaction of viruses as the «gold standard» of serological methods, it is appropriate to clarify the results of immunoenzyme analysis. Since the results contain a signi ficant number of false positive results, it is necessary to carry out comprehensive studies using both serological and molecular genetics methods.

  15. Polyclonal Antibody Therapies for Clostridium difficile Infection

    Directory of Open Access Journals (Sweden)

    Michael R. Simon

    2014-10-01

    Full Text Available Clostridium difficile infection has emerged as a growing worldwide health problem. The colitis of Clostridium difficile infection results from the synergistic action of C. difficile secreted toxins A and B upon the colon mucosa. A human monoclonal IgG anti-toxin has demonstrated the ability in combination therapy to reduce mortality in C. difficile challenged hamsters. This antibody is currently in a clinical trial for the treatment of human Clostridium difficile infection. More than one group of investigators has considered using polyclonal bovine colostral antibodies to toxins A and B as an oral passive immunization. A significant proportion of the healthy human population possesses polyclonal antibodies to the Clostridium difficile toxins. We have demonstrated that polyclonal IgA derived from the pooled plasma of healthy donors possesses specificity to toxins A and B and can neutralize these toxins in a cell-based assay. This suggests that secretory IgA prepared from such pooled plasma IgA may be able to be used as an oral treatment for Clostridium difficile infection.

  16. Monoclonal antibodies based on hybridoma technology.

    Science.gov (United States)

    Yagami, Hisanori; Kato, Hiroshi; Tsumoto, Kanta; Tomita, Masahiro

    2013-03-01

    Based on the size and scope of the present global market for medicine, monoclonal antibodies (mAbs) have a very promising future, with applications for cancers through autoimmune ailments to infectious disease. Since mAbs recognize only their target antigens and not other unrelated proteins, pinpoint medical treatment is possible. Global demand is dramatically expanding. Hybridoma technology, which allows production of mAbs directed against antigens of interest is therefore privileged. However, there are some pivotal points for further development to generate therapeutic antibodies. One is selective generation of human mAbs. Employment of transgenic mice producing human antibodies would overcome this problem. Another focus is recognition sites and conformational epitopes in antigens may be just as important as linear epitopes, especially when membrane proteins such as receptors are targeted. Recognition of intact structures is of critical importance for medical purposes. In this review, we describe patent related information for therapeutic mAbs based on hybridoma technology and also discuss new advances in hybridoma technology that facilitate selective production of stereospecific mAbs. PMID:24237029

  17. Serum Antibody Repertoire Profiling Using In Silico Antigen Screen.

    Science.gov (United States)

    Liu, Xinyue; Hu, Qiang; Liu, Song; Tallo, Luke J; Sadzewicz, Lisa; Schettine, Cassandra A; Nikiforov, Mikhail; Klyushnenkova, Elena N; Ionov, Yurij

    2013-01-01

    Serum antibodies are valuable source of information on the health state of an organism. The profiles of serum antibody reactivity can be generated by using a high throughput sequencing of peptide-coding DNA from combinatorial random peptide phage display libraries selected for binding to serum antibodies. Here we demonstrate that the targets of immune response, which are recognized by serum antibodies directed against sequential epitopes, can be identified using the serum antibody repertoire profiles generated by high throughput sequencing. We developed an algorithm to filter the results of the protein database BLAST search for selected peptides to distinguish real antigens recognized by serum antibodies from irrelevant proteins retrieved randomly. When we used this algorithm to analyze serum antibodies from mice immunized with human protein, we were able to identify the protein used for immunizations among the top candidate antigens. When we analyzed human serum sample from the metastatic melanoma patient, the recombinant protein, corresponding to the top candidate from the list generated using the algorithm, was recognized by antibodies from metastatic melanoma serum on the western blot, thus confirming that the method can identify autoantigens recognized by serum antibodies. We demonstrated also that our unbiased method of looking at the repertoire of serum antibodies reveals quantitative information on the epitope composition of the targets of immune response. A method for deciphering information contained in the serum antibody repertoire profiles may help to identify autoantibodies that can be used for diagnosing and monitoring autoimmune diseases or malignancies. PMID:23826227

  18. IBC's 23rd Antibody Engineering and 10th Antibody Therapeutics Conferences and the Annual Meeting of The Antibody Society: December 2-6, 2012, San Diego, CA.

    Science.gov (United States)

    Marquardt, John; Begent, Richard H J; Chester, Kerry; Huston, James S; Bradbury, Andrew; Scott, Jamie K; Thorpe, Philip E; Veldman, Trudi; Reichert, Janice M; Weiner, Louis M

    2012-01-01

    Now in its 23rd and 10th years, respectively, the Antibody Engineering and Antibody Therapeutics conferences are the Annual Meeting of The Antibody Society. The scientific program covers the full spectrum of challenges in antibody research and development from basic science through clinical development. In this preview of the conferences, the chairs provide their thoughts on sessions that will allow participants to track emerging trends in (1) the development of next-generation immunomodulatory antibodies; (2) the complexity of the environment in which antibodies must function; (3) antibody-targeted central nervous system (CNS) therapies that cross the blood brain barrier; (4) the extension of antibody half-life for improved efficacy and pharmacokinetics (PK)/pharmacodynamics (PD); and (5) the application of next generation DNA sequencing to accelerate antibody research. A pre-conference workshop on Sunday, December 2, 2012 will update participants on recent intellectual property (IP) law changes that affect antibody research, including biosimilar legislation, the America Invents Act and recent court cases. Keynote presentations will be given by Andreas Plückthun (University of Zürich), who will speak on engineering receptor ligands with powerful cellular responses; Gregory Friberg (Amgen Inc.), who will provide clinical updates of bispecific antibodies; James D. Marks (University of California, San Francisco), who will discuss a systems approach to generating tumor targeting antibodies; Dario Neri (Swiss Federal Institute of Technology Zürich), who will speak about delivering immune modulators at the sites of disease; William M. Pardridge (University of California, Los Angeles), who will discuss delivery across the blood-brain barrier; and Peter Senter (Seattle Genetics, Inc.), who will present his vision for the future of antibody-drug conjugates. For more information on these meetings or to register to attend, please visit www.IBCLifeSciences.com/Antibody

  19. Monoclonal Antibodies Attached to Carbon Nanotube Transistors for Paclitaxel Detection

    Science.gov (United States)

    Lee, Wonbae; Lau, Calvin; Richardson, Mark; Rajapakse, Arith; Weiss, Gregory; Collins, Philip; UCI, Molecular Biology; Biochemistry Collaboration; UCI, Departments of Physics; Astronomy Collaboration

    Paclitaxel is a naturally-occurring pharmaceutical used in numerous cancer treatments, despite its toxic side effects. Partial inhibition of this toxicity has been demonstrated using weakly interacting monoclonal antibodies (3C6 and 8A10), but accurate monitoring of antibody and paclitaxel concentrations remains challenging. Here, single-molecule studies of the kinetics of antibody-paclitaxel interactions have been performed using single-walled carbon nanotube field-effect transistors. The devices were sensitized with single antibody attachments to record the single-molecule binding dynamics of paclitaxel. This label-free technique recorded a range of dynamic interactions between the antibody and paclitaxel, and it provided sensitive paclitaxel detection for pM to nM concentrations. Measurements with two different antibodies suggest ways of extending this working range and uncovering the mechanistic differences among different antibodies.

  20. Mechanism of human antibody-mediated neutralization of Marburg virus.

    Science.gov (United States)

    Flyak, Andrew I; Ilinykh, Philipp A; Murin, Charles D; Garron, Tania; Shen, Xiaoli; Fusco, Marnie L; Hashiguchi, Takao; Bornholdt, Zachary A; Slaughter, James C; Sapparapu, Gopal; Klages, Curtis; Ksiazek, Thomas G; Ward, Andrew B; Saphire, Erica Ollmann; Bukreyev, Alexander; Crowe, James E

    2015-02-26

    The mechanisms by which neutralizing antibodies inhibit Marburg virus (MARV) are not known. We isolated a panel of neutralizing antibodies from a human MARV survivor that bind to MARV glycoprotein (GP) and compete for binding to a single major antigenic site. Remarkably, several of the antibodies also bind to Ebola virus (EBOV) GP. Single-particle EM structures of antibody-GP complexes reveal that all of the neutralizing antibodies bind to MARV GP at or near the predicted region of the receptor-binding site. The presence of the glycan cap or mucin-like domain blocks binding of neutralizing antibodies to EBOV GP, but not to MARV GP. The data suggest that MARV-neutralizing antibodies inhibit virus by binding to infectious virions at the exposed MARV receptor-binding site, revealing a mechanism of filovirus inhibition. PMID:25723164

  1. Determination of thyroid antimicrosomal antibodies by solid-phase radioimmunoassay

    Energy Technology Data Exchange (ETDEWEB)

    Lacka, K.; Kasowicz, J.; Furmaniak-Wehr, J.; Gembicki, M. (Akademia Medyczna, Poznan (Poland))

    1984-01-01

    Thyroid antimicrosomal antibodies have been detected in patient's serum by immunofluorescence and hemagglutination techniques as well as by radioimmunoassay. The aim was to introduce the solid-phase radioimmunoassay for thyroid antimicrosomal antibodies determination. The microsomal fraction was adsorbed on the walls of polystyrene tubes. In the next stage patient's serum was added. The retention of antibodies bound to microsomes on the tube walls was detected by adding /sup 125/I-protein A. The occurrence of antimicrosomal antibodies in the group of patients with Graves' disease was 88%; in case of myxedema autoantibodies were found in 70%. In patients with chronic thyroiditis antimicrosomal antibodies were present in 90% and in the group of non-toxic goiter the incidence was 25%. In the control group antimicrosomal antibodies were found in 8%. The solid-phase radioimmunoassay for thyroid antimicrosomal antibody determination proved to be a useful, sensitive and reproducible method for clinical studies.

  2. Anticardiolipin antibodies in proliferative diabetic retinopathy: An additional risk factor

    International Nuclear Information System (INIS)

    To report the prevalence of anticardiolipin antibodies in patients with proliferative diabetic retinopathy (PDR) having high-risk criteria (HRC). Diabetic patients having PDR with HRC and diabetics free of retinopathy were compared for the presence of anticardiolipin antibodies. Among the 34 patients, 6 (17.7%) of diabetics having PDR with HRC were positive for anticardiolipin antibodies. There was no significant association of aCL antibodies with sex or type of diabetes. Using Pearson's correlation test, no significant associations of aCL antibodies with duration of diabetes or age of patients were found. All patients who were positive for anticardiolipin antibodies had PDR with HRC. The difference was statistically significant. Presence of anticardiolipin antibodies may represent an additional risk factor for PDR. (author)

  3. Quantitative assessment of antibody internalization with novel monoclonal antibodies against Alexa fluorophores.

    Directory of Open Access Journals (Sweden)

    Sindy Liao-Chan

    Full Text Available Antibodies against cell surface antigens may be internalized through their specific interactions with these proteins and in some cases may induce or perturb antigen internalization. The anti-cancer efficacy of antibody-drug conjugates is thought to rely on their uptake by cancer cells expressing the surface antigen. Numerous techniques, including microscopy and flow cytometry, have been used to identify antibodies with desired cellular uptake rates. To enable quantitative measurements of internalization of labeled antibodies, an assay based on internalized and quenched fluorescence was developed. For this approach, we generated novel anti-Alexa Fluor monoclonal antibodies (mAbs that effectively and specifically quench cell surface-bound Alexa Fluor 488 or Alexa Fluor 594 fluorescence. Utilizing Alexa Fluor-labeled mAbs against the EphA2 receptor tyrosine kinase, we showed that the anti-Alexa Fluor reagents could be used to monitor internalization quantitatively over time. The anti-Alexa Fluor mAbs were also validated in a proof of concept dual-label internalization assay with simultaneous exposure of cells to two different mAbs. Importantly, the unique anti-Alexa Fluor mAbs described here may also enable other single- and dual-label experiments, including label detection and signal enhancement in macromolecules, trafficking of proteins and microorganisms, and cell migration and morphology.

  4. The INNs and outs of antibody nonproprietary names.

    Science.gov (United States)

    Jones, Tim D; Carter, Paul J; Plückthun, Andreas; Vásquez, Max; Holgate, Robert G E; Hötzel, Isidro; Popplewell, Andrew G; Parren, Paul W H I; Enzelberger, Markus; Rademaker, Hendrik J; Clark, Michael R; Lowe, David C; Dahiyat, Bassil I; Smith, Victoria; Lambert, John M; Wu, Herren; Reilly, Mary; Haurum, John S; Dübel, Stefan; Huston, James S; Schirrmann, Thomas; Janssen, Richard A J; Steegmaier, Martin; Gross, Jane A; Bradbury, Andrew R M; Burton, Dennis R; Dimitrov, Dimiter S; Chester, Kerry A; Glennie, Martin J; Davies, Julian; Walker, Adam; Martin, Steve; McCafferty, John; Baker, Matthew P

    2016-01-01

    An important step in drug development is the assignment of an International Nonproprietary Name (INN) by the World Health Organization (WHO) that provides healthcare professionals with a unique and universally available designated name to identify each pharmaceutical substance. Monoclonal antibody INNs comprise a -mab suffix preceded by a substem indicating the antibody type, e.g., chimeric (-xi-), humanized (-zu-), or human (-u-). The WHO publishes INN definitions that specify how new monoclonal antibody therapeutics are categorized and adapts the definitions to new technologies. However, rapid progress in antibody technologies has blurred the boundaries between existing antibody categories and created a burgeoning array of new antibody formats. Thus, revising the INN system for antibodies is akin to aiming for a rapidly moving target. The WHO recently revised INN definitions for antibodies now to be based on amino acid sequence identity. These new definitions, however, are critically flawed as they are ambiguous and go against decades of scientific literature. A key concern is the imposition of an arbitrary threshold for identity against human germline antibody variable region sequences. This leads to inconsistent classification of somatically mutated human antibodies, humanized antibodies as well as antibodies derived from semi-synthetic/synthetic libraries and transgenic animals. Such sequence-based classification implies clear functional distinction between categories (e.g., immunogenicity). However, there is no scientific evidence to support this. Dialog between the WHO INN Expert Group and key stakeholders is needed to develop a new INN system for antibodies and to avoid confusion and miscommunication between researchers and clinicians prescribing antibodies. PMID:26716992

  5. Anticardiolipin antibodies in pathogenesis of infertility

    Directory of Open Access Journals (Sweden)

    Lončar Dragan

    2010-01-01

    Full Text Available Background/Aim. Antiphospholipid syndrome (APS is an autoimmune disorder clinically characterized by arterial or venous thrombosis and/or specific obstetric complications and presence of antiphospholipid antibodies (aPL in the serum. It occurs in 0.3% of pregnant women, while 1% of them have two spontaneous abortions. The aim of this study was to analyze the frequency of biphospholipid antibodies in pregnant women with recurrent spontaneous abortions. Methods. We analyzed 60 pregnant women who had two or more recurrent miscarriages. The control group included 60 healthy pregnant women. We analyzed titres of anticardiolipin (aCL IgG and/or IgM with high titres (> 20 U/mL, lupus anticoagulant (LAC antibodies and anti-beta-2 glycoprotein (b2-GP1 IgG as well as parameters of coagulation status of pregnant women. Results. Analyzing Spearman's rank correlation coefficient in a group of affected patients, we noticed a slightly positive correlation of lupus anticoagulants (LAC with aCL antibodies of both classes, while the correlation with b2GP1 IgG was negative. Both classes of aCL antibodies and antib2GP1 IgG were in a discrete positive correlation with the given variables. In the control group, there was a lack of consistency in correlation of the study variables with LAC-aCl IgG, compared to the affected patients, and there was a standard negative coefficient of correlation with anti-b2GP1 IgG. The correlation ratio of anti-b2GP1 IgG was negative for all studied test parameters. Analysis of hemostatic parameters showed a statistically significant difference in the concentration of fibrinogen (p < 0.01 and thrombocyte count (p < 0.05 between the study and the control group of pregnant women. Lower mean values of fibrinogen (2.90 ± 0.45 g/L and lower thrombocyte count [(179.20 ± 6.00 × 109] were found in the study group of pregnant women with secondary infertility compared to the mean values of fibrinogen (3.60 ± 0.55 g/L and thrombocyte count

  6. Antibody induction versus placebo, no induction, or another type of antibody induction for liver transplant recipients

    DEFF Research Database (Denmark)

    Penninga, Luit; Wettergren, André; Wilson, Colin H;

    2014-01-01

    BACKGROUND: Liver transplantation is an established treatment option for end-stage liver failure. To date, no consensus has been reached on the use of immunosuppressive T-cell antibody induction for preventing rejection after liver transplantation. OBJECTIVES: To assess the benefits and harms...... of immunosuppressive T-cell specific antibody induction compared with placebo, no induction, or another type of T-cell specific antibody induction for prevention of acute rejection in liver transplant recipients. SEARCH METHODS: We searched The Cochrane Hepato-Biliary Group Controlled Trials Register, the Cochrane...... Central Register of Controlled Trials (CENTRAL), MEDLINE, EMBASE, Science Citation Index Expanded, and the World Health Organization (WHO) International Clinical Trials Registry Platform (ICTRP) until September 2013. SELECTION CRITERIA: Randomised clinical trials assessing immunosuppression with T...

  7. An efficient method for isolating antibody fragments against small peptides by antibody phage display

    DEFF Research Database (Denmark)

    Duan, Zhi; Siegumfeldt, Henrik

    2010-01-01

    not always bind efficiently to passive adsorption surfaces, and we developed a simple method to quantify the binding capacity of surfaces with the peptides. Background binding (the binding of scFvs to the background matrix) is an obstacle for successful selection, and we evaluated two methods......We generated monoclonal scFv (single chain variable fragment) antibodies from an antibody phage display library towards three small synthetic peptides derived from the sequence of s1-casein. Key difficulties for selection of scFv-phages against small peptides were addressed. Small peptides do...

  8. Platelet stimulation by antifibronectin antibodies requires the Fc region of antibody.

    OpenAIRE

    Holderbaum, D; Culp, L. A.; Bensusan, H B; Gershman, H.

    1982-01-01

    Anti-human fibronectin antibodies produced in a goat or in rabbits stimulate the release of serotonin from washed or gelatin/Sepharose-treated human platelets in a dose-dependent manner. This finding led us to propose that fibronectin on the platelet plasma membrane might serve as a collagen receptor on these cells [Bensusan, H. B., Koh, T. L., Henry, K. G., Murray, B. A. & Culp, L. A. (1978) Proc. Natl. Acad. Sci. USA 75, 5864-5868]. To determine whether direct interaction of the antibody wi...

  9. Antigen-Specific Antibody Glycosylation Is Regulated via Vaccination.

    Science.gov (United States)

    Mahan, Alison E; Jennewein, Madeleine F; Suscovich, Todd; Dionne, Kendall; Tedesco, Jacquelynne; Chung, Amy W; Streeck, Hendrik; Pau, Maria; Schuitemaker, Hanneke; Francis, Don; Fast, Patricia; Laufer, Dagna; Walker, Bruce D; Baden, Lindsey; Barouch, Dan H; Alter, Galit

    2016-03-01

    Antibody effector functions, such as antibody-dependent cellular cytotoxicity, complement deposition, and antibody-dependent phagocytosis, play a critical role in immunity against multiple pathogens, particularly in the absence of neutralizing activity. Two modifications to the IgG constant domain (Fc domain) regulate antibody functionality: changes in antibody subclass and changes in a single N-linked glycan located in the CH2 domain of the IgG Fc. Together, these modifications provide a specific set of instructions to the innate immune system to direct the elimination of antibody-bound antigens. While it is clear that subclass selection is actively regulated during the course of natural infection, it is unclear whether antibody glycosylation can be tuned, in a signal-specific or pathogen-specific manner. Here, we show that antibody glycosylation is determined in an antigen- and pathogen-specific manner during HIV infection. Moreover, while dramatic differences exist in bulk IgG glycosylation among individuals in distinct geographical locations, immunization is able to overcome these differences and elicit antigen-specific antibodies with similar antibody glycosylation patterns. Additionally, distinct vaccine regimens induced different antigen-specific IgG glycosylation profiles, suggesting that antibody glycosylation is not only programmable but can be manipulated via the delivery of distinct inflammatory signals during B cell priming. These data strongly suggest that the immune system naturally drives antibody glycosylation in an antigen-specific manner and highlights a promising means by which next-generation therapeutics and vaccines can harness the antiviral activity of the innate immune system via directed alterations in antibody glycosylation in vivo.  .

  10. Antigen-Specific Antibody Glycosylation Is Regulated via Vaccination.

    Directory of Open Access Journals (Sweden)

    Alison E Mahan

    2016-03-01

    Full Text Available Antibody effector functions, such as antibody-dependent cellular cytotoxicity, complement deposition, and antibody-dependent phagocytosis, play a critical role in immunity against multiple pathogens, particularly in the absence of neutralizing activity. Two modifications to the IgG constant domain (Fc domain regulate antibody functionality: changes in antibody subclass and changes in a single N-linked glycan located in the CH2 domain of the IgG Fc. Together, these modifications provide a specific set of instructions to the innate immune system to direct the elimination of antibody-bound antigens. While it is clear that subclass selection is actively regulated during the course of natural infection, it is unclear whether antibody glycosylation can be tuned, in a signal-specific or pathogen-specific manner. Here, we show that antibody glycosylation is determined in an antigen- and pathogen-specific manner during HIV infection. Moreover, while dramatic differences exist in bulk IgG glycosylation among individuals in distinct geographical locations, immunization is able to overcome these differences and elicit antigen-specific antibodies with similar antibody glycosylation patterns. Additionally, distinct vaccine regimens induced different antigen-specific IgG glycosylation profiles, suggesting that antibody glycosylation is not only programmable but can be manipulated via the delivery of distinct inflammatory signals during B cell priming. These data strongly suggest that the immune system naturally drives antibody glycosylation in an antigen-specific manner and highlights a promising means by which next-generation therapeutics and vaccines can harness the antiviral activity of the innate immune system via directed alterations in antibody glycosylation in vivo.  .

  11. Cerebellar Ataxia and Glutamic Acid Decarboxylase Antibodies

    Science.gov (United States)

    Ariño, Helena; Gresa-Arribas, Nuria; Blanco, Yolanda; Martínez-Hernández, Eugenia; Sabater, Lidia; Petit-Pedrol, Mar; Rouco, Idoia; Bataller, Luis; Dalmau, Josep O.; Saiz, Albert; Graus, Francesc

    2016-01-01

    IMPORTANCE Current clinical and immunologic knowledge on cerebellar ataxia (CA) with glutamic acid decarboxylase 65 antibodies (GAD65-Abs) is based on case reports and small series with short-term follow-up data. OBJECTIVE To report the symptoms, additional antibodies, prognostic factors, and long-term outcomes in a cohort of patients with CA and GAD65-Abs. DESIGN, SETTING, AND PARTICIPANTS Retrospective cohort study and laboratory investigations at a center for autoimmune neurologic disorders among 34 patients with CA and GAD65-Abs, including 25 with long-term follow-up data (median, 5.4 years; interquartile range, 3.1-10.3 years). MAIN OUTCOMES AND MEASURES Analysis of clinicoimmunologic features and predictors of response to immunotherapy. Immunochemistry on rat brain, cultured neurons, and human embryonic kidney cells expressing GAD65, GAD67, α1-subunit of the glycine receptor, and a repertoire of known cell surface autoantigens were used to identify additional antibodies. Twenty-eight patients with stiff person syndrome and GAD65-Abs served as controls. RESULTS The median age of patients was 58 years (range, 33-80 years); 28 of 34 patients (82%) were women. Nine patients (26%) reported episodes of brainstem and cerebellar dysfunction or persistent vertigo several months before developing CA. The clinical presentation was subacute during a period of weeks in 13 patients (38%). Nine patients (26%) had coexisting stiff person syndrome symptoms. Systemic organ-specific autoimmunities (type 1 diabetes mellitus and others) were present in 29 patients (85%). Twenty of 25 patients with long-term follow-up data received immunotherapy (intravenous immunoglobulin in 10 and corticosteroids and intravenous immunoglobulin or other immunosuppressors in 10), and 7 of them (35%) improved. Predictors of clinical response included subacute onset of CA (odds ratio [OR], 0.50; 95% CI, 0.25-0.99; P = .047) and prompt immunotherapy (OR, 0.98; 95% CI, 0.96-0.99; P = .01). Similar

  12. A universal strategy for stable intracellular antibodies.

    Science.gov (United States)

    Shaki-Loewenstein, Shelly; Zfania, Rahely; Hyland, Stephen; Wels, Winfried S; Benhar, Itai

    2005-08-01

    The expression of intracellular antibodies (intrabodies) in mammalian cells has provided a powerful tool to manipulate microbial and cellular signalling pathways in a highly precise manner. However, several technical hurdles have thus far restricted their more widespread use. In particular, single-chain antibodies (scFvs) have been reported to fold poorly in the reducing environment of the cytoplasm and as such there has been a reluctance to use scFv-phage libraries as a source of intrabodies unless a preselection step was applied to identify these rare scFvs that could fold properly in the absence of disulfide bonds. Recently, we reported that scFvs can be efficiently expressed within the cytoplasm of bacteria when fused at the C-terminus of the Escherichia coli maltose-binding protein (MBP). Here, we demonstrate that such MBP-scFvs are similarly stabilized when expressed in the mammalian cell cytoplasm as well as other compartments. This was demonstrated by comparing MBP-scFv fusions to the corresponding unfused scFvs that activate a defective beta-galactosidase enzyme, others that neutralize the wild-type beta-galactosidase enzyme, and an antibody that blocks the epidermal growth factor receptor. In all cases, the MBP-scFvs significantly outperformed their unfused counterparts. Our results suggest that fusion of scFvs to MBP, and possibly to other "chaperones in the context of a fusion protein", may provide a universal approach for efficient expression of intrabodies in the mammalian cell cytoplasm. This strategy should allow investigators to bypass much of the in vitro scFv characterization that is often not predictive of in vivo intrabody function and provide a more efficient use of large native and synthetic scFv-phage libraries already in existence to identify intrabodies that will be active in vivo.

  13. Intratypic heterologous vaccination of calves can induce an antibody response in presence of maternal antibodies against foot-and-mouth disease virus

    OpenAIRE

    Dekker, A.; Eble, P.L.; Stockhofe-Zurwieden, N; Chenard, G.

    2014-01-01

    Background - Maternal antibodies can interfere with foot-and-mouth disease vaccination. In this study we determined whether intratypic heterologous vaccination could help to improve herd immunity. Results - In unvaccinated calves, a half-life of maternal antibodies of 21 days was determined. At two weeks of age, calves without maternal antibodies showed a good antibody response against both vaccines used in the trial, while in calves with maternal antibodies no antibody response to homologous...

  14. Presence of Autoimmune Antibody in Chikungunya Infection

    Directory of Open Access Journals (Sweden)

    Wirach Maek-a-nantawat

    2009-01-01

    Full Text Available Chikungunya infection has recently re-emerged as an important arthropod-borne disease in Thailand. Recently, Southern Thailand was identified as a potentially endemic area for the chikungunya virus. Here, we report a case of severe musculoskeletal complication, presenting with muscle weakness and swelling of the limbs. During the investigation to exclude autoimmune muscular inflammation, high titers of antinuclear antibody were detected. This is the report of autoimmunity detection associated with an arbovirus infection. The symptoms can mimic autoimmune polymyositis disease, and the condition requires close monitoring before deciding to embark upon prolonged specific treatment with immunomodulators.

  15. Antipolymer antibody in Italian fibromyalgic patients

    OpenAIRE

    Bazzichi, Laura; Giacomelli, Camillo; De Feo, Francesca; Giuliano, Tiziana; Rossi, Alessandra; Doveri, Marica; Tani, Chiara; Wilson, Russell B.; Bombardieri, Stefano

    2007-01-01

    The objectives of the present study were to evaluate the presence of antipolymer antibody (APA) seropositivity in 285 Italian patients affected by primary fibromyalgia (FM) and to verify whether APA levels correlate with disease severity and with cytokine levels. APA levels were determined on serum samples by an indirect ELISA kit that detects IgG APA. Cytokines (IL-1, IL-6, IL-8, IL-10 and TNFα) were measured by ELISA in plasma. The impact of FM on the quality of life was estimated using the...

  16. Preparation, characterization and radiolabelling of antigranulocytemonoclonal antibody

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    The human granulocytes were isolated. Using hybridoma techniques, a hybridoma cell line(HSN) producing monoclonal antibody (McAb) against human granulocyte was obtained.The antibodybelonged to IgG1 subclass. It was confirmed byimmunohistochemical tests that HSN reacted selectivelynot only with human granulocytes, but also with their bone marrow precursors. Whereas humanlymphocytesand red blood cells retained negative in the tests. No cross-reaction was observedwith theperipheral blood cells in other animals. Its affinity constant was5.7×108L/mol, and the number of epitopesper granulocyte was 4.7×105. Monoclonal antibodydisplayed no loss of immunoreactivity after labelledwith 99mTc.

  17. Development of Anti-Isoproturon Polyclonal Antibody

    Institute of Scientific and Technical Information of China (English)

    LI Fang-shi; SUN Feng; LIU Xian-jin; CUI Heng-hua

    2007-01-01

    A competitive enzyme-linked immunosorbent assay (ELISA) suitable for the determination of the urea herbicide isoproturon,3-(4-isopropylphenyl)-1,1-dimethylurea, in food and environmental samples was developed. Two haptens named 1-(3-carboxypropyl)-3-(4-isopropylphenyl)-1-methylurca (hapten 4C) and 1-(5-carboxypentyl)-3-(4-isopropylphenyl)-1-methylurea (hapten 6C) were synthesized. The haptens were coupled to bovine serum albumin (BSA) and ovalbumin(OVA), respectively, using the N-hydroxysuccinimide reaction. The hapten 6C-BSA conjugate was used as the immunogen,with which a high-titer anti-isoproturon polyclonal antibody (pAb) was successfully obtained by immunization of New Zealand white rabbits. The hapten 4C-OVA conjugate was used as coating antigen and a method of the indirect competitive ELISA for isoproturon was established. The haptens were confirmed with TLC, IR, and 1H NMR. The conjugation molar ratios of hapten 4C to OVA and hapten 6C to BSA were 36:1 and 46:1, respectively, as calculated by a UV spectrophotometry.The highest titer of the anti-isoproturon sera determined by a non-competitive indirect ELISA procedure was 1.6×105. The optimal concentrations of the coating antigen and the dilution of the anti-isoproturon sera used in the ELISA were 0.1 mg L-1 and 1.0 × 105, respectively. The concentration of isoproturon that inhibits 50% of antibody-antigen binding (IC50) was 0.07 mg mL-1.The cross-reactivities of six urea herbicides including chlorbromuron, fluometuron, monolinuron were lower than 0.1%. Isoproturon is a small molecule without immune activity and active functional group for attaching to carrier protein. To produce an antibody against isoproturon with high titer and high specificity is the most important step in the development of an immunochemical method for the determination of isoproturon in food and environmental samples. The two haptens synthesized in this study have carboxyl groups and accommodate different lengths of spacer arms, and

  18. Production of monoclonal antibodies for radioimmunoassays

    International Nuclear Information System (INIS)

    Specific antibodies (Abs) have proven most useful and versatile tools for the identification, quantification, and localization of minute amounts of small and large molecules in biologic materials, e.g., body fluids, specific cells, and other body components. So far the most widely used technique for the production of specific Abs consists in immunization of animals like rabbits, goats, or horses, monitoring of Ab formation in the serum, and selection of animals which produce serum containing Abs sufficiently specific for the use envisaged. Although this approach has yielded many valuable results, it has some deficiencies

  19. Automatic Identification of Antibodies in the Protein Data Bank

    Institute of Scientific and Technical Information of China (English)

    LI Xun; WANG Renxiao

    2009-01-01

    An automatic method has been developed for identifying antibody entries in the protein data bank (PDB). Our method, called KIAb (Keyword-based Identification of Antibodies), parses PDB-format files to search for particular keywords relevant to antibodies, and makes judgment accordingly. Our method identified 780 entries as antibodies on the entire PDB. Among them, 767 entries were confirmed by manual inspection, indicating a high success rate of 98.3%. Our method recovered basically all of the entries compiled in the Summary of Antibody Crystal Structures (SACS) database. It also identified a number of entries missed by SACS. Our method thus provides a more com-plete mining of antibody entries in PDB with a very low false positive rate.

  20. The investigation of relationship between preeclampsia and antiphospholipid antibody syndrome

    Directory of Open Access Journals (Sweden)

    Mehmet Tayyar

    2013-03-01

    Full Text Available Aim. The aim of this study was evaluate the relationship between preeclampsia and antiphospholipid antibodies. Methods. A total of 116 pregnant women between 20th and 40th weeks of gestation admitted to our department were investigated. 63 of them were allocated our preeclampsia group and 53 of them were allocated our control group. Lupus anticoagulant, anti-cardiolipin antibodies (IG G ve M and antiphosphatidylserine antibodies (IG G ve M were measured. Results. There was no statistical significance between preeclampsia and control group for antiphospholipid antibodies but these were two times higher in preeclamptic group compared to control group. (22.2% in preeclampsia, 11.3% in control group p=0.193. Conclusions. In an unselected population we were not able to demonstrate an association between preeclampsia and antiphospholipid antibody syndrome but antiphospholipid antibody ratio elevated in women with preeclampsia. These findings show that, there is a need for large scale studies.

  1. Production of monoclonal antibody with Celline-350 bioreactor

    International Nuclear Information System (INIS)

    Monoclonal antibodies are protein that are highly specific and sensitive in their reaction with specific sites on target molecules that they have become reagents of central importance in the diagnostic and treatment of human diseases. This paper reports the use of CELLine-350 bioreactor to produce continuous supply of serum-free breast cancer monoclonal antibody. Initial volume of 5ml (1.5 x 106 viable cells/ml) is inoculated into the bioreactor and harvesting is done every 5 days to obtain high yield monoclonal antibody. The serum-free supernatant is precipitated with 50% saturated ammonia sulfate and the antibody is purified by protein-G affinity chromatography. The concentration of monoclonal antibody successfully produced by the bioreactor is 0.91mg/ml respectively and it is measured by the Lowry method. This result shows that bioreactor Celline-350 is easy to handle and cost effective for the continuous production of serum free monoclonal antibody. (Author)

  2. Generation and characterization of novel stromal specific antibodies

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    Rheumatoid synovial fibroblasts were used as an immunogen to produce monoclonal antibodies selected for their reactivity with stromal cell antigens. Mice were immunised with low passage whole cell preparations and the subsequent hybridomas were screened by immunohistochemistry on rheumatoid synovium and tonsil sections. The aim was to identify those antibodies that recognised antigens that were restricted to stromal cells and were not expressed on CD45 positive leucocytes. A significant number of antibodies detected antigen that identified endothelial cells. These antibodies were further characterised to determine whether the vessels identified by these antibodies were vascular or lymphatic.From five fusions clones were identified with predominant reactivity with: 1) fibroblasts and endothelial cells; or 2)broad stromal elements (fibroblast, endothelium, epithelium, follicular dendritic cells). A fibroblast-specific antibody that did not also identify vessels was not generated. Examples of each reactivity pattern are discussed.

  3. Rational Design of CXCR4 Specific Antibodies with Elongated CDRs

    Science.gov (United States)

    2015-01-01

    The bovine antibody (BLV1H12) which has an ultralong heavy chain complementarity determining region 3 (CDRH3) provides a novel scaffold for antibody engineering. By substituting the extended CDRH3 of BLV1H12 with modified CXCR4 binding peptides that adopt a β-hairpin conformation, we generated antibodies specifically targeting the ligand binding pocket of CXCR4 receptor. These engineered antibodies selectively bind to CXCR4 expressing cells with binding affinities in the low nanomolar range. In addition, they inhibit SDF-1-dependent signal transduction and cell migration in a transwell assay. Finally, we also demonstrate that a similar strategy can be applied to other CDRs and show that a CDRH2-peptide fusion binds CXCR4 with a Kd of 0.9 nM. This work illustrates the versatility of scaffold-based antibody engineering and could greatly expand the antibody functional repertoire in the future. PMID:25041362

  4. Design and manufacture of monoclonal antibodies for radioimmunotherapy

    International Nuclear Information System (INIS)

    Appropriate design and manufacture of monoclonal antibodies is fundamental to their use for radioimmunotherapy. Besides the right selection of antibody specificity and affinity, recombinant antibodies can be designed to simplify manufacture and minimise unwanted side effects. Although many innovative new technologies have been developed in recent years, antibodies are still most commonly produced from mammalian cells and purified by column chromatography. Purification methods have to be designed and validated to remove potential contaminants, especially retroviruses which in principle might be present in mammalian cell lines. Adherence to relevant Good Manufacturing Practice is mandatory in the production of any medicinal product and there are numerous guidelines regarding the manufacture of antibodies. This article outlines some methods used for fermentation, purification and quality control of antibodies intended for radiolabelling

  5. Engineering broadly neutralizing antibodies for HIV prevention and therapy.

    Science.gov (United States)

    Hua, Casey K; Ackerman, Margaret E

    2016-08-01

    A combination of advances spanning from isolation to delivery of potent HIV-specific antibodies has begun to revolutionize understandings of antibody-mediated antiviral activity. As a result, the set of broadly neutralizing and highly protective antibodies has grown in number, diversity, potency, and breadth of viral recognition and neutralization. These antibodies are now being further enhanced by rational engineering of their anti-HIV activities and coupled to cutting edge gene delivery and strategies to optimize their pharmacokinetics and biodistribution. As a result, the prospects for clinical use of HIV-specific antibodies to treat, clear, and prevent HIV infection are gaining momentum. Here we discuss the diverse methods whereby antibodies are being optimized for neutralization potency and breadth, biodistribution, pharmacokinetics, and effector function with the aim of revolutionizing HIV treatment and prevention options. PMID:26827912

  6. Antibody engineering: facing new challenges in cancer therapy

    Institute of Scientific and Technical Information of China (English)

    Laura SANZ; (A)ngel M CUESTA; Marta COMPTE; Luis (A)LVAREZ-VALLINA

    2005-01-01

    Antibody-based therapeutics are beginning to realize the promise enclosed in their early denomination as "magic bullets". Initial disappointment has turned into clinical and commercial success, and engineered antibodies currently represent over 30% of biopharmaceuticals in clinical trials. Recent structural and functional data have allowed the design of a new generation of therapeutic antibodies, with strategies ranging from complement-mediated and antibody-dependant cellular cytotoxicity enhancement to improved cytotoxic payloads using toxins, drugs,radionucleids and viral delivery. This review considers the structure of different types of recombinant antibodies, their mechanism of action and how their efficacy has been increased using a broad array of approaches. We will also focus on the additional benefits offered by the use of gene therapy methods for the in vivo production of therapeutic antibodies.

  7. Fc-galactosylation modulates antibody-dependent cellular cytotoxicity of therapeutic antibodies.

    Science.gov (United States)

    Thomann, Marco; Reckermann, Katharina; Reusch, Dietmar; Prasser, Jessica; Tejada, Max L

    2016-05-01

    The therapeutic activity of monoclonal antibodies can involve immune cell mediated effector functions including antibody-dependent cellular cytotoxicity (ADCC), an activity that is modulated by the structure of Fc-glycans, and in particular the lack of core fucose. The heterogeneity of these glycostructures and the inherent variability of traditional PBMC-based in vitro ADCC assays, have made it challenging to quantitatively assess the impact of other glycostructures on ADCC activity. We applied a quantitative NK cell based assay to generate a database consisting of Fc-glycostructure and ADCC data from 54 manufacturing batches of a CHO-derived monoclonal antibody. Explorative analysis of the data indicated that, apart from afucosylation, galactosylation levels could influence ADCC activity. We confirmed this hypothesis by demonstrating enhanced ADCC upon enzymatic hypergalactosylation of four different monoclonal antibodies derived using standard CHO manufacturing processes. Furthermore we quantitatively compare the effects of galactosylation and afucosylation in the context of glycan heterogeneity and demonstrate that while galactose can influence ADCC activity, afucosylation remains the primary driver of this activity. PMID:27058641

  8. An Insertion Mutation That Distorts Antibody Binding Site Architecture Enhances Function of a Human Antibody

    Energy Technology Data Exchange (ETDEWEB)

    Krause, Jens C.; Ekiert, Damian C.; Tumpey, Terrence M.; Smith, Patricia B.; Wilson, Ian A.; Crowe, Jr., James E. (Vanderbilt); (Scripps); (CDC)

    2011-09-02

    The structural and functional significance of somatic insertions and deletions in antibody chains is unclear. Here, we demonstrate that a naturally occurring three-amino-acid insertion within the influenza virus-specific human monoclonal antibody 2D1 heavy-chain variable region reconfigures the antibody-combining site and contributes to its high potency against the 1918 and 2009 pandemic H1N1 influenza viruses. The insertion arose through a series of events, including a somatic point mutation in a predicted hot-spot motif, introduction of a new hot-spot motif, a molecular duplication due to polymerase slippage, a deletion due to misalignment, and additional somatic point mutations. Atomic resolution structures of the wild-type antibody and a variant in which the insertion was removed revealed that the three-amino-acid insertion near the base of heavy-chain complementarity-determining region (CDR) H2 resulted in a bulge in that loop. This enlarged CDR H2 loop impinges on adjacent regions, causing distortion of the CDR H1 architecture and its displacement away from the antigen-combining site. Removal of the insertion restores the canonical structure of CDR H1 and CDR H2, but binding, neutralization activity, and in vivo activity were reduced markedly because of steric conflict of CDR H1 with the hemagglutinin antigen.

  9. Antibody structural modeling with prediction of immunoglobulin structure (PIGS)

    DEFF Research Database (Denmark)

    Marcatili, Paolo; Olimpieri, Pier Paolo; Chailyan, Anna;

    2014-01-01

    Antibodies (or immunoglobulins) are crucial for defending organisms from pathogens, but they are also key players in many medical, diagnostic and biotechnological applications. The ability to predict their structure and the specific residues involved in antigen recognition has several useful appl...... on average) to build a structural model of an antibody. It is based on the concept of canonical structures of antibody loops and on our understanding of the way light and heavy chains pack together....

  10. Antibody-mediated Xenograft Injury: Mechanisms and Protective Strategies

    OpenAIRE

    Pierson, Richard N.

    2009-01-01

    The use of porcine organs for clinical transplantation is a promising potential solution to the shortage of human organs. Preformed anti-pig antibody is the primary cause of hyperacute rejection, while elicited antibody can contribute to subsequent “delayed” xenograft rejection. This article will review recent progress to overcome antibody mediated xenograft rejection, through modification of the host immunity and use of genetically engineered pig organs.

  11. Distinction between antinuclear antibody and P-ANCA.

    OpenAIRE

    Lee, S. S.; Lawton, J W; Chak, W

    1991-01-01

    To differentiate between perinuclear immunofluorescence staining of antinuclear antibody (ANA) and the perinuclear form of anti-neutrophil cytoplasmic antibody (P-ANCA), the pattern after formaldehyde vapour fixation of normal human neutrophils was compared with that of standard ethanol fixation. Fifteen out of 17 myeloperoxidase antibody positive sera showed cytoplasmic staining on formaldehyde vapour fixed cells; 30 of the 32 ANA positive samples became negative or gave weak nuclear stainin...

  12. Critical evaluation of monoclonal antibody staining in breast carcinoma.

    OpenAIRE

    Parham, D M; Coghill, G; Robertson, A.J.

    1989-01-01

    The immunoperoxidase staining of 84 primary invasive breast carcinomas with four monoclonal antibodies (BRST-1, HMFG1, EMA, B72.3) was evaluated by semiquantitative light microscopical examination and quantitative image analysis. Major differences in the staining of the tumours for each of the monoclonal antibodies was observed. Correlation between monoclonal antibody staining and patient age, survival, histological grade, tumour diameter and cellularity was also carried out. This showed a si...

  13. Antibody avidity in swine lymphocyte antigen-defined miniature pigs.

    OpenAIRE

    Appleyard, G D; Mallard, B A; Kennedy, B. W.; Wilkie, B. N.

    1992-01-01

    Antibody avidity to hen egg white lysozyme (HEWL) was measured by thiocyanate ion elution enzyme-linked immunosorbent assay (ELISA) in swine lymphocyte antigen (SLA) defined miniature pigs. Serum antibody avidity was evaluated on day 14 and 30 after primary (day 0) and secondary (day 14) immunizations in eight to ten week old miniature pigs previously typed for swine lymphocyte antigen genotype. The effect of SLA genotype, litter, and gender on anti-HEWL antibody avidity was determined by lea...

  14. Development of antibodies to human embryonic stem cell antigens

    OpenAIRE

    Stanley Marisa; Rao Mahendra S; Olson Judith M; Cai Jingli; Taylor Eva; Ni Hsiao-Tzu

    2005-01-01

    Abstract Background Using antibodies to specific protein antigens is the method of choice to assign and identify cell lineage through simultaneous analysis of surface molecules and intracellular markers. Embryonic stem cell research can be benefited from using antibodies specific to transcriptional factors/markers that contribute to the "stemness" phenotype or critical for cell lineage. Results In this report, we have developed and validated antibodies (either monoclonal or polyclonal) specif...

  15. Anti-collagen antibodies in sera from rheumatoid arthritis patients.

    OpenAIRE

    Beard, H K; Ryvar, R; Skingle, J; Greenbury, C. L.

    1980-01-01

    Anti-cartilage antibodies, demonstrable by immunofluorescence, were found in 3.3% of rheumatoid arthritis patients. In most of these patients antibodies to type II collagen were detected. In specificity studies on these anti-collagen antibodies, they appeared to be type specific, showing no reaction with collagen types I and III. Denatured type II collagen reacted much less well than native type II, but isolated peptides from different regions of the collagen molecule were differentiated by i...

  16. Monoclonal Antibody Expression and Novel Purification in Nicotiana benthamiana

    OpenAIRE

    Fulton, Andrew Dale

    2011-01-01

    Over the past few decades researchers and industrial professionals alike have realized the vast potential of monoclonal antibodies to treat diseases ranging from arthritis, immune and infectious diseases to cancer. There are a number of antibodies on the market that constitute a large portion of the biopharmaceutical niche in the drug industry. Blockbuster drugs (selling greater than $1 billion/year), include antibodies such as Avastin (bevacizumab), Herceptin (trastuzumab), Rituxan (rituxi...

  17. Mechanisms involved in antibody- and complement-mediated allograft rejection

    OpenAIRE

    Wasowska, Barbara A.

    2010-01-01

    Antibody-mediated rejection has become critical clinically because this form of rejection is usually unresponsive to conventional anti-rejection therapy, and therefore, it has been recognized as a major cause of allograft loss. Our group developed experimental animal models of vascularized organ transplantation to study pathogenesis of antibody- and complement-mediated endothelial cell injury leading to graft rejection. In this review, we discuss mechanisms of antibody-mediated graft rejectio...

  18. Monoclonal antibodies against plant proteins recognise animal intermediate filaments.

    Science.gov (United States)

    Parke, J M; Miller, C C; Cowell, I; Dodson, A; Dowding, A; Downes, M; Duckett, J G; Anderton, B J

    1987-01-01

    Four monoclonal antibodies were raised against polypeptides present in a high-salt detergent-insoluble fraction from cells of Chlamydomonas reinhardtii. Indirect immunofluorescence microscopy of fibroblasts and epithelial cells grown in culture using these plant antibodies revealed staining arrays identical to those obtained with well characterised antibodies to animal intermediate filaments. Immunofluorescence microscopy of Chlamydomonas with these monoclonal antibodies and a monoclonal antibody that recognises all animal intermediate filaments (anti-IFA) gave a diffuse, patchy cytoplasmic staining pattern. Both the plant antibodies and anti-IFA stained interphase onion root tip cells in a diffuse perinuclear pattern. In metaphase through to telophase, the labelling patterns colocalised with those of microtubules. Labelling of the phragmoplast was also detected but not staining of the preprophase band. On Western blots of various animal cell lines and tissues, all the antibodies labelled known intermediate filament proteins. On Western blots of whole Chlamydomonas proteins, all the antibodies labelled a broad band in the 57,000 Mr range, and three antibodies labelled bands around 66,000 and 140,000 Mr but with variable intensities. On Western blots of whole onion root tip proteins, all the antibodies labelled 50,000 Mr (two to three bands) polypeptides and a diffuse band around 60,000 Mr and three of the antibodies also labelled several polypeptides in the 90,000-200,000 Mr range. The consistent labelling of these different bands by several different monoclonal antibodies recognising animal intermediate filaments makes these polypeptides putative plant intermediate filament proteins. PMID:2446785

  19. Goat serums for fluorescent antibody conjugates to chlamydial antigens.

    OpenAIRE

    Tessler, J.

    1984-01-01

    Serums from goats hyperimmunized with Chlamydia psittaci consistently produce antichlamydial fluorescent antibody conjugate of high titer. The titer of the fluorescent antibody conjugate prepared from a given serum correlated well with the titer obtained by agar gel precipitin, but not with the complement fixation. The agar gel precipitin test can be used to predict whether a given serum is satisfactory for use in production of a conjugate for direct fluorescent antibody tests. Serums with an...

  20. Purification of Murine Monoclonal IgM Antibody

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    This paper presents the purification of a monoclonal IgM antibody against human tumor associated antigen Lewis-Y by ion exchange chromatography and gel filtration.Enzyme-linked immunosorbent assay (ELISA) and SDS-polyacrylamide gel electrophoresis (PAGE) were used to identify purified IgM antibody.In flow cytometry analysis, the purified IgM antibody recognizes human breast tumor cell line MCF-7 which expresses Lewis-Y antigen.This work presents a new way for the purification of murine monoclonal IgM antibody.

  1. Antibody binding to p-Si using LANL SAM chemistry

    Energy Technology Data Exchange (ETDEWEB)

    Anderson, Aaron S [Los Alamos National Laboratory

    2010-12-06

    This NMSBA-sponsored project involves the attachment of antibodies to polymeric silicon (p-Si) surfaces, with the ultimate goal of attaching antibodies to nanowires for Vista Therapeutics, Inc. (Santa Fe, NM). This presentation describes the functionalization of p-Si surfaces. the activation of terminal carboxylates on these surfaces, the conjugation of antibodies, and the analyses undertaken at each step. The results of this work show that antibody conjugation is possible on p-Si coatings using the well-known EDC/NHS activation chemistry.

  2. Antibody-Based Strategies to Prevent and Treat Influenza

    Directory of Open Access Journals (Sweden)

    Ram eSasisekharan

    2015-07-01

    Full Text Available Passive immunization using antibodies has been suggested to offer several benefits in comparison to other antiviral treatment options. The potential for seasonal protection arising from a single injection of antibodies is appealing and has been pursued for a number of infectious agents. However, until recently, antibody-based strategies to combat infectious agents has been hampered due to the fact that typical antibodies have been found to be strain-specific, with the virus evolving resistance in many cases. The discovery of broadly neutralizing antibodies (bNAbs in, for example, influenza, dengue virus, and HIV, which bind to multiple, structurally-diverse strains has provided renewed interest in this area. This review will focus on new technologies that enable the discovery of bNAbs, the challenges and opportunities of immunotherapies as an important addition to existing antiviral therapy, and the role of antibody discovery in informing rational vaccine discovery – with agents targeting influenza specifically addressed. Multiple agents have entered the clinic and raise the possibility that a single antibody or small combination of antibodies can effectively neutralize a wide variety of strains. However, challenges remain - including combating escape variants, pharmacodynamics of antibody distribution, and development of efficacy biomarkers beyond virologic endpoints.

  3. Biophysical characterization of a model antibody drug conjugate.

    Science.gov (United States)

    Arakawa, Tsutomu; Kurosawa, Yasunori; Storms, Michael; Maruyama, Toshiaki; Okumura, C J; Maluf, Nasib Karl

    2016-01-01

    Antibody drug conjugates (ADC) are important next-generation biopharmaceuticals and thus require stringent structure characterization as is the case for monoclonal antibodies. We have tested several biophysical techniques, i.e., circular dichroism, analytical ultracentrifugation, differential scanning calorimetry and fluorescence spectroscopy, to characterize a fluorescein-labeled monoclonal antibody as a model ADC. These techniques indicated possible small structure and stability changes by the conjugation, while largely retaining the tertiary structure of the antibody, consistent with unaltered biological activities. Thus, the above biophysical techniques are effective at detecting changes in the structural properties of ADC. PMID:27534450

  4. Clinical analyses of perinuclear antineutrophil cytoplasmic antibody associated hypertrophic pachymeningitis

    Institute of Scientific and Technical Information of China (English)

    赵鹏

    2013-01-01

    Objective To explore the clinical characteristics,diagnosis,treatment and prognosis of perinuclear antineutrophil cytoplasmic antibody(p-ANCA)associated hypertrophic pachymeningitis.Methods A retrospective study

  5. Antibody Conjugates: From Heterogeneous Populations to Defined Reagents

    Directory of Open Access Journals (Sweden)

    Patrick Dennler

    2015-08-01

    Full Text Available Monoclonal antibodies (mAbs and their derivatives are currently the fastest growing class of therapeutics. Even if naked antibodies have proven their value as successful biopharmaceuticals, they suffer from some limitations. To overcome suboptimal therapeutic efficacy, immunoglobulins are conjugated with toxic payloads to form antibody drug conjugates (ADCs and with chelating systems bearing therapeutic radioisotopes to form radioimmunoconjugates (RICs. Besides their therapeutic applications, antibody conjugates are also extensively used for many in vitro assays. A broad variety of methods to functionalize antibodies with various payloads are currently available. The decision as to which conjugation method to use strongly depends on the final purpose of the antibody conjugate. Classical conjugation via amino acid residues is still the most common method to produce antibody conjugates and is suitable for most in vitro applications. In recent years, however, it has become evident that antibody conjugates, which are generated via site-specific conjugation techniques, possess distinct advantages with regard to in vivo properties. Here, we give a comprehensive overview on existing and emerging strategies for the production of covalent and non-covalent antibody conjugates.

  6. Macrophages are critical effectors of antibody therapies for cancer.

    Science.gov (United States)

    Weiskopf, Kipp; Weissman, Irving L

    2015-01-01

    Macrophages are innate immune cells that derive from circulating monocytes, reside in all tissues, and participate in many states of pathology. Macrophages play a dichotomous role in cancer, where they promote tumor growth but also serve as critical immune effectors of therapeutic antibodies. Macrophages express all classes of Fcγ receptors, and they have immense potential to destroy tumors via the process of antibody-dependent phagocytosis. A number of studies have demonstrated that macrophage phagocytosis is a major mechanism of action of many antibodies approved to treat cancer. Consequently, a number of approaches to augment macrophage responses to therapeutic antibodies are under investigation, including the exploration of new targets and development of antibodies with enhanced functions. For example, the interaction of CD47 with signal-regulatory protein α (SIRPα) serves as a myeloid-specific immune checkpoint that limits the response of macrophages to antibody therapies, and CD47-blocking agents overcome this barrier to augment phagocytosis. The response of macrophages to antibody therapies can also be enhanced with engineered Fc variants, bispecific antibodies, or antibody-drug conjugates. Macrophages have demonstrated success as effectors of cancer immunotherapy, and further investigation will unlock their full potential for the benefit of patients. PMID:25667985

  7. Surface immobilization of antibody on silk fibroin through conformational transition.

    Science.gov (United States)

    Lu, Qiang; Wang, Xiaoqin; Zhu, Hesun; Kaplan, David L

    2011-07-01

    In recent studies silk fibroin has been explored as a new material platform for biosensors. Based on these developments, a procedure for the immobilization of antibodies on silk fibroin substrates was developed as a route to functionalizing these biosensor systems. By controlling the conformational transition of the silk fibroin, a primary antibody was immobilized and enriched at the surface of silk fibroin substrates under mild reaction conditions to maintain antibody function. Compared to chemical crosslinking, the immobilization efficiency in the present approach was increased significantly. This method, achieving high loading of antibody while retaining function, improves the feasibility of silk fibroin as a platform material for biosensor applications.

  8. The Biochemical Properties of Antibodies and Their Fragments.

    Science.gov (United States)

    Hnasko, Robert M

    2015-01-01

    Immunoglobulins (Ig) or antibodies are powerful molecular recognition tools that can be used to identify minute quantities of a given target analyte. Their antigen-binding properties define both the sensitivity and selectivity of an immunoassay. Understanding the biochemical properties of this class of protein will provide users with the knowledge necessary to select the appropriate antibody composition to maximize immunoassay results. Here we define the general biochemical properties of antibodies and their similarities and differences, explain how these properties influence their functional relationship to an antigen target, and describe a method for the enzymatic fragmentation of antibodies into smaller functional parts.

  9. The antibody response in Lyme disease.

    Science.gov (United States)

    Craft, J. E.; Grodzicki, R. L.; Shrestha, M.; Fischer, D. K.; García-Blanco, M.; Steere, A. C.

    1984-01-01

    We determined the antibody response against the Ixodes dammini spirochete in Lyme disease patients by indirect immunofluorescence and an enzyme-linked immunosorbent assay (ELISA). The specific IgM response became maximal three to six weeks after disease onset, and then declined, although titers sometimes remained elevated during later disease. Specific IgM levels correlated directly with total serum IgM. The specific IgG response, often delayed initially, was nearly always present during neuritis and arthritis, and frequently remained elevated after months of remission. Although results obtained by indirect immunofluorescence and the ELISA were similar, the ELISA was more sensitive and specific. Cross-reactive antibodies from patients with other spirochetal infections were blocked by absorption of sera with Borrelia hermsii, but titers of Lyme disease sera were also decreased. To further characterize the specificity of the humoral immune response against the I. dammini spirochete, 35S-methionine-labeled spirochetal antigens were identified by immunoprecipitation with sera from Lyme arthritis patients. These polypeptides had molecular weights of 62, 60, 47, 37, 22, 18, and 15 kDa, and were not recognized by control sera. We conclude that the ELISA, without absorption, is the best method to assay the humoral immune response in Lyme disease, and we have identified methionine-containing spirochetal polypeptides that may be important in Lyme arthritis. PMID:6393607

  10. Drug Development of Therapeutic Monoclonal Antibodies.

    Science.gov (United States)

    Mould, Diane R; Meibohm, Bernd

    2016-08-01

    Monoclonal antibodies (MAbs) have become a substantial part of many pharmaceutical company portfolios. However, the development process of MAbs for clinical use is quite different than for small-molecule drugs. MAb development programs require careful interdisciplinary evaluations to ensure the pharmacology of both the MAb and the target antigen are well-understood. Selection of appropriate preclinical species must be carefully considered and the potential development of anti-drug antibodies (ADA) during these early studies can limit the value and complicate the performance and possible duration of preclinical studies. In human studies, many of the typical pharmacology studies such as renal or hepatic impairment evaluations may not be needed but the pharmacokinetics and pharmacodynamics of these agents is complex, often necessitating more comprehensive evaluation of clinical data and more complex bioanalytical assays than might be used for small molecules. This paper outlines concerns and strategies for development of MAbs from the early in vitro assessments needed through preclinical and clinical development. This review focuses on how to develop, submit, and comply with regulatory requirements for MAb therapeutics. PMID:27342605

  11. Array tomography: immunostaining and antibody elution.

    Science.gov (United States)

    Micheva, Kristina D; O'Rourke, Nancy; Busse, Brad; Smith, Stephen J

    2010-11-01

    Array tomography is a volumetric microscopy method based on physical serial sectioning. Ultrathin sections of a plastic-embedded tissue are cut using an ultramicrotome, bonded in an ordered array to a glass coverslip, stained as desired, and imaged. The resulting two-dimensional image tiles can then be reconstructed computationally into three-dimensional volume images for visualization and quantitative analysis. The minimal thickness of individual sections permits high-quality rapid staining and imaging, whereas the array format allows reliable and convenient section handling, staining, and automated imaging. Also, the physical stability of the arrays permits images to be acquired and registered from repeated cycles of staining, imaging, and stain elution, as well as from imaging using multiple modalities (e.g., fluorescence and electron microscopy). Array tomography makes it possible to visualize and quantify previously inaccessible features of tissue structure and molecular architecture. However, careful preparation of the tissue is essential for successful array tomography; these steps can be time-consuming and require some practice to perfect. In this protocol, tissue arrays are prepared for imaging by tagging with primary antibodies against specific cellular targets, followed by labeling with fluorescent secondary antibodies. Alternatively, fluorescent proteins that have been introduced into the tissue before dissection can be used. PMID:21041398

  12. Anti IH: An antibody worth mention.

    Science.gov (United States)

    Mohanan, Nithya; Henry, Nittin; Rafi, Aboobacker Mohamed; Innah, Susheela J

    2016-01-01

    A 72-year-old female with co-morbidities posted for surgical correction of fracture neck of femur without any history of transfusions was noted to have a hemoglobin level of 7 g/dl and packed red blood cells transfusion was ordered. Pretransfusion tests demonstrated A1B group with D positive on forward grouping. Reverse grouping showed a varying grade of agglutination with A, B, and O cells. Agglutination being stronger at 4°C. Antibody screening showed pan-agglutination, direct Coomb's test and auto control were negative. The serum reacted with adult O cells (OIadult) but not with adult Bombay cells (Oh Iadult) or O cord (Oicord) cells. A possibility of a compound cold antibody anti IH was made and A1B compatible cells were transfused to the patient. This case report illustrates anti-IH cold agglutinin with broad thermal amplitude. Uniqueness of this case report was O group incompatibility with A1B group, which was detected earlier and a catastrophic transfusion reaction being subverted.

  13. Anti IH: An antibody worth mention

    Directory of Open Access Journals (Sweden)

    Nithya Mohanan

    2016-01-01

    Full Text Available A 72-year-old female with co-morbidities posted for surgical correction of fracture neck of femur without any history of transfusions was noted to have a hemoglobin level of 7 g/dl and packed red blood cells transfusion was ordered. Pretransfusion tests demonstrated A1B group with D positive on forward grouping. Reverse grouping showed a varying grade of agglutination with A, B, and O cells. Agglutination being stronger at 4°C. Antibody screening showed pan-agglutination, direct Coomb's test and auto control were negative. The serum reacted with adult O cells (OIadult but not with adult Bombay cells (Oh Iadult or O cord (Oicord cells. A possibility of a compound cold antibody anti IH was made and A1B compatible cells were transfused to the patient. This case report illustrates anti-IH cold agglutinin with broad thermal amplitude. Uniqueness of this case report was O group incompatibility with A1B group, which was detected earlier and a catastrophic transfusion reaction being subverted.

  14. Antibody responses in allogeneic radiation chimeras

    International Nuclear Information System (INIS)

    The construction of long-lived allogeneic radiation chimeras, free of graft-versus-host disease, has been achieved using serologic elimination of Thy 1+ cells from donor bone marrow. Humoral immune function was not restored in these animals as evidenced by lack of primary antibody responses to a T cell-dependent antigen, namely, sheep erythrocytes (SRBC) both in vivo and in vitro. No evidence for a suppressor cell-mediated mechanism was found. Using separated chimera spleen cell populations and specific helper cell soluble mediators, the functional capabilities of chimera B cells, T cells, and macrophages were assessed. These findings suggested that the failure of chimeras to produce antibody is not the result of impaired B cell, T cell, or macrophage function, but rather, that it is due to ineffective cellular interactions. Physiologic cellular interactions depend upon the sharing of major histocompatibility complex (MHC) determinants between interacting cells. However, the self-recognition repertoire of developing T cells may be influenced by the environment which these cells differentiate such that they learn to recognize host MHC determinants as self. These findings support the interpretation that the immunologic hyporeactivity of allogeneic bone marrow chimeras reflects the role of the host environment in restricting the interactive capabilities of donor-derived cells

  15. Seropositivity of Dengue Antibodies during Pregnancy

    Directory of Open Access Journals (Sweden)

    Nor Azlin Mohamed Ismail

    2014-01-01

    Full Text Available Purpose. Malaysia a dengue endemic country with dengue infections in pregnancy on the rise. The present study was aimed at determining dengue seroprevalence (IgG or IgM during pregnancy and its neonatal transmission in dengue seropositive women. Methods. Maternal with paired cord blood samples were tested for dengue antibodies (IgG and IgM using an enzyme-linked immunosorbent assay (ELISA. Maternal age, parity, occupation, ethnic group, and gestational age were recorded. Data on neonatal Apgar score and admissions to the Neonatal Intensive Care Unit (NICU were analyzed. Results. Out of 358 women recruited, about 128 (35.8% patients were seropositive. Twelve patients (3.4% had recent infections (IgM positive and another 116 women (32.4% were with past infections (IgG positive. All babies born to seropositive mothers had positive IgG paired cord blood; however, no IgM seropositivity was observed. All neonates had good Apgar scores and did not require NICU admission. Conclusion. In this study, 35.8% pregnant women were found to be dengue seropositive. However, transplacental transfer of IgG antibodies had no detrimental effect on the neonatal outcomes.

  16. Thyroid antibody-negative euthyroid Graves’ ophthalmopathy

    Directory of Open Access Journals (Sweden)

    Arshiya Tabasum

    2016-06-01

    Full Text Available TSH receptor antibodies (TRAbs are the pathological hallmark of Graves’ disease, present in nearly all patients with the disease. Euthyroid Graves’ ophthalmopathy (EGO is a well-recognized clinical entity, but its occurrence in patients with negative TRAbs is a potential source of diagnostic confusion. A 66-year-old female presented to our endocrinology clinic with right eye pain and diplopia in the absence of thyroid dysfunction. TRAbs were negative, as measured with a highly sensitive third-generation thyrotropin-binding inhibitory immunoglobulin (TBII ELISA assay. CT and MRI scans of the orbit showed asymmetrical thickening of the inferior rectus muscles but no other inflammatory or malignant orbital pathology. Graves’ ophthalmopathy (GO was diagnosed on the basis of the clinical and radiological features, and she underwent surgical recession of the inferior rectus muscle with complete resolution of the diplopia and orbital pain. She remained euthyroid over the course of follow-up but ultimately developed overt clinical and biochemical hyperthyroidism, 24 months after the initial presentation. By this time, she had developed positive TRAb as well as thyroid peroxidase antibodies. She responded to treatment with thionamides and remains euthyroid. This case highlights the potential for negative thyroid-specific autoantibodies in the presentation of EGO and underscores the variable temporal relationship between the clinical expression of thyroid dysfunction and orbital disease in the natural evolution of Graves’ disease.

  17. IgE antibodies against snake venoms.

    Science.gov (United States)

    Alonso, A; Scavini, L M; Marino, G A; Rodríguez, S M

    1995-01-01

    A similar event was detected in the clinical records of a small group of atopic patients living in the northern provinces of Argentina, i.e., they were bitten by a snake of the Bothrops species (or yarará) during their rural activities (woodcutters, cattle-drivers and farmers). Those who were bitten twice suffered an acute episode of hives and angioedema within 15 minutes after the snake bite. The presence of specific antibodies against Bothrops alternata (Ba) extract was detected by means of RAST for IgE and Ouchterlony and Boyden for IgG. The Ouchterlony also demonstrated crossreactivity among the venoms of the Bothrops species and the positivity of the six fractions obtained by DEAE-cellulose column fractionation against the horse anti-Ba serum. The Ba antigen induced a definite inhibition of the RAST. We presume that hives and angioedema in atopic patients immediately after a second snake bite could be attributed to the presence of a specific IgE antibody against the venom, and must not be misinterpreted with the toxic effects that appear later. PMID:7551202

  18. Licensed monoclonal antibodies and associated challenges.

    Science.gov (United States)

    Khan, Amjad Hayat; Sadroddiny, Esmaeil

    2015-12-23

    Monoclonal antibodies (mAbs) are the leading class of targeted therapeutics and remarkably effective in addressing autoimmune diseases, inflammations, infections, and various types of cancer. Several mAbs approved by US food and drug administration (FDA), are available on the market and a number are pending for approval. Luckily, FDA approved mAbs have played a pivotal role in the treatment and prevention of lethal diseases. However, claiming that licensed mAbs are 100% safe is still debatable, because infections, malignancies, anaphylactoid, and anaphylactic reactions are the more frequently associated adverse events. To evaluate benefit to risk ratio of mAbs, it is important for the clinical research staff or physicians to monitor and follow-up the patients who are receiving mAbs dozes. It is recommended that patients, physicians, biopharmaceutical companies, and researchers should keep in touch to highlight and resolve antibody-based adverse events. In this review we underscore the associated challenges of mAbs, approved by FDA from 2007-2014. PMID:27472864

  19. Antibody-Mediated Autoimmune Encephalopathies and Immunotherapies.

    Science.gov (United States)

    Gastaldi, Matteo; Thouin, Anaïs; Vincent, Angela

    2016-01-01

    Over the last 15 years it has become clear that rare but highly recognizable diseases of the central nervous system (CNS), including newly identified forms of limbic encephalitis and other encephalopathies, are likely to be mediated by antibodies (Abs) to CNS proteins. The Abs are directed against membrane receptors and ion channel-associated proteins that are expressed on the surface of neurons in the CNS, such as N-methyl D-aspartate receptors and leucine-rich, glioma inactivated 1 protein and contactin-associated protein like 2, that are associated with voltage-gated potassium channels. The diseases are not invariably cancer-related and are therefore different from the classical paraneoplastic neurological diseases that are associated with, but not caused by, Abs to intracellular proteins. Most importantly, the new antibody-associated diseases almost invariably respond to immunotherapies with considerable and sometimes complete recovery, and there is convincing evidence of their pathogenicity in the relatively limited studies performed so far. Treatments include first-line steroids, intravenous immunoglobulins, and plasma exchange, and second-line rituximab and cyclophosphamide, followed in many cases by steroid-sparing agents in the long-term. This review focuses mainly on N-methyl D-aspartate receptor- and voltage-gated potassium channel complex-related Abs in adults, the clinical phenotypes, and treatment responses. Pediatric cases are referred to but not reviewed in detail. As there have been very few prospective studies, the conclusions regarding immunotherapies are based on retrospective studies. PMID:26692392

  20. A Neutralizing Antibody Assay Based on a Reporter of Antibody-Dependent Cell-Mediated Cytotoxicity.

    Science.gov (United States)

    Wu, Yuling; Li, Jia J; Kim, Hyun Jun; Liu, Xu; Liu, Weiyi; Akhgar, Ahmad; Bowen, Michael A; Spitz, Susan; Jiang, Xu-Rong; Roskos, Lorin K; White, Wendy I

    2015-11-01

    Benralizumab is a humanized anti-IL5 receptor α (IL5Rα) monoclonal antibody (mAb) with enhanced (afucosylation) antibody-dependent cell-mediated cytotoxicity (ADCC) function. An ADCC reporter cell-based neutralizing antibody (NAb) assay was developed and characterized to detect NAb against benralizumab in human serum to support the clinical development of benralizumab. The optimal ratio of target cells to effector cells was 3:1. Neither parental benralizumab (fucosylated) nor benralizumab Fab resulted in ADCC activity, confirming the requirement for ADCC activity in the NAb assay. The serum tolerance of the cells was determined to be 2.5%. The cut point derived from normal and asthma serum samples was comparable. The effective range of benralizumab was determined, and 35 ng/mL [80% maximal effective concentration (EC80)] was chosen as the standard concentration to run in the assessment of NAb. An affinity purified goat anti-benralizumab polyclonal idiotype antibody preparation was shown to have NAb since it inhibited ADCC activity in a dose-dependent fashion. The low endogenous concentrations of IL5 and soluble IL5 receptor (sIL5R) did not demonstrate to interfere with the assay. The estimated assay sensitivities at the cut point were 1.02 and 1.10 μg/mL as determined by the surrogate neutralizing goat polyclonal and mouse monoclonal anti-drug antibody (ADA) controls, respectively. The assay can detect NAb (at 2.5 μg/mL) in the presence of 0.78 μg/mL benralizumab. The assay was not susceptible to non-specific matrix effects. This study provides an approach and feasibility of developing an ADCC cell-based NAb assay to support biopharmaceuticals with an ADCC function. PMID:26205082

  1. Role of antiviral antibodies in resistance against coxsackievirus B3 infection: interaction between preexisting antibodies and an interferon inducer.

    OpenAIRE

    Cho, C T; Feng, K. K.; McCarthy, V P; Lenahan, M F

    1982-01-01

    An experimental model of coxsackievirus B3 infection in newborn mice was utilized to examine the protective role of antiviral antibodies and an interferon inducer, polyinosinic acid-polycytidylic acid [poly(I:C)]. Subcutaneous administration to the infected mice of specific antiviral antibodies resulted in significant protection against coxsackievirus B3 infection. Antibody-treated animals had shortened viremia, early clearance of virus from tissues, and a reduced mortality rate. Dose respons...

  2. DARPins: a true alternative to antibodies.

    Science.gov (United States)

    Stumpp, Michael T; Amstutz, Patrick

    2007-03-01

    Designed ankyrin repeat proteins (DARPins) are a promising class of non-immunoglobulin proteins that can offer advantages over antibodies for target binding in drug discovery and drug development. DARPins have been successfully used, for example, for the inhibition of kinases, proteases and drug-exporting membrane proteins. DARPins specifically targeting the cancer marker HER2 have also been generated and were shown to function in both in vitro diagnostics and in vivo tumor targeting. DARPins are ideally suited for in vivo imaging or delivery of toxins or other therapeutic payloads because of their favorable molecular properties, including small size and high stability. The low-cost production in bacteria and the rapid generation of many target-specific DARPins make the DARPin approach useful for drug discovery. Additionally, DARPins can be easily generated in multispecific formats, offering the potential to target an effector DARPin to a specific organ or to target multiple receptors with one molecule composed of several DARPins.

  3. SPECT assay of radiolabeled monoclonal antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Jaszczak, R.J.

    1992-02-01

    The long-term goal of this research project is to develop methods to improve the utility of single photon emission computed tomography (SPECI) to quantify the biodistribution of monoclonal antibodies (MoAbs) labeled with clinically relevant radionuclides ({sup 123}I, {sup 131}I, and {sup 111}In) and with another radionuclide,{sup 211}At, recently used in therapy. We describe here our progress in developing quantitative SPECT methodology for {sup 111}In and {sup 123}I. We have focused our recent research thrusts on the following aspects of SPECT: (1) The development of improved SPECT hardware, such as improved acquisition geometries. (2) The development of better reconstruction methods that provide accurate compensation for the physical factors that affect SPECT quantification. (3) The application of carefully designed simulations and experiments to validate our hardware and software approaches.

  4. DARPins: a true alternative to antibodies.

    Science.gov (United States)

    Stumpp, Michael T; Amstutz, Patrick

    2007-03-01

    Designed ankyrin repeat proteins (DARPins) are a promising class of non-immunoglobulin proteins that can offer advantages over antibodies for target binding in drug discovery and drug development. DARPins have been successfully used, for example, for the inhibition of kinases, proteases and drug-exporting membrane proteins. DARPins specifically targeting the cancer marker HER2 have also been generated and were shown to function in both in vitro diagnostics and in vivo tumor targeting. DARPins are ideally suited for in vivo imaging or delivery of toxins or other therapeutic payloads because of their favorable molecular properties, including small size and high stability. The low-cost production in bacteria and the rapid generation of many target-specific DARPins make the DARPin approach useful for drug discovery. Additionally, DARPins can be easily generated in multispecific formats, offering the potential to target an effector DARPin to a specific organ or to target multiple receptors with one molecule composed of several DARPins. PMID:17436550

  5. SPECT assay of radiolabeled monoclonal antibodies

    International Nuclear Information System (INIS)

    The long-term goal of this research project is to develop methods to improve the utility of single photon emission computed tomography (SPECI) to quantify the biodistribution of monoclonal antibodies (MoAbs) labeled with clinically relevant radionuclides (123I, 131I, and 111In) and with another radionuclide,211At, recently used in therapy. We describe here our progress in developing quantitative SPECT methodology for 111In and 123I. We have focused our recent research thrusts on the following aspects of SPECT: (1) The development of improved SPECT hardware, such as improved acquisition geometries. (2) The development of better reconstruction methods that provide accurate compensation for the physical factors that affect SPECT quantification. (3) The application of carefully designed simulations and experiments to validate our hardware and software approaches

  6. Monoclonal antibodies for the control of influenza virus vaccines.

    NARCIS (Netherlands)

    H.J.M. van de Donk; M.F. van Olderen; A.D.M.E. Osterhaus (Albert); J.C. de Jong (Jan)

    1984-01-01

    textabstractHybridomas producing haemagglutination inhibiting monoclonal antibodies against influenza A/Texas/1/77 H3N2 were developed. One hybridoma producing antibodies reacting with Victoria/3/75, Texas/1/77 Bangkok/1/79 and England/496/80 was selected to determine the potency of influenza virusv

  7. Detection of novel diagnostic antibodies in ankylosing spondylitis: An overview.

    Science.gov (United States)

    Quaden, Dana H F; De Winter, Liesbeth M; Somers, Veerle

    2016-08-01

    Ankylosing spondylitis (AS) is a debilitating, chronic, rheumatic disease characterized by inflammation and new bone formation resulting in fusion of the spine and sacroiliac joints. Since early treatment is impeded by a delayed diagnosis, it is highly important to find new biomarkers that improve early diagnosis and may also contribute to a better assessment of disease activity, prognosis and therapy response in AS. Because of the absence of rheumatoid factor, AS was long assumed to have a seronegative character and antibodies are thus not considered a hallmark of the disease. However, emerging evidence suggests plasma cells and autoantibodies to be involved in the disease course. In this review, the role of B cells and antibodies in AS is discussed. Furthermore, an overview is provided of antibodies identified in AS up till now, and their diagnostic potential. Many of these antibody responses were based on small study populations and further validation is lacking. Moreover, most were identified by a hypothesis-driven approach and thus limited to antibodies against targets that are already known to be involved in AS pathogenesis. Hence, we propose an unbiased approach to identify novel diagnostic antibodies. The already successfully applied techniques cDNA phage display and serological antigen selection will be used to identify antibodies against both known and new antigen targets in AS plasma. These newly identified antibodies will enhance early diagnosis of AS and provide more insight into the underlying disease pathology, resulting in a more effective treatment strategy and eventually an improved disease outcome. PMID:27288842

  8. Monoclonal antibodies to Herpes Simplex Virus Type 2

    International Nuclear Information System (INIS)

    In this thesis the production and characterisation of monoclonal antibodies to Herpes Simplex Virus Type 2 is described. The development of a suitable radioimmunoassay for the detection of anti-HSV-2 antibodies, and the selection of an optimal immunisation schedule, is given. Three assay systems are described and their reliability and sensitivity compared. (Auth.)

  9. Human-like antibodies neutralizing Western equine encephalitis virus

    Science.gov (United States)

    Hülseweh, Birgit; Rülker, Torsten; Pelat, Thibaut; Langermann, Claudia; Frenzel, Andrè; Schirrmann, Thomas; Dübel, Stefan; Thullier, Philippe; Hust, Michael

    2014-01-01

    This study describes the development of the first neutralizing antibodies against Western equine encephalitis virus (WEEV), a member of the genus Alphavirus. WEEV is transmitted by mosquitoes and can spread to the human central nervous system, causing symptoms ranging from mild febrile reactions to life-threatening encephalitis. WEEV has been classified as a biological warfare agent by the US Centers for Disease Control and Prevention. No anti-WEEV drugs are currently commercially available. Neutralizing antibodies are useful for the pre- and post-exposure treatment of WEEV infections. In this study, two immune antibody gene libraries were constructed from two macaques immunized with inactivated WEEV. Four antibodies were selected from these libraries and recloned as scFv-Fc, with a human Fc part. These antibodies bound WEEV specifically in ELISA with little or no cross-reaction with other alphaviruses. They were further analyzed by immunohistochemistry. All binders were suitable for the intracellular detection of WEEV particles. Neutralizing activity was determined in vitro. Three of the four antibodies were found to be neutralizing; about 1 ng/mL of the best antibody (ToR69–3A2) neutralized 50% of 5x104 TCID50/mL. Due to its human-like nature with a germinality index of 89% (VH) and 91% (VL), the ToR69–3A2 antibody is a promising candidate for future passive vaccine development. PMID:24518197

  10. Structural Comparison of Different Antibodies Interacting with Parvovirus Capsids

    Energy Technology Data Exchange (ETDEWEB)

    Hafenstein, Susan; Bowman, Valorie D.; Sun, Tao; Nelson, Christian D.S.; Palermo, Laura M.; Chipman, Paul R.; Battisti, Anthony J.; Parrish, Colin R.; Rossmann, Michael G.; Cornell; Purdue

    2009-05-13

    The structures of canine parvovirus (CPV) and feline parvovirus (FPV) complexed with antibody fragments from eight different neutralizing monoclonal antibodies were determined by cryo-electron microscopy (cryoEM) reconstruction to resolutions varying from 8.5 to 18 {angstrom}. The crystal structure of one of the Fab molecules and the sequence of the variable domain for each of the Fab molecules have been determined. The structures of Fab fragments not determined crystallographically were predicted by homology modeling according to the amino acid sequence. Fitting of the Fab and virus structures into the cryoEM densities identified the footprints of each antibody on the viral surface. As anticipated from earlier analyses, the Fab binding sites are directed to two epitopes, A and B. The A site is on an exposed part of the surface near an icosahedral threefold axis, whereas the B site is about equidistant from the surrounding five-, three-, and twofold axes. One antibody directed to the A site binds CPV but not FPV. Two of the antibodies directed to the B site neutralize the virus as Fab fragments. The differences in antibody properties have been linked to the amino acids within the antibody footprints, the position of the binding site relative to the icosahedral symmetry elements, and the orientation of the Fab structure relative to the surface of the virus. Most of the exposed surface area was antigenic, although each of the antibodies had a common area of overlap that coincided with the positions of the previously mapped escape mutations.

  11. Monoclonal Antibodies Specific for Hippurate Hydrolase of Campylobacter jejuni

    OpenAIRE

    Steele, Marina; Gyles, Carlton; Chan, Voon Loong; Odumeru, Joseph

    2002-01-01

    Eleven monoclonal antibodies raised against recombinant Campylobacter jejuni hippurate hydrolase were tested for binding to lysates from 19 C. jejuni strains, 12 other Campylobacter strains, and 21 non-Campylobacter strains. Several monoclonal antibodies bound to C. jejuni but not to other Campylobacter species and may be useful in a species-specific immunoassay.

  12. High throughput production of mouse monoclonal antibodies using antigen microarrays

    DEFF Research Database (Denmark)

    De Masi, Federico; Chiarella, P.; Wilhelm, H.;

    2005-01-01

    Recent advances in proteomics research underscore the increasing need for high-affinity monoclonal antibodies, which are still generated with lengthy, low-throughput antibody production techniques. Here we present a semi-automated, high-throughput method of hybridoma generation and identification...

  13. Antibody function in neutralization and protection against HIV-1

    NARCIS (Netherlands)

    Hessell, A.J.

    2009-01-01

    The ability to induce neutralizing antibodies is generally thought to be of great importance for vaccine efficacy. In HIV-1 research this quality has been elusive as the HIV-1 virus has evolved multiple mechanisms to evade neutralizing antibodies. This thesis traces studies with four broadly neutral

  14. Plasma antibody levels in periodontitis patients and controls

    NARCIS (Netherlands)

    Graswinckel, JEM; van der Velden, U; van Winkelhoff, AJ; Hoek, FJ; Loos, BG

    2004-01-01

    Background: A major aspect of the adaptive host response in periodontitis is the production of antibodies. Several risk and susceptibility factors for periodontitis, including smoking, age and composition of the subgingival microflora, have also been suggested to influence antibody production. Aim:

  15. Anticarbohydrate antibodies in patients with inflammatory bowel disease

    Directory of Open Access Journals (Sweden)

    Mohammed Fakhraldeen

    2010-06-01

    Full Text Available Evaluating the prevalence of antichitobioside carbohydrate antibody (ACCA, antilaminaribioside carbohydrate antibodies (ALCA, antimannobioside carbohydrate antibodies (AMCA and anti-Saccharomyces cerevisiae antibodies (ASCA, in patients with inflammatory bowel disease (IBD. Subjects and methods: 268 serum samples were used; 115 Crohn's disease (CD, 83 ulcerative colitis, and 70 healthy control samples. All samples were evaluated using enzyme-linked immunosorbent assay for the following four anticarbohydrate antibodies: ACCA, ALCA, AMCA, and ASCA. Results: In patients with Crohn's disease the prevalence of the anticarbohydrate antibodies was: ASCA 69%, AMCA 32%, ACCA 28% and ALCA 24% with the highest prevalence being for ASCA (P-value<0.0001 while in patients with ulcerative colitis the prevalence was: ACCA 46%, AMCA 35%, ALCA 23% and ASCA 15% with the highest prevalence being for ACCA (P-value<0.001. Conclusion: Anticarbohydrate antibodies are significantly present in patients with IBD. The use of a panel of anticarbohydrate antibodies may provide additional help in distinguishing IBD from non-IBD disease patterns and narrow the range of differential diagnosis in these patients.

  16. Antibody and B cell responses to Plasmodium sporozoites

    Directory of Open Access Journals (Sweden)

    Johanna N Dups

    2014-11-01

    Full Text Available Antibodies are capable of blocking infection of the liver by Plasmodium sporozoites. Accordingly the induction of anti-sporozoite antibodies is a major aim of various vaccine approaches to malaria. In recent years our knowledge of the specificity and quantities of antibodies required for protection has been greatly expanded by clinical trials of various whole sporozoite and subunit vaccines. Moreover, the development of humanized mouse models and transgenic parasites have also aided our ability to assess the specificity of antibodies and their ability to block infection. Nonetheless, considerable gaps remain in our knowledge - in particular in understanding what antigens are recognized by infection blocking antibodies and in knowing how we can induce robust, long-lived antibody responses. Maintaining high levels of circulating antibodies is likely to be of primary importance, as antibodies must block infection in the short time it takes for sporozoites to reach the liver from the skin. It is clear that a better understanding of the development of protective B cell-mediated immunity will aid the development and refinement of malaria vaccines.

  17. Detection of novel diagnostic antibodies in ankylosing spondylitis: An overview.

    Science.gov (United States)

    Quaden, Dana H F; De Winter, Liesbeth M; Somers, Veerle

    2016-08-01

    Ankylosing spondylitis (AS) is a debilitating, chronic, rheumatic disease characterized by inflammation and new bone formation resulting in fusion of the spine and sacroiliac joints. Since early treatment is impeded by a delayed diagnosis, it is highly important to find new biomarkers that improve early diagnosis and may also contribute to a better assessment of disease activity, prognosis and therapy response in AS. Because of the absence of rheumatoid factor, AS was long assumed to have a seronegative character and antibodies are thus not considered a hallmark of the disease. However, emerging evidence suggests plasma cells and autoantibodies to be involved in the disease course. In this review, the role of B cells and antibodies in AS is discussed. Furthermore, an overview is provided of antibodies identified in AS up till now, and their diagnostic potential. Many of these antibody responses were based on small study populations and further validation is lacking. Moreover, most were identified by a hypothesis-driven approach and thus limited to antibodies against targets that are already known to be involved in AS pathogenesis. Hence, we propose an unbiased approach to identify novel diagnostic antibodies. The already successfully applied techniques cDNA phage display and serological antigen selection will be used to identify antibodies against both known and new antigen targets in AS plasma. These newly identified antibodies will enhance early diagnosis of AS and provide more insight into the underlying disease pathology, resulting in a more effective treatment strategy and eventually an improved disease outcome.

  18. MONOCLONAL ANTIBODIES TO IDENTIFY TOMATO MOSAIC TOBAMOVIRUS (TOMV

    Directory of Open Access Journals (Sweden)

    Duarte Keila M.R.

    2001-01-01

    Full Text Available Monoclonal antibodies were obtained against Tomato mosaic tobamovirus (ToMV isolated in Brazil. One antibody (8G7G2 isotyped as IgG2b (kappa light chain showed strong specificity and very low cross reaction with the Tobacco mosaic virus (TMV. It can be used in identification of tomato mosaic virus (ToMV.

  19. Antibodies to Berne virus in horses and other animals

    NARCIS (Netherlands)

    Horzinek, M.C.; Weiss, Marianne; Steck, F.; Kaderli, R.

    1984-01-01

    After inoculation into 2 foals, Berne virus induced neutralizing antibody, but did not cause clinical symptoms. In a horizontal study of seropositive mares and their offspring, a decline of maternal antibodies and a sudden synchronous seroconversion in all foals were observed, again without clinical

  20. Antibody phage display applications for nuclear medicine imaging and therapy

    Energy Technology Data Exchange (ETDEWEB)

    Winthrop, M.D.; Denardo, G.L.; Denardo, S.J. [Sacramento Univ. of California Davis Medical Center, Sacramento, CA (United States). Dept. of Internal Medicine, Div. of Radiodiagnosis and Terapy

    2000-09-01

    Antibody-based constructs genetically engineered from genes of diverse origin provide a remarkable opportunity to develop functional molecular imaging techniques and specific molecular targeted radionuclide therapies. Phage display libraries of antibody fragment genes can be used to select antibody-based constructs that bind any chosen epitope. A large naive human antibody-based library was used to illustrate binding of antibody constructs to a variety of common and unique antigens. Antibody-based libraries from hybridoma cells, lymphocytes from immunized humans or from mice and human antibody repertoires produced in transgenic mice have also been described. Several orders of magnitude of affinity enhancement can be achieved by random or site specific mutations of the selected binding peptide domains of the scFv. Affinities (K{sub d}) as high as 10{sup -}11 M (10 pM) for affinity-matured scFv have been documented. Such gene libraries thus offer an almost limitless variety of antibody-based molecular binding peptide modules that can be used in creative ways for the construction of new targeting agents for functional or molecular imaging and therapy.

  1. Anti-GM-CSF antibodies in paediatric pulmonary alveolar proteinosis

    OpenAIRE

    Latzin, P; Tredano, M.; Wust, Y; J. de Blic; Nicolai, T; Bewig, B; Stanzel, F.; Kohler, D.; Bahuau, M.; Griese, M

    2005-01-01

    Background: Auto-antibodies against granulocyte-macrophage colony stimulating factor (GM-CSF) may be central to the pathogenesis of adult sporadic pulmonary alveolar proteinosis (PAP). The role of anti-GM-CSF auto-antibodies in paediatric forms of PAP is as yet unclear.

  2. Cell-Free Synthesis Meets Antibody Production: A Review

    Directory of Open Access Journals (Sweden)

    Marlitt Stech

    2015-01-01

    Full Text Available Engineered antibodies are key players in therapy, diagnostics and research. In addition to full size immunoglobulin gamma (IgG molecules, smaller formats of recombinant antibodies, such as single-chain variable fragments (scFv and antigen binding fragments (Fab, have emerged as promising alternatives since they possess different advantageous properties. Cell-based production technologies of antibodies and antibody fragments are well-established, allowing researchers to design and manufacture highly specific molecular recognition tools. However, as these technologies are accompanied by the drawbacks of being rather time-consuming and cost-intensive, efficient and powerful cell-free protein synthesis systems have been developed over the last decade as alternatives. So far, prokaryotic cell-free systems have been the focus of interest. Recently, eukaryotic in vitro translation systems have enriched the antibody production pipeline, as these systems are able to mimic the natural pathway of antibody synthesis in eukaryotic cells. This review aims to overview and summarize the advances made in the production of antibodies and antibody fragments in cell-free systems.

  3. Postbooster Antibodies from Humans as Source of Diphtheria Antitoxin

    OpenAIRE

    Bermejo-Martin, Jesús F; Avila-Alonso, Ana; González-Rivera, Milagros; Tamayo, Eduardo; Eiros, Jose María; Almansa, Raquel

    2016-01-01

    Diphtheria antitoxin for therapeutic use is in limited supply. A potential source might be affinity-purified antibodies originally derived from plasma of adults who received a booster dose of a vaccine containing diphtheria toxoid. These antibodies might be useful for treating even severe cases of diphtheria.

  4. Postbooster Antibodies from Humans as Source of Diphtheria Antitoxin.

    Science.gov (United States)

    Bermejo-Martin, Jesús F; Avila-Alonso, Ana; González-Rivera, Milagros; Tamayo, Eduardo; Eiros, Jose María; Almansa, Raquel

    2016-07-01

    Diphtheria antitoxin for therapeutic use is in limited supply. A potential source might be affinity-purified antibodies originally derived from plasma of adults who received a booster dose of a vaccine containing diphtheria toxoid. These antibodies might be useful for treating even severe cases of diphtheria. PMID:27314309

  5. Development of Biodegradable Nanocarriers Loaded with a Monoclonal Antibody

    Directory of Open Access Journals (Sweden)

    Andrew Gdowski

    2015-02-01

    Full Text Available Treatments utilizing monoclonal antibody therapeutics against intracellular protein-protein interactions in cancer cells have been hampered by several factors, including poor intracellular uptake and rapid lysosomal degradation. Our current work examines the feasibility of encapsulating monoclonal antibodies within poly(lactic-co-glycolic acid (PLGA nanoparticles using a water/oil/water double emulsion solvent evaporation technique. This method can be used to prepare protective polymeric nanoparticles for transporting functional antibodies to the cytoplasmic compartment of cancer cells. Nanoparticles were formulated and then characterized using a number of physical and biological parameters. The average nanoparticle size ranged from 221 to 252 nm with a low polydispersity index. Encapsulation efficiency of 16%–22% and antibody loading of 0.3%–1.12% were observed. The antibody molecules were released from the nanoparticles in a sustained manner and upon release maintained functionality. Our studies achieved successful formulation of antibody loaded polymeric nanoparticles, thus indicating that a PLGA-based antibody nanoformulation is a promising intracellular delivery vehicle for a large number of new intracellular antibody targets in cancer cells.

  6. Functionally fused antibodies--a novel adjuvant fusion system

    DEFF Research Database (Denmark)

    Larsen, Martin; Jensen, Kim Bak; Christensen, Peter Astrup;

    2008-01-01

    Antibodies capable of recognizing key molecular targets isolated e.g. by phage display technology have been used in the pursuit of new and improved therapies for prevalent human diseases. These approaches often take advantage of non-immunogenic antibody fragments to achieve specific toxin-, radio...

  7. Dissecting antibodies with regards to linear and conformational epitopes.

    Science.gov (United States)

    Forsström, Björn; Axnäs, Barbara Bisławska; Rockberg, Johan; Danielsson, Hanna; Bohlin, Anna; Uhlen, Mathias

    2015-01-01

    An important issue for the performance and specificity of an antibody is the nature of the binding to its protein target, including if the recognition involves linear or conformational epitopes. Here, we dissect polyclonal sera by creating epitope-specific antibody fractions using a combination of epitope mapping and an affinity capture approach involving both synthesized peptides and recombinant protein fragments. This allowed us to study the relative amounts of antibodies to linear and conformational epitopes in the polyclonal sera as well as the ability of each antibody-fraction to detect its target protein in Western blot assays. The majority of the analyzed polyclonal sera were found to have most of the target-specific antibodies directed towards linear epitopes and these were in many cases giving Western blot bands of correct molecular weight. In contrast, many of the antibodies towards conformational epitopes did not bind their target proteins in the Western blot assays. The results from this work have given us insights regarding the nature of the antibody response generated by immunization with recombinant protein fragments and has demonstrated the advantage of using antibodies recognizing linear epitopes for immunoassay involving wholly or partially denatured protein targets. PMID:25816293

  8. Dissecting antibodies with regards to linear and conformational epitopes.

    Directory of Open Access Journals (Sweden)

    Björn Forsström

    Full Text Available An important issue for the performance and specificity of an antibody is the nature of the binding to its protein target, including if the recognition involves linear or conformational epitopes. Here, we dissect polyclonal sera by creating epitope-specific antibody fractions using a combination of epitope mapping and an affinity capture approach involving both synthesized peptides and recombinant protein fragments. This allowed us to study the relative amounts of antibodies to linear and conformational epitopes in the polyclonal sera as well as the ability of each antibody-fraction to detect its target protein in Western blot assays. The majority of the analyzed polyclonal sera were found to have most of the target-specific antibodies directed towards linear epitopes and these were in many cases giving Western blot bands of correct molecular weight. In contrast, many of the antibodies towards conformational epitopes did not bind their target proteins in the Western blot assays. The results from this work have given us insights regarding the nature of the antibody response generated by immunization with recombinant protein fragments and has demonstrated the advantage of using antibodies recognizing linear epitopes for immunoassay involving wholly or partially denatured protein targets.

  9. Double-antibody radioimmunoassay for staphylococcal enterotoxin C2

    International Nuclear Information System (INIS)

    A sensitive double-antibody radioimmunoassay for staphylococcal enterotoxin C2 is described. The assay procedure employs anti-rabbit gamma globulin, prepared in goats, to precipitate the antigen-antibody complex of enterotoxin C2 and anti-enterotoxin C2. The test is sensitive to 100 pg of enterotoxin

  10. A Quantitative Method for Comparing the Brightness of Antibody-dye Reagents and Estimating Antibodies Bound per Cell.

    Science.gov (United States)

    Kantor, Aaron B; Moore, Wayne A; Meehan, Stephen; Parks, David R

    2016-01-01

    We present a quantitative method for comparing the brightness of antibody-dye reagents and estimating antibodies bound per cell. The method is based on complementary binding of test and fill reagents to antibody capture microspheres. Several aliquots of antibody capture beads are stained with varying amounts of the test conjugate. The remaining binding sites on the beads are then filled with a second conjugate containing a different fluorophore. Finally, the fluorescence of the test conjugate compared to the fill conjugate is used to measure the relative brightness of the test conjugate. The fundamental assumption of the test-fill method is that if it takes X molecules of one test antibody to lower the fill signal by Y units, it will take the same X molecules of any other test antibody to give the same effect. We apply a quadratic fit to evaluate the test-fill signal relationship across different amounts of test reagent. If the fit is close to linear, we consider the test reagent to be suitable for quantitative evaluation of antibody binding. To calibrate the antibodies bound per bead, a PE conjugate with 1 PE molecule per antibody is used as a test reagent and the fluorescence scale is calibrated with Quantibrite PE beads. When the fluorescence per antibody molecule has been determined for a particular conjugate, that conjugate can be used for measurement of antibodies bound per cell. This provides comparisons of the brightness of different conjugates when conducted on an instrument whose statistical photoelectron (Spe) scales are known. © 2016 by John Wiley & Sons, Inc.

  11. Antibody response and antibody affinity maturation in cats with experimental proliferative immune complex glomerulonephritis.

    Science.gov (United States)

    Bishop, S A; Bailey, M; Lucke, V M; Stokes, C R

    1992-07-01

    An experimental model of proliferative glomerulonephritis (GN) in the cat, which closely resembles human proliferative forms of GN, has been used to study the role of antibody and antibody affinity in the development of immune complex-mediated renal disease. The serum IgG and IgM antibody response to antigen, average antibody affinity (avidity) and affinity heterogeneity of the IgG and IgM populations was assessed at varying times after commencement of chronic immunization with the antigen, human serum albumin (HSA), by enzyme immunoassay. Cats could be classified according to whether they were "low", "intermediate" or "high" IgG responders, by quantification of serum IgG values. Cats with the lowest serum IgG values failed to develop glomerulonephritis. However, there was no relationship between actual IgG values and the severity of the induced disease. In contrast to IgG, there was no division of cats into low or high IgM anti-HSA responders. Again, cats with the lowest IgM values failed to develop GN, but, more interestingly, a late, marked increase in serum IgM anti-HSA occurred only in cats that developed clinical signs of GN (anterior uveitis and nephrotic syndrome). Maturation of average, functional IgG affinity (avidity) for HSA following chronic immunization was clearly demonstrated for all cats. At the end of the experiment, all cats had IgG of high affinity for HSA and the average affinity heterogeneity of the IgG populations was less than in measurements taken earlier. Values of IgG affinity at the end of the experiment were very similar both in cats which developed GN and in those which remained clinically, biochemically and pathologically normal. In contrast to IgG antibody, some cats developed IgM of increased affinity, whilst others produced antibody of reduced affinity, following chronic immunization. There was no correlation between the development of disease and the production of either low or high affinity IgM antibody. Data indicated that an

  12. Novel antimalarial antibodies highlight the importance of the antibody Fc region in mediating protection.

    Science.gov (United States)

    Pleass, Richard J; Ogun, Solabomi A; McGuinness, David H; van de Winkel, Jan G J; Holder, Anthony A; Woof, Jenny M

    2003-12-15

    Parasite drug resistance and difficulties in developing effective vaccines have precipitated the search for alternative therapies for malaria. The success of passive immunization suggests that immunoglobulin (Ig)-based therapies are effective. To further explore the mechanism(s) by which antibody mediates its protective effect, we generated human chimeric IgG1 and IgA1 and a single-chain diabody specific for the C-terminal 19-kDa region of Plasmodium yoelii merozoite surface protein 1 (MSP119), a major target of protective immune responses. These novel human reagents triggered in vitro phagocytosis of merozoites but, unlike their parental mouse IgG2b, failed to protect against parasite challenge in vivo. Therefore, the Fc region appears critical for mediating protection in vivo, at least for this MSP119 epitope. Such antibodies may serve as prototype therapeutic agents, and as useful tools in the development of in vitro neutralization assays with Plasmodium parasites. PMID:12855589

  13. Maturation Pathways of Cross-Reactive HIV-1 Neutralizing Antibodies

    Directory of Open Access Journals (Sweden)

    Dimiter S. Dimitrov

    2009-11-01

    Full Text Available Several human monoclonal antibodies (hmAbs and antibody fragments, including the best characterized in terms of structure-function b12 and Fab X5, exhibit relatively potent and broad HIV-1 neutralizing activity. However, the elicitation of b12 or b12-like antibodies in vivo by vaccine immunogens based on the HIV-1 envelope glycoprotein (Env has not been successful. B12 is highly divergent from the closest corresponding germline antibody while X5 is less divergent. We have hypothesized that the relatively high degree of specific somatic hypermutations may preclude binding of the HIV-1 envelope glycoprotein (Env to closest germline antibodies, and that identifying antibodies that are intermediates in the pathways to maturation could help design novel vaccine immunogens to guide the immune system for their enhanced elicitation. In support of this hypothesis we have previously found that a germline-like b12 (monovalent and bivalent scFv as an Fc fusion protein or IgG lacks measurable binding to an Env as measured by ELISA with a sensitivity in the μM range [1]; here we present evidence confirming and expanding these findings for a panel of Envs. In contrast, a germline-like scFv X5 bound Env with high (nM affinity. To begin to explore the maturation pathways of these antibodies we identified several possible b12 intermediate antibodies and tested their neutralizing activity. These intermediate antibodies neutralized only some HIV-1 isolates and with relatively weak potency. In contrast, germline-like scFv X5 neutralized a subset of the tested HIV-1 isolates with comparable efficiencies to that of the mature X5. These results could help explain the relatively high immunogenicity of the coreceptor binding site on gp120 and the abundance of CD4-induced (CD4i antibodies in HIV-1-infected patients (X5 is a CD4i antibody as well as the maturation pathway of X5. They also can help identify antigens that can bind specifically to b12 germline and

  14. Bispecific antibodies and their use in applied research

    Directory of Open Access Journals (Sweden)

    Harshit Verma

    Full Text Available Bispecific antibodies (BsAb can, by virtue of combining two binding specificities, improve the selectivity and efficacy of antibody-based treatment of human disease. Antibodies with two distinct binding specificities have great potential for a wide range of clinical applications as targeting agents for in vitro and in vivo immunodiagnosis, therapy and for improving immunoassays. They have shown great promise for targeting cytotoxic effector cells, delivering radionuclides, toxins or cytotoxic drugs to specific targets, particularly tumour cells. The development of BsAb research goes through three main stages: chemical cross linking of murine-derived monoclonal antibody, hybrid hybridomas and engineered BsAb. This article is providing the potential applications of bispecific antibodies. [Vet World 2012; 5(12.000: 775-780

  15. Modification of monoclonal antibodies by polymers possessing chelating properties

    Energy Technology Data Exchange (ETDEWEB)

    Torchilin, V.P.; Khaw, B.A.; Klibanov, A.L.; Slinkin, M.A.; Haber, E.; Smirnov, V.N.

    1986-12-01

    This paper describes a basically new approach to obtaining diagnostic antibodies, consisting of a one-point modification of the antibody, without loss of its activity, by a high-molecular-weight synthetic polymer with the ability of effectively chelating ions of heavy metals. As a result of this approach, preparations of active antibodies containing some tens of atoms of the metal per protein molecule can be obtained. The concentration of radioactive metal (/sup 111/In) was determined with a gamma-counter and the Mn and Cd concentrations by spectroscopy. Gel-filtration of polymer-modified antibodies after binding of /sup 111/InCl/sub 3/ is shown. Also, solid-phase radioimmunoassay of antibodies and Fab fragments, native and modified by chelating polymers, is presented.

  16. Prokaryotic Expression and Polyclonal Antibody Preparation of PRL3

    Institute of Scientific and Technical Information of China (English)

    SHEN Xing-gui; LI Wan-nan; WANG Jing; JIANG Yi-qun; LI Qing-shan; FU Xue-qi

    2007-01-01

    Phosphatase of regenerating liver 3(PRL3), which belongs to the superfamily of protein tyrosine phosphatases (PTPs), represents a group of low molecular weight PTPs that participate in tumorigenesis and metastasis processes.Presented here are the results of cloning, prokaryotic expression, purification, and polyclonal antibody preparation of PRL3. To obtain a specific polyclonal antibody against PRL3, the authors have prepared GST-PRL3 to immunize rabbits and purify an anti-PRL3 polyclonal antibody by negative selection affinity columns. Western blot analysis shows that the anti-PRL3 polyclonal antibody has a specific binding ability with PRL3 protein. The anti-PRL3 polyclonal antibody provides a good tool to further study the function of PRL3.

  17. ELISA Detection of Francisella tularensis using Polyclonaland Monoclonal Antibodies

    Directory of Open Access Journals (Sweden)

    Miroslav Pohanka

    2008-09-01

    Full Text Available The mouse monoclonal and polyclonal antibodies were produced for the detection of intracellular pathogenand potential warfare agent Francisella tularensis. Antibody titers obtained were 1:640 for polyclonal antibodiesand 1:320 for monoclonal antibodies. Both antibodies were used in the indirect enzyme-linked immunosorbentassay (ELISA found to detect F. tularensis whole cells. The limit of detection was 5.4×106 CFU/ml for polyclonalantibodies and 6.9×106 CFU/ml for monoclonal antibodies. The value sample could  be distinguished from anyconcentration of another gram-negative bacterium: Escherichia coli.Defence Science Journal, 2008, 58(5, pp.698-702, DOI:http://dx.doi.org/10.14429/dsj.58.1693

  18. [A spectrum of neurological diseases with anti-VGKC antibody].

    Science.gov (United States)

    Arimura, Kimiyoshi; Watanabe, Osamu; Nagado, Tatsui

    2007-11-01

    Anti-VGKC antibody causing peripheral nerve hyperexcitability is already an established clinical entity. Recently, many patients with non-herpetic limbic encephalitis (NHLE) with anti-VGKC antibody have been reported. The characteristic clinical features are low serum Na+ concentration and good response to immunotherapy. Anti-VGK antibody positive NHLE is relatively frequent among immune-mediated NHLE. It is important to know that this disease is responsive to immunotherapy. Furthermore, anti-VGKC antibody is also positive in some intractable epilepsies. These findings suggest that anti-VGKC is correlated with hyperexcitability in both the peripheral and central nervous system and that the spectrum of anti-VGKC antibody syndrome is now expanding.

  19. Phosphokinase antibody arrays on dendron-coated surface.

    Directory of Open Access Journals (Sweden)

    Ju-Won Kwak

    Full Text Available Monitoring protein phosphorylation at the cellular level is important to understand the intracellular signaling. Among the phosphoproteomics methods, phosphokinase antibody arrays have emerged as preferred tools to measure well-characterized phosphorylation in the intracellular signaling. Here, we present a dendron-coated phosphokinase antibody array (DPA in which the antibodies are immobilized on a dendron-coated glass slide. Self-assembly of conically shaped dendrons well-controlled in size and structure resulted in precisely controlled lateral spacing between the immobilized phosphosite-specific antibodies, leading to minimized steric hindrance and improved antigen-antibody binding kinetics. These features increased sensitivity, selectivity, and reproducibility in measured amounts of protein phosphorylation. To demonstrate the utility of the DPA, we generated the phosphorylation profiles of brain tissue samples obtained from Alzheimer's disease (AD model mice. The analysis of the profiles revealed signaling pathways deregulated during the course of AD progression.

  20. The Role of Complement in Antibody Therapy for Infectious Diseases.

    Science.gov (United States)

    Wibroe, Peter P; Helvig, Shen Y; Moein Moghimi, S

    2014-04-01

    The complement system is part of the innate immune system, eliciting central immunoregulatory functions. Detection of foreign surfaces is either achieved through complement-specific patternrecognition molecules or mediated by antigen recognition of antibodies. Immunoglobulin A (IgA), IgG, and IgM all have the potential to initiate a complement response, with the efficiency and response development closely related to the antibody isotype, multimeric state, and degree of glycosylation. A group of serum proteins constitutes the central effector functions of complement, thus allowing direct cell lysis, opsonization, and inflammation. These effector functions can be used in antibody therapies, especially against infectious diseases, as the target membranes lack complement regulatory proteins. The relative contribution of each function and the interplay with direct antibody-mediated clearance is not fully exploited, thus suggesting an option for further rational optimization of antibody therapies.

  1. Chronic Renal Allograft Dysfunction Antibody-Mediated: An Update

    Directory of Open Access Journals (Sweden)

    Maurizio Salvadori,

    2014-07-01

    Full Text Available This paper reviews the most important studies on chronic antibody-mediated rejection (cABMR, which is an important cause of late graft dysfunction after renal transplantation. Several antibodies seem to be responsible for chronic rejection; new techniques have allowed us to identify these antibodies in circulation. The pathogenetic role of the antibodies generally includes the complement pathway, but may also be complement-independent. This paper also examines the pathogenesis of chronic endothelial lesions, as well as the histopathological aspects. Antibodies responsible for chronic rejection may preexist before transplantation or may develop after transplantation. The possible therapeutic approaches are poor and principally based on early identification and desensitisation techniques. New B cell targeting drugs are aimed at an improved control of the relevant condition.

  2. Triiodothyronine uptake test using specific antibody as the secondary binder

    International Nuclear Information System (INIS)

    We describe a specific and reliable triiodithyronine uptake (T3-U) method for the estimation of free thyroxine index(FT4I) using precipitated anti-T3-antibody and second-antibody (anti-rabbit-IgG raised in goat) complex. Since the method measures the partitioning of T3-125I between the binding proteins and antibody, separation of the portion of T3-125I taken up by the antibody from that bound to the binding proteins is achieved by the use of pre-incubated primary and second antibody complex in the form of suspension, and separation of the bound complex was carried out by means of centrifugation. The T3-U method developed is clinically evaluated and compared with the reputed commercial kit and a correlation of 0.96 was obtained. (author)

  3. Development of radioactivity labelling method of new antibody by using the antibody engineering

    International Nuclear Information System (INIS)

    With an aim to develop a method to produce labelled antibodies with low immunogenicity, two recombinant fusion proteins; scFv-His and scFv-MTβ were produced using gene engineering techniques. The former was constructed with scFv-antibody and histidine hexamer, a metal-chelated protein (or peptide). The latter was done with scFv-antibody and β-domain of metallothionein. Then, antigen-binding activity and metal-binding activity of these fusion proteins were determined using gel-filtration chromatography and ELISA. The main antigen-binding activity of scFv-His preparation was detected in a domain of about 25-30 kDa, which agreed with the peak of 29 kDa corresponding to the presumed molecular weight for the protein. Whereas the antigen-binding activity of scFv-MTβ was found in a domain of 30-35 kDa, which agreed with 32 kDa, the presumed molecular weight of scFv-MTβ. Gel-filtration chromatography of scFv-His preparation after the addition of Cu2+ ion revealed an optical absorption at 280 nm and a Cu-peak near at 14 kDa. These results suggested that the metal affinity of the histidine-hexamer was too weak to chelate Cu2+ in a solution. The chromatography of scFv-MTβ preparation added with Cd2+ showed a peak of Cd appeared around a position of about 20 kDa but the peak was not coincident with that of the antigen-binding activity (ca. 30 kDa), suggesting that the present preparation of scFv-MTβ had no Cd-binding activity due to metal-exchange reaction. Based on these results, problems on the production of recombinant scFv-antibody fused with metal-binding domain of cystein-binding type or histidine-binding one were discussed. (M.N.)

  4. Clearance of injected radioactively labelled antibodies to tumour products by liposome-bound second antibodies

    International Nuclear Information System (INIS)

    Liposomes containing anti-goat immunoglobulin were injected 24 h after administration of 125I-labelled goat antibody against the carcinoembryonic antigen (anti-CEA) to groups of nude mice bearing human tumour xenografts, and normal mice. Controls in each group received radioactively labelled anti-CEA only. In liposome-treated mice, blood 125I levels were lower than those of controls 30 min to 24 h after liposome administration, with corresponding accumulation of 125I activity in the liver and spleen for the first 2 h after liposome injection. [14C]Cholesterol or sup(99m)Tc labels in the bilayer were eliminated rapidly from the blood, with uptake in the liver and spleen. In xenograft-bearing mice, 125I activity detected in the tumours up to 6 h after liposome injection was identical to that detected in the tumours of controls. However, 24 h after liposome injection a reduction in the tumour concentration of 125I-labelled anti-CEA was obtained, but the tumour/blood radioactivity was still increased. In two mice given 27 μmol lipid, the blood radioactivity count after 24 h was only 5% of that in the controls. In rabbits, 2 h after administration of liposomes containing anti-goat second antibody, the circulating 125I activity had dropped by 28-40%. The results suggest that administration of liposome-entrapped second antibody approximately 2 h prior to external scintigraphy may clear circulating radioactively labelled primary antibody by up to 50%. (Auth.)

  5. Overcoming Instability of Antibody-Nanomaterial Conjugates: Next Generation Targeted Nanomedicines Using Bispecific Antibodies.

    Science.gov (United States)

    Howard, Christopher B; Fletcher, Nicholas; Houston, Zachary H; Fuchs, Adrian V; Boase, Nathan R B; Simpson, Joshua D; Raftery, Lyndon J; Ruder, Tim; Jones, Martina L; de Bakker, Christopher J; Mahler, Stephen M; Thurecht, Kristofer J

    2016-08-01

    Targeted nanomaterials promise improved therapeutic efficacy, however their application in nanomedicine is limited due to complexities associated with protein conjugations to synthetic nanocarriers. A facile method to generate actively targeted nanomaterials is developed and exemplified using polyethylene glycol (PEG)-functional nanostructures coupled to a bispecific antibody (BsAb) with dual specificity for methoxy PEG (mPEG) epitopes and cancer targets such as epidermal growth factor receptor (EGFR). The EGFR-mPEG BsAb binds with high affinity to recombinant EGFR (KD : 1 × 10(-9) m) and hyperbranched polymer (HBP) consisting of mPEG (KD : 10 × 10(-9) m) and demonstrates higher avidity for HBP compared to linear mPEG. The binding of BsAb-HBP bioconjugate to EGFR on MDA-MB-468 cancer cells is investigated in vitro using a fluorescently labeled polymer, and in in vivo xenograft models by small animal optical imaging. The antibody-targeted nanostructures show improved accumulation in tumor cells compared to non-targeted nanomaterials. This demonstrates a facile approach for tuning targeting ligand density on nanomaterials, by modulating surface functionality. Antibody fragments are tethered to the nanomaterial through simple mixing prior to administration to animals, overcoming the extensive procedures encountered for developing targeted nanomedicines.

  6. Polyclonal anti-idiotypic antibodies mimicking the small cell lung carcinoma antigen cluster-5A interact with a panel of antibodies and induce specific immune response in animals.

    OpenAIRE

    Zwicky, C.; Stahel, R A; Jaksche, H.; Waibel, R.; Lehmann, H. P.; Loibner, H

    1991-01-01

    Polyclonal anti-idiotypic antibodies (ab2) were generated by immunising goats with the murine IgG2a monoclonal antibody SWA20 which recognises the SCLC antigen cluster-5A, a tumour-associated sialoglycoprotein. Ab2 was shown to bind specifically to antibody SWA20, but not to isotype matched control antibodies. Pre-incubation with ab2 completely inhibited target cell binding of antibody SWA20 and of four other antibodies to cluster-5A antigen, while no effect was seen with antibodies to cluste...

  7. Antiphospholipid antibodies in Brazilian hepatitis C virus carriers

    Directory of Open Access Journals (Sweden)

    A.M. Atta

    2008-06-01

    Full Text Available Hepatitis C, a worldwide viral infection, is an important health problem in Brazil. The virus causes chronic infection, provoking B lymphocyte dysfunction, as represented by cryoglobulinemia, non-organ-specific autoantibody production, and non-Hodgkin's lymphoma. The aim of this research was to screen for the presence of antiphospholipid autoantibodies in 109 Brazilian hepatitis C virus carriers without clinical history of antiphospholipid syndrome. Forty healthy individuals were used as the control group. IgA, IgG, and IgM antibodies against cardiolipin and β2-glycoprotein I were measured with an enzyme-linked immunosorbent assay, using a cut-off point of either 20 UPL or 20 SBU. While 24 (22.0% hepatitis C carriers had moderate titers of IgM anticardiolipin antibodies (median, 22.5 MPL; 95%CI: 21.5-25.4 MPL, only three carriers (<3% had IgG anticardiolipin antibodies (median, 23 GPL; 95%CI: 20.5-25.5 GPL. Furthermore, IgA anticardiolipin antibodies were not detected in these individuals. Male gender and IgM anticardiolipin seropositivity were associated in the hepatitis C group (P = 0.0004. IgA anti-β2-glycoprotein-I antibodies were detected in 29 of 109 (27.0% hepatitis C carriers (median, 41 SAU; 95%CI: 52.7-103.9 SAU. Twenty patients (18.0% had IgM anti-β2-glycoprotein I antibodies (median, 27.6 SMU; 95%CI: 23.3-70.3 SMU, while two patients had IgG antibodies against this protein (titers, 33 and 78 SGU. Antiphospholipid antibodies were detected in only one healthy individual, who was seropositive for IgM anticardiolipin. We concluded that Brazilian individuals chronically infected with hepatitis C virus present a significant production of antiphospholipid antibodies, mainly IgA anti-β2-glycoprotein I antibodies, which are not associated with clinical manifestations of antiphospholipid syndrome.

  8. Anti-DNA antibody mediated catalysis is isotype dependent.

    Science.gov (United States)

    Xia, Yumin; Eryilmaz, Ertan; Zhang, Qiuting; Cowburn, David; Putterman, Chaim

    2016-01-01

    Anti-DNA antibodies are the serological hallmark of systemic lupus erythematosus, and participate in the pathogenesis of lupus nephritis by cross-reacting with multiple renal antigens. Previously, using a panel of murine anti-DNA IgGs that share identical variable regions but that differ in the constant regions, we demonstrated that the cross-reaction and renal pathogenicity of anti-DNA antibodies are isotype dependent. In this study, we investigated the catalytic potential of this anti-DNA antibody panel, and determined its isotype dependency. The three isotype switch variants (IgG1, IgG2a, IgG2b) and the parent IgG3 PL9-11 anti-DNA antibodies were compared in their catalysis of 500 base pair linear double stranded DNA and a 12-mer peptide (ALWPPNLHAWVP), by gel analysis, MALDI-TOF mass spectrometry, and nuclear magnetic resonance spectroscopy. The binding affinity of anti-DNA antibodies to double stranded DNA and peptide antigens were assessed by ELISA and surface plasmon resonance. We found that the PL9-11 antibody isotypes vary significantly in their potential to catalyze the cleavage of both linear and double stranded DNA and the proteolysis of peptides. The degree of the cleavage and proteolysis increases with the incubation temperature and time. While different PL9-11 isotypes have the same initial attack sites within the ALWPPNLHAWVP peptide, there was no correlation between binding affinity to the peptide and proteolysis rates. In conclusion, the catalytic properties of anti-DNA antibodies are isotype dependent. This finding provides further evidence that antibodies that share the same variable region, but which have different constant regions, are functionally distinct. The catalytic effects modulated by antibody constant regions need to be considered in the design of therapeutic antibodies (abzymes) and peptides designed to block pathogenic autoantibodies.

  9. Rifampicin-dependent antibodies bind a similar or identical epitope to glycoprotein IX-specific quinine-dependent antibodies

    NARCIS (Netherlands)

    Burgess, J K; Lopez, J A; Gaudry, L E; Chong, B H

    2000-01-01

    The drug-dependent antibody of a patient with rifampicin-induced thrombocytopenia was characterized using the antigen-capture enzyme-linked immunosorbent assay (MAIPA assay), flow cytometry, and immunoprecipitation. The antibody was found to bind glycoprotein (GP) Ib-IX but not GPIIb-IIIa because (1

  10. ANTIBODIES DEFINING RAT ENDOTHELIAL-CELLS - RECA-1, A PAN-ENDOTHELIAL CELL-SPECIFIC MONOCLONAL-ANTIBODY

    NARCIS (Netherlands)

    DUIJVESTIJN, AM; VANGOOR, H; KLATTER, F; MAJOOR, GD; VANBUSSEL, E; VRIESMAN, PJCV

    1992-01-01

    We have been searching for antibodies reactive with rat endothelial cells. Two monoclonal antibodies (mAb), named RECA-1 and RECA-2 were produced and tested in immunoperoxidase staining on frozen sections of various rat tissues. Staining patterns were compared to those obtained with the mAbs OX-2, O

  11. Differentiation of Behcet's disease from inflammatory bowel diseases: Anti-saccharomyces cerevisiae antibody and anti-neutrophilic cytoplasmic antibody

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The differential diagnosis of Behcet's disease (BD) from inflammatory bowel disease (IBD) is sometimes difficult and challenging. Hereby, we suggested the utility of anti-saccharomyces cerevisiae antibody (ASCA) and anti-neutrophilic cytoplasmic antibody (p-ANCA) in the differential diagnosis of BD from IBD.

  12. Utilisation of antibody microarrays for the selection of specific and informative antibodies from recombinant library binders of unknown quality.

    Science.gov (United States)

    Kibat, Janek; Schirrmann, Thomas; Knape, Matthias J; Helmsing, Saskia; Meier, Doris; Hust, Michael; Schröder, Christoph; Bertinetti, Daniela; Winter, Gerhard; Pardes, Khalid; Funk, Mia; Vala, Andrea; Giese, Nathalia; Herberg, Friedrich W; Dübel, Stefan; Hoheisel, Jörg D

    2016-09-25

    Many diagnostic and therapeutic concepts require antibodies of high specificity. Recombinant binder libraries and related selection approaches allow the efficient isolation of antibodies against almost every target of interest. Nevertheless, it cannot be guaranteed that selected antibodies perform well and interact specifically enough with analytes unless an elaborate characterisation is performed. Here, we present an approach to shorten this process by combining the selection of suitable antibodies with the identification of informative target molecules by means of antibody microarrays, thereby reducing the effort of antibody characterisation by concentrating on relevant molecules. In a pilot scheme, a library of 456 single-chain variable fragment (scFv) binders to 134 antigens was used. They were arranged in a microarray format and incubated with the protein content of clinical tissue samples isolated from pancreatic ductal adenocarcinoma and healthy pancreas, as well as recurrent and non-recurrent bladder tumours. We observed significant variation in the expression of the E3 ubiquitin-protein ligase (CHFR) as well as the glutamate receptor interacting protein 2 (GRIP2), for example, always with more than one of the scFvs binding to these targets. Only the relevant antibodies were then characterised further on antigen microarrays and by surface plasmon resonance experiments so as to select the most specific and highest affinity antibodies. These binders were in turn used to confirm a microarray result by immunohistochemistry analysis. PMID:26709003

  13. Combining Phage and Yeast Cell Surface Antibody Display to Identify Novel Cell Type-Selective Internalizing Human Monoclonal Antibodies.

    Science.gov (United States)

    Bidlingmaier, Scott; Su, Yang; Liu, Bin

    2015-01-01

    Using phage antibody display, large libraries can be generated and screened to identify monoclonal antibodies with affinity for target antigens. However, while library size and diversity is an advantage of the phage display method, there is limited ability to quantitatively enrich for specific binding properties such as affinity. One way of overcoming this limitation is to combine the scale of phage display selections with the flexibility and quantitativeness of FACS-based yeast surface display selections. In this chapter we describe protocols for generating yeast surface antibody display libraries using phage antibody display selection outputs as starting material and FACS-based enrichment of target antigen-binding clones from these libraries. These methods should be widely applicable for the identification of monoclonal antibodies with specific binding properties. PMID:26060069

  14. A simple vector system to improve performance and utilisation of recombinant antibodies

    OpenAIRE

    Vincent Karen J; Mitchell Joanne N; Rojas Gertrudis; Martin Cecile D; Wu Jiahua; McCafferty John; Schofield Darren J

    2006-01-01

    Abstract Background Isolation of recombinant antibody fragments from antibody libraries is well established using technologies such as phage display. Phage display vectors are ideal for efficient display of antibody fragments on the surface of bacteriophage particles. However, they are often inefficient for expression of soluble antibody fragments, and sub-cloning of selected antibody populations into dedicated soluble antibody fragment expression vectors can enhance expression. Results We ha...

  15. Discovery and characterization of antibody variants using mass spectrometry-based comparative analysis for biosimilar candidates of monoclonal antibody drugs.

    Science.gov (United States)

    Li, Wenhua; Yang, Bin; Zhou, Dongmei; Xu, Jun; Ke, Zhi; Suen, Wen-Chen

    2016-07-01

    Liquid chromatography mass spectrometry (LC-MS) is the most commonly used technique for the characterization of antibody variants. MAb-X and mAb-Y are two approved IgG1 subtype monoclonal antibody drugs recombinantly produced in Chinese hamster ovary (CHO) cells. We report here that two unexpected and rare antibody variants have been discovered during cell culture process development of biosimilars for these two approved drugs through intact mass analysis. We then used comprehensive mass spectrometry-based comparative analysis including reduced light, heavy chains, and domain-specific mass as well as peptide mapping analysis to fully characterize the observed antibody variants. The "middle-up" mass comparative analysis demonstrated that the antibody variant from mAb-X biosimilar candidate was caused by mass variation of antibody crystalline fragment (Fc), whereas a different variant with mass variation in antibody antigen-binding fragment (Fab) from mAb-Y biosimilar candidate was identified. Endoproteinase Lys-C digested peptide mapping and tandem mass spectrometry analysis further revealed that a leucine to glutamine change in N-terminal 402 site of heavy chain was responsible for the generation of mAb-X antibody variant. Lys-C and trypsin coupled non-reduced and reduced peptide mapping comparative analysis showed that the formation of the light-heavy interchain trisulfide bond resulted in the mAb-Y antibody variant. These two cases confirmed that mass spectrometry-based comparative analysis plays a critical role for the characterization of monoclonal antibody variants, and biosimilar developers should start with a comprehensive structural assessment and comparative analysis to decrease the risk of the process development for biosimilars. PMID:27214604

  16. PK/PD analysis of a novel pH-dependent antigen-binding antibody using a dynamic antibody-antigen binding model.

    Science.gov (United States)

    Haraya, Kenta; Tachibana, Tatsuhiko; Iwayanagi, Yuki; Maeda, Atsuhiko; Ozeki, Kazuhisa; Nezu, Junichi; Ishigai, Masaki; Igawa, Tomoyuki

    2016-04-01

    Previously, we have reported novel engineered antibody with pH-dependent antigen-binding (recycling antibody), and with both pH-dependent antigen-binding and increased FcRn-binding at neutral pH (sweeping antibody). The purpose of this study is to perform PK/PD predictions to better understand the potential applications of the antibodies as therapeutics. To demonstrate the applicability of recycling and sweeping antibodies over conventional antibodies, PK/PD analyses were performed. PK/PD parameters for antibody and antigen dynamics were estimated from the results of a pharmacokinetic study in human FcRn transgenic mice. A simulation study was performed using the estimated PK/PD parameters with various target antigen profiles. In comparison to conventional antibody, recycling antibody enhanced antibody-antigen complex clearance by 3 folds, while sweeping antibody accelerated antigen clearance by 10 folds in a pharmacokinetic study. Simulation results showed that recycling and sweeping antibodies can improve dosage frequency and reduce the required dose for target antigens with various clearances, plasma concentrations or binding kinetics. Moreover, importance of the association rate constant to enhance the beneficial effect of antibodies was shown. These results support the conclusion that recycling and sweeping antibodies can be applied to various target antigens with different profiles, and expand the number of antigens that antibodies can target. PMID:26944099

  17. Tumor targeting of radiolabeled antibodies using HYNIC chelate

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Tae Sup; Chung, Wee Sup; Woo, Kwang Sun; Choi, Tae Hyun; Chung, Hye Kyung; Lee, Myung Jin; Kim, So Yeon; Jung, Jae Ho; Choi, Chang Woon; Lim, Sang Moo [KIRAMS, Seoul (Korea, Republic of); Darwati, Siti [National Nuclear Energy Agency, Tangerang (Indonesia)

    2004-07-01

    There is an increasing interest in the use of labeled antibodies for diagnosis of cancers as well as for therapy. Various radiolabeling methods have been used in order to obtain better tumor specific targeting for detection and therapy. It was generally used to tumor targeted immunotherapy and immunodetection that lym-1, mouse monoclonal antibody, was specific binding to surface antigen of Raji. The 3E8 antibody was produced from humanized anti-TAG-72 monoclonal antibody (AKA) by amino acid change in 95-99 residues of heavy chain complementary determinant regions (HCDRs) 3 using phage displayed library technology. In this study, we are investigating the usefulness of HYNIC chelate as a bifunctional chelating agent in radioimmunodetecton of tumor. Two types of antibodies, Lym-1 and 3E8, were used for the conjugation with HYNIC chelate. Lym-1 and 3E8 are specific antibodies to surface antigen of Non-Hogkin's lymphoma and TAG-72 antigen of colorectal carcinoma, respectively. We prepare HYNIC-antibody conjugates, determine radiolabeling yield with {sup 99m}Tc and evaluate tumor targeting in tumor bearing nude mice model.

  18. Antibody dependent enhancement of frog virus 3 infection

    Directory of Open Access Journals (Sweden)

    Penny Emily

    2010-02-01

    Full Text Available Abstract Background Viruses included in the family Iridoviridae are large, icosahedral, dsDNA viruses that are subdivided into 5 genera. Frog virus 3 (FV3 is the type species of the genus Ranavirus and the best studied iridovirus at the molecular level. Typically, antibodies directed against a virus act to neutralize the virus and limit infection. Antibody dependent enhancement occurs when viral antibodies enhance infectivity of the virus rather than neutralize it. Results Here we show that anti-FV3 serum present at the time of FV3 infection enhances infectivity of the virus in two non-immune teleost cell lines. We found that antibody dependent enhancement of FV3 was dependent on the Fc portion of anti-FV3 antibodies but not related to complement. Furthermore, the presence of anti-FV3 serum during an FV3 infection in a non-immune mammalian cell line resulted in neutralization of the virus. Our results suggest that a cell surface receptor specific to teleost cell lines is responsible for the enhancement. Conclusions This report represents the first evidence of antibody dependent enhancement in iridoviruses. The data suggests that anti-FV3 serum can either neutralize or enhance viral infection and that enhancement is related to a novel antibody dependent enhancement pathway found in teleosts that is Fc dependent.

  19. Quantifying serum antibody in bird fanciers' hypersensitivity pneumonitis

    Directory of Open Access Journals (Sweden)

    Anderson Kenneth

    2006-06-01

    Full Text Available Abstract Background Detecting serum antibody against inhaled antigens is an important diagnostic adjunct for hypersensitivity pneumonitis (HP. We sought to validate a quantitative fluorimetric assay testing serum from bird fanciers. Methods Antibody activity was assessed in bird fanciers and control subjects using various avian antigens and serological methods, and the titer was compared with symptoms of HP. Results IgG antibody against pigeon serum antigens, quantified by fluorimetry, provided a good discriminator of disease. Levels below 10 mg/L were insignificant, and increasing titers were associated with disease. The assay was unaffected by total IgG, autoantibodies and antibody to dietary hen's egg antigens. Antigens from pigeon serum seem sufficient to recognize immune sensitivity to most common pet avian species. Decreasing antibody titers confirmed antigen avoidance. Conclusion Increasing antibody titer reflected the likelihood of HP, and decreasing titers confirmed antigen avoidance. Quantifying antibody was rapid and the increased sensitivity will improve the rate of false-negative reporting and obviate the need for invasive diagnostic procedures. Automated fluorimetry provides a method for the international standardization of HP serology thereby improving quality control and improving its suitability as a diagnostic adjunct.

  20. A mechanistic compartmental model for total antibody uptake in tumors.

    Science.gov (United States)

    Thurber, Greg M; Dane Wittrup, K

    2012-12-01

    Antibodies are under development to treat a variety of cancers, such as lymphomas, colon, and breast cancer. A major limitation to greater efficacy for this class of drugs is poor distribution in vivo. Localization of antibodies occurs slowly, often in insufficient therapeutic amounts, and distributes heterogeneously throughout the tumor. While the microdistribution around individual vessels is important for many therapies, the total amount of antibody localized in the tumor is paramount for many applications such as imaging, determining the therapeutic index with antibody drug conjugates, and dosing in radioimmunotherapy. With imaging and pretargeted therapeutic strategies, the time course of uptake is critical in determining when to take an image or deliver a secondary reagent. We present here a simple mechanistic model of antibody uptake and retention that captures the major rates that determine the time course of antibody concentration within a tumor including dose, affinity, plasma clearance, target expression, internalization, permeability, and vascularization. Since many of the parameters are known or can be estimated in vitro, this model can approximate the time course of antibody concentration in tumors to aid in experimental design, data interpretation, and strategies to improve localization. PMID:22974563

  1. Phospho-Specific Antibody Probes of Intermediate Filament Proteins.

    Science.gov (United States)

    Goto, Hidemasa; Tanaka, Hiroki; Kasahara, Kousuke; Inagaki, Masaki

    2016-01-01

    Intermediate filaments (IFs) form one of the major cytoskeletal systems in the cytoplasm or beneath the nuclear membrane. Accumulating data have suggested that IF protein phosphorylation dramatically changes IF structure/dynamics in cells. For the production of an antibody recognizing site-specific protein phosphorylation (a site- and phosphorylation state-specific antibody), we first employed a strategy to immunize animals with an in vitro-phosphorylated polypeptide or a phosphopeptide (corresponding to a phosphorylated residue and its surrounding sequence of amino acids), instead of a phosphorylated protein. Our established methodology not only improves the chance of obtaining a phospho-specific antibody but also has the advantage that one can predesign a targeted phosphorylation site. It is now applied to the production of an antibody recognizing other types of site-specific posttranslational modification, such as acetylation or methylation. The use of such an antibody in immunocytochemistry enables us to analyze spatiotemporal distribution of site-specific IF protein phosphorylation. The antibody is of great use to identify a protein kinase responsible for in vivo IF protein phosphorylation and to monitor intracellular kinase activities through IF protein phosphorylation. Here, we present an overview of our methodology and describe stepwise approaches for the antibody characterization. We also provide some examples of analyses for IF protein phosphorylation involved in mitosis and signal transduction.

  2. Pathogenic and Epiphenomenal Anti-DNA Antibodies in SLE

    Directory of Open Access Journals (Sweden)

    Mirjana Pavlovic

    2010-01-01

    Full Text Available The discoveries of natural and the development of manufactured highly efficient catalytic antibodies (abzymes opens the door to many practical applications. One of the most fascinating is the use of such antibodies in human therapy and prevention (vaccination, of cancer, AIDS, autoimmune diseases. A special entity of naturally occurring DNA hydrolytic anti-DNA antibodies is emerging within past decades linked to autoimmune and lymphoproliferative disorders, such as systemic lupus erythematosus (SLE, multiple sclerosis (MS, Sjogren Syndrome (SS, B - Chronic lymphocytic leucosis (B-CLL, and Multiple Myeloma (MM. The origin of the antibodies is unknown. The underlying mechanisms of these activities are suggested to be penetration into the living cells and translocation in the nucleus, with recognition of the specific binding sites at particular (ss or ds DNA. There are controversies in the literature whether hydrolysis is a sequence-specific event. The interplay between anti-DNA antibodies and DNA is not yet elucidated. This molecular “twist” also suggests that anti-DNA antibodies with DNA hydrolytic capacity could be the organism's immune response to a microbial attack, with microbial DNA, or specific genes within microbial DNA sequence, as a target for neutralization. The catalytic antibody-based approach can become a key tool in selective chemotherapeutic strategies.

  3. Seroprevalence of dengue virus antibodies in healthy Jamaicans.

    Science.gov (United States)

    Brown, Michelle G; Vickers, Ivan E; Salas, Rose Alba; Smikle, Monica F

    2009-01-01

    Dengue fever, a mosquito borne viral infection, is endemic to Jamaica. The seroprevalence of dengue IgG and IgM antibodies were determined in 277 healthy Jamaicans by enzyme linked immunosorbent assay (ELISA). The seroprevalence of dengue IgG antibodies was 100% (277/277) while dengue IgM antibodies were found in 3.6% (10/277). A statistically significant association was found between the presence of dengue IgM antibodies and gender (males 10/105, 9.5% vs females 0/172, 0.0%); chi(2) = 17.0, p=0.000.The high seroprevalence rate of dengue IgG antibodies and the presence of dengue IgM in the healthy population are in keeping with the endemicity of the virus in Jamaica. Therefore tests for dengue IgG antibodies are of limited usefulness in Jamaica and can be safely excluded from diagnostic testing as a cost saving measure. Serological diagnosis of current dengue infection should be centred around the dengue IgM tests although the limitations in the predictive values of such tests should also be considered. The results also suggest that the risk of emergence of the more severe forms of dengue, dengue haemorrhagic fever (DHF) and dengue shock syndrome (DSS) in the Jamaican population, due to the presence of enhancing antibodies, is high. PMID:19996526

  4. Isolation of Potent CGRP Neutralizing Antibodies Using Four Simple Assays.

    Science.gov (United States)

    Neal, Frances; Arnold, Joanne; Rossant, Christine J; Podichetty, Sadhana; Lowne, David; Dobson, Claire; Wilkinson, Trevor; Colley, Caroline; Howes, Rob; Vaughan, Tristan J

    2016-01-01

    Calcitonin gene-related peptide (CGRP) is a small neuropeptide and a potent vasodilator that is widely associated with chronic pain and migraine. An antibody that inhibits CGRP function would be a potential therapeutic for treatment of these disorders. Here we describe the isolation of highly potent antibodies to CGRP from phage and ribosome display libraries and characterization of their epitope, species cross-reactivity, kinetics, and functional activity. Homogenous time-resolved fluorescence (HTRF) binding assays identified antibodies with the desired species cross-reactivity from naïve libraries, and HTRF epitope competition assays were used to characterize and group scFv by epitope. The functional inhibition of CGRP and species cross-reactivity of purified scFv and antibodies were subsequently confirmed using cAMP assays. We show that epitope competition assays could be used as a surrogate for functional cell-based assays during affinity maturation, in combination with scFv off-rate ranking by biolayer interferometry (BLI). This is the first time it has been shown that off-rate ranking can be predictive of functional activity for anti-CGRP antibodies. Here we demonstrate how, by using just four simple assays, diverse panels of antibodies to CGRP can be identified. These assay formats have potential utility in the identification of antibodies to other therapeutic targets. PMID:26450103

  5. Anti-Cardiolipin Antibody in Acute Myocardial Infarction

    Directory of Open Access Journals (Sweden)

    Abdolreza S. Jahromi

    2010-01-01

    Full Text Available Problem statement: Myocardial infarction is the combined result of environmental and personal factors. Data concerning the relation between anti-Phospholipid (aPL antibodies and myocardial infarction in subjects without evidence of overt autoimmune disease are conflicting. Anticardiolipin antibody is detected in various diseases like rheumatoid arthritis, systemic lupus erythematosus and anti-phospholipid antibody syndrome. The study of Anticardiolipin antibody in Acute Myocardial Infarction (AMI might shed light on etiologic mechanisms in the pathogenesis of acute coronary syndromes. The purpose of the present study was to determine association of plasma aPL antibodies, namely, anti-Cardiolipin (aCL antibodies, with AMI. Approach: This study recruited 45 patients with the diagnosis of AMI according to WHO criteria in their first 24 h of admission. Thirty six matched individuals were studied as the control group with normal coronary artery angiography. Samples were tested for IgG-class antibodies to cardiolipin by an ELISA and the results were compared. Results: There were not significant differences between plasma level of aCLAs IgG in the patients with AMI on admission ant the control group. Also aCLAs IgG was not correlated with hypertension, diabetes mellitus, hyperlipidemia, sex, age and smoking. Conclusion: Our findings suggest that aCLAs IgG are not indicative of hypercoagulable state in patients with AMI.

  6. Expression and assembly of a fully active antibody in algae

    Science.gov (United States)

    Mayfield, Stephen P.; Franklin, Scott E.; Lerner, Richard A.

    2003-01-01

    Although combinatorial antibody libraries have solved the problem of access to large immunological repertoires, efficient production of these complex molecules remains a problem. Here we demonstrate the efficient expression of a unique large single-chain (lsc) antibody in the chloroplast of the unicellular, green alga, Chlamydomonas reinhardtii. We achieved high levels of protein accumulation by synthesizing the lsc gene in chloroplast codon bias and by driving expression of the chimeric gene using either of two C. reinhardtii chloroplast promoters and 5' and 3' RNA elements. This lsc antibody, directed against glycoprotein D of the herpes simplex virus, is produced in a soluble form by the alga and assembles into higher order complexes in vivo. Aside from dimerization by disulfide bond formation, the antibody undergoes no detectable posttranslational modification. We further demonstrate that accumulation of the antibody can be modulated by the specific growth regime used to culture the alga, and by the choice of 5' and 3' elements used to drive expression of the antibody gene. These results demonstrate the utility of alga as an expression platform for recombinant proteins, and describe a new type of single chain antibody containing the entire heavy chain protein, including the Fc domain.

  7. Association of anti-phospholipid antibodies with connective tissue diseases

    Directory of Open Access Journals (Sweden)

    Reena Rai

    2015-01-01

    Full Text Available Background: The antiphospholipid antibodies (APLA are directed against phospholipids and their binding proteins and are frequently found in association with connective tissue disorders. Systemic lupus erythematoses (SLE with APLA may cause a diagnostic dilemma as there are several manifestations like haemolytic anemia, thrombocytopenia, neurologic manifestations, leg ulcerations, serositis proteinuria which overlap in both these conditions. We conducted a study to find out the association of antiphospholipid antibodies with connective tissue diseases and compared the clinical and laboratory parameters between antiphoshpolipid antibody positive and antiphoshpolipid antibody negative group. Materials and Methods: This study was carried out in 102 patients diagnosed with connective tissue diseases. APLA testing was done at baseline and for those positive, the test was repeated after 12 weeks. Results: 14.7 % of patients with connective tissue diseases tissue had positive antiphoshpolipid antibodies. Positive antiphoshpolipid antibody was detected in 73.3% of patients with SLE group, 13.3% of patients with mixed connective tissue disease (MCTD and 13.3% of patients with systemic sclerosis. APLA positivity was seen in SLE patients with leg ulcers (87.2%, neurologic manifestation (72.7%, hemolytic anemia (62.3%, thrombocytopenia (72.7%, serositis (27.8% and proteinuria(19.6%. Conclusions: Antiphoshpolipid antibodies should be tested in all patients with connective tissue disease.

  8. Phosphorylcholine allows for evasion of bactericidal antibody by Haemophilus influenzae.

    Directory of Open Access Journals (Sweden)

    Sarah E Clark

    Full Text Available The human pathogen Haemophilus influenzae has the ability to quickly adapt to different host environments through phase variation of multiple structures on its lipooligosaccharide (LPS, including phosphorylcholine (ChoP. During colonization with H. influenzae, there is a selection for ChoP+ phase variants. In a murine model of nasopharyngeal colonization, this selection is lost in the absence of adaptive immunity. Based on previous data highlighting the importance of natural antibody in limiting H. influenzae colonization, the effect of ChoP expression on antibody binding and its bactericidal activity was investigated. Flow cytometric analysis revealed that ChoP+ phase variants had decreased binding of antibody to LPS epitopes compared to ChoP- phase variants. This difference in antibody binding correlated with increased survival of ChoP+ phase variants in the presence of antibody-dependent, complement-mediated killing. ChoP+ phase variants were also more resistant to trypsin digestion, suggesting a general effect on the physical properties of the outer membrane. Moreover, ChoP-mediated protection against antibody binding correlated with increased resilience of outer membrane integrity. Collectively, these data suggest that ChoP expression provides a selective advantage during colonization through ChoP-mediated effects on the accessibility of bactericidal antibody to the cell surface.

  9. Serum Treponema IgM Antibody Test for Syphilis Diagnosis

    Institute of Scientific and Technical Information of China (English)

    郑占才; 张荣富; 溪茜

    2003-01-01

    Objective: To evaluate the clinical utility of testing serum anti-treponema pallidum IgM antibody in the diagnosis of syphilis patients. Methods: Seventy-two cases of syphilis were tested for specific IgM antibody with ELISA, and the results were compared with RPR and TPPA.Results: The sensitivity of IgM antibody was 73.3 %(11/15) in primary syphilis, 88.9% (16/18) in sec-ondary syphilis, and there was no significant differ-ence between these values (x2=1.6363, P>0.10). The sensitivity of IgM antibody in diagnosing latent syphi-lis was only 26.1% (6/23), much lower than the detec-tion rate in symptomatic earlv svDhilis (x2=17.6189. P<0.005). RPR and TPPA were both 100% sensitive in latent and early symptomatic syphilis. Two were posi,five for IgM in the 16 cases who had received regular treatments 2 to 24 months before enrolled.Conclusions: Specific IgM antibody detection doees not appear superior to RPR and TPPA in diagnosing primary syphilis. The diagnosis of latent syphilis should mainly rely on RPR and TPPA, since there are low titers of IgM antibody at that stage. IgM antibody testing alone should not be recommended for monitor-ing syphilis development or treatment efficacy. Fur-ther studies should be concerned.

  10. The use of combinations of monoclonal antibodies in clinical oncology.

    Science.gov (United States)

    Henricks, Linda M; Schellens, Jan H M; Huitema, Alwin D R; Beijnen, Jos H

    2015-12-01

    Treatment with monoclonal antibodies is becoming increasingly important in clinical oncology. These antibodies specifically inhibit signaling pathways in tumor growth and/or induce immunological responses against tumor cells. By combining monoclonal antibodies several pathways may be targeted simultaneously, potentially leading to additive or synergistic effects. Theoretically, antibodies are very suitable for use in combination therapy, because of limited overlapping toxicity and lack of pharmacokinetic interactions. In this article an overview is given of preclinical and clinical data on twenty-five different combinations of antibodies in oncology. Some of these combinations have proven clinical benefit, for example the combination of trastuzumab and pertuzumab in HER2-positive breast cancer, which exemplifies an additive or synergistic effect on antitumor activity in clinical studies and the combination of nivolumab and ipilimumab, which results in significant increases in progression-free and overall survival in patients with advanced melanoma. However, other combinations may lead to unfavorable results, such as bevacizumab with cetuximab or panitumumab in advanced colorectal cancer. These combinations result in shorter progression-free survival and increased toxicity compared to therapy with a single antibody. In summary, the different published studies showed widely varying results, depending on the combination of antibodies, indication and patient population. More preclinical and clinical studies are necessary to unravel the mechanisms behind synergistic or antagonistic effects of combining monoclonal antibodies. Most research on combination therapies is still in an early stage, but it is expected that for several tumor types the use of combination therapy of antibodies will become standard of care in the near future.

  11. Anti-microbial antibodies in celiac disease: Trick or treat?

    Institute of Scientific and Technical Information of China (English)

    Maria Papp; Ildiko Foldi; Istvan Altorjay; Eszter Palyu; Miklos Udvardy; Judit Tumpek; Sandor Sipka; Ilma Rita Korponay-Szabo; Eva Nemes; Gabor Veres; Tamas Dinya; Attila Tordai; Hajnalka Andrikovics; Gary L Norman; Peter Laszlo Lakatos

    2009-01-01

    AIM: To determine the prevalence of a new set of anti-glycan and anti-outer membrane protein (anti- OMP) antibodies in a Hungarian cohort of adult Celiac disease (CD) patients. METHODS: 190 consecutive CD patients [M/F: 71/119, age:39.9 (SD:14.1) years], 100 healthy, and 48 gastrointestinal controls were tested for glycan anti- Saccharomyces cerevisiae (gASCA), anti-laminaribioside (ALCA), anti-chitobioside, anti-mannobioside, anti-OMP antibodies and major NOD2/CARD15 mutations. Thirty out of 82 CD patients enrolled at the time of diagnosis were re-evaluated for the same antibodies after longstanding gluten-free diet (GFD).RESULTS: 65.9% of the CD patients were positive for at least one of the tested antibodies at the time of the diagnosis. Except anti-OMP and ALCA, antimicrobial antibodies were exclusively seen in untreated CD; however, the overall sensitivity was low. Any glycan positivity (LR+: 3.13; 95% CI: 2.08-4.73)was associated with an increased likelihood ratio for diagnosing CD. Significant correlation was found between the levels of anti-glycan and anti-endomysial or anti-transglutaminase antibodies. Anti-glycan positivity was lost after longstanding GFD. Anti-glycan antibody titers were associated with symptoms at presentation, but not the presence of NOD2/CARD15 mutations. Patients with severe malabsorption more frequently had multiple antibodies at diagnosis ( P = 0.019).CONCLUSION: The presence of anti-glycan antibodies in CD seems to be secondary to the impaired small bowel mucosa which can lead to increased antigen presentation.Furthermore, anti-glycan positivity may be considered an additional marker of CD and dietary adherence.

  12. Anti-DNA antibodies: Sequencing, cloning, and expression

    Energy Technology Data Exchange (ETDEWEB)

    Barry, M.M.

    1992-01-01

    To gain some insight into the mechanism of systemic lupus erythematosus, and the interactions involved in proteins binding to DNA four anti-DNA antibodies have been investigated. Two of the antibodies, Hed 10 and Jel 242, have previously been prepared from female NZB/NZW mice which develop an autoimmune disease resembling human SLE. The remaining two antibodies, Jel 72 and Jel 318, have previously been produced via immunization of C57BL/6 mice. The isotypes of the four antibodies investigated in this thesis were determined by an enzyme-linked-immunosorbent assay. All four antibodies contained [kappa] light chains and [gamma]2a heavy chains except Jel 318 which contains a [gamma]2b heavy chain. The complete variable regions of the heavy and light chains of these four antibodies were sequenced from their respective mRNAs. The gene segments and variable gene families expressed in each antibody were identified. Analysis of the genes used in the autoimmune anti-DNA antibodies and those produced by immunization indicated no obvious differences to account for their different origins. Examination of the amino acid residues present in the complementary-determining regions of these four antibodies indicates a preference for aromatic amino acids. Jel 72 and Jel 242 contain three arginine residues in the third complementary-determining region. A single-chain Fv and the variable region of the heavy chain of Hed 10 were expressed in Escherichia coli. Expression resulted in the production of a 26,000 M[sub r] protein and a 15,000 M[sub r] protein. An immunoblot indicated that the 26,000 M[sub r] protein was the Fv for Hed 10, while the 15,000 M[sub r] protein was shown to bind poly (dT). The contribution of the heavy chain to DNA binding was assessed.

  13. Human Dengue Antibodies against Structural and Nonstructural Proteins

    OpenAIRE

    Valdés, Katia; Alvarez, Mayling; Pupo, Maritza; Vázquez, Susana; Rodríguez, Rayner; Guzmán, María G.

    2000-01-01

    Antibodies against dengue virus type 2 and 4 proteins in acute-phase sera of 10 primary and 10 secondary dengue fever and dengue hemorrhagic fever patients were studied by Western blotting. In the first group the immune response was barely detectable, while in the second group more proteins were detected, with a very strong reaction. Anti-NS1 and -NS3 antibodies were detected mainly in secondary cases. Anti-E, -NS3, and -NS5 antibodies were detected in a high number of cases. The possibility ...

  14. Antibody-based protein detection using piezoresistive cantilever arrays

    Science.gov (United States)

    Dauksaite, Vita; Lorentzen, Martin; Besenbacher, Flemming; Kjems, Jørgen

    2007-03-01

    A piezoresistive cantilever array platform with electrical read-out was applied for protein detection using GST (glutathione-S-transferase) and GST antibodies as a model system. Sensing was performed in the static deflection mode under constant flow conditions. The GST antibodies were directly immobilized on the cantilever gold surface by means of free thiol groups. The setup allowed simultaneous deflection measurements with sensor and control-antibody-immobilized reference cantilevers and enabled detection of 1 ng µl-1 (40 nM) of GST protein, which is similar to the sensitivity reported for cantilever sensors using an optical read-out system.

  15. Measuring an antibody affinity distribution molecule by molecule

    Energy Technology Data Exchange (ETDEWEB)

    Bradbury, Andrew M [Los Alamos National Laboratory; Werner, James H [Los Alamos National Laboratory; Temirov, Jamshid [INVITROGEN

    2008-01-01

    Single molecule fluorescence mIcroscopy was used to observe the binding and unbinding of hapten decorated quantum dots with individual surface immobilized antibodies. The fluorescence time history from an individual antibody site can be used to calculate its binding affinity. While quantum dot blinking occurs during these measurements, we describe a simple empirical method to correct the apparent/observed affinity to account for the blinking contribution. The combination of many single molecule affinity measurements from different antibodies yields not only the average affinity, it directly measures the full shape and character of the surface affinity distribution function.

  16. Stimulation of antibody synthesis induced by surgical trauma in rats.

    Science.gov (United States)

    Kinnaert, P; Mahieu, A; Van Geertruyden, N

    1978-01-01

    The effect of a standard laparatomy on antibody synthesis was studied in Wistar R/A rats recieving an intravenous injection of 10(9) sheep red blood cells (SRBC) during the surgical procedure. Anti-SRBC antibody titres were significantly higher in operated animals than in controls. When SRBC were given 2 hr after the surgical procedure, stimulation of antibody synthesis still persisted, but when the antigen was administered 24 hr after laparotomy, no significant difference could be detected between the operated animals and controls. Surgery also enhances the secondary humoral response. PMID:668200

  17. Selective detection of antibodies in microstructured polymer optical fibers

    DEFF Research Database (Denmark)

    Jensen, Jesper Bo Damm; Hoiby, P.E.; Emiliyanov, Grigoriy Andreev;

    2005-01-01

    We demonstrate selective detection of fluorophore labeled antibodies from minute samples probed by a sensor layer of complementary biomolecules immobilized inside the air holes of microstructured Polymer Optical Fiber (mPOF). The fiber core is defined by a ring of 6 air holes and a simple procedure...... was applied to selectively capture either α-streptavidin or α-CRP antibodies inside these air holes. A sensitive and easy-to-use fluorescence method was used for the optical detection. Our results show that mPOF based biosensors can provide reliable and selective antibody detection in ultra small sample...

  18. Dosimetry of radiolabeled monoclonal antibodies used for therapy

    International Nuclear Information System (INIS)

    The present state of radiotherapy using labeled antibodies is reviewed. From the point of view of dosimetry, antibody therapy does not seem to have reached a stable and practicable enough state to provide an input to any but rather tentative dosimetry models. These, therefore, should not be taken too far until the problems of antibody targeting have been more fully developed. Some of the instrumental techniques for acquiring dosimetric data under clinical conditions are discussed as are some of the techniques of therapy in use today. 8 references, 3 figures

  19. Heterohybridoma for the production of non murine monoclonal antibodies

    Directory of Open Access Journals (Sweden)

    Kh.Victoria Chanu and M. Ayub Ali

    Full Text Available Hybridoma technology described by kohler and Milstein produce only mouse immunoglobulins. Such immunoglobulins have limited use due to its negative side effects such as the recipient’s immune response. The production of a non murine monoclonal antibody to combat the problems of murine monoclonal antibody is again difficult due to the lack of a suitable myeloma cell line. Heterohybridoma formed by the fusion of lymphocyte of one species with the myeloma cell of a different species is the solution, which can be used for the production of non murine monoclonal antibodies. [Veterinary World 2010; 3(8.000: 390-392

  20. Monoclonal antibody against a Burkitt lymphoma-associated antigen.

    OpenAIRE

    Wiels, J; Fellous, M.; Tursz, T

    1981-01-01

    A monoclonal antibody, referred to as 38.13, was obtained by fusing murine myeloma cells with Lewis rat splenocytes sensitized with Daudi cells (human Burkitt lymphoma containing Epstein--Barr virus genome but lacking HLA-A, -B, and -C and beta 2-microglobulin molecules at the cell surface). 38.13 antibody was demonstrated to be a rat IgM. By complement-dependent microcytotoxicity and indirect immunofluorescence assays, 38.13 antibody was shown to react specifically with cells derived from Bu...

  1. Characterization of antibodies against ferret immunoglobulins, cytokines and CD markers

    DEFF Research Database (Denmark)

    Martel, Cyril Jean-Marie; Aasted, Bent

    2009-01-01

    immunoglobulins, we identified and characterized polyclonal antibodies towards ferret IgG, IgM and IgA. We also identified 22 monoclonal antibodies (mAbs) raised mostly against human CD markers which cross-reacted with ferret leukocytes. These antibodies were originally specific against human CD8, CD9, CD14, CD18......, CD25, CD29, CD32, CD44, CD61, CD71, CD79b, CD88, CD104, CD172a and mink CD3. Finally, we identified 4 cross-reacting mAbs with specificities against ferret interferon-gamma, TNF-alpha, interleukin-4 and interleukin-8....

  2. Genetically delivered antibody protects against West Nile virus

    OpenAIRE

    Pereboev, Alexander; Borisevich, Viktoriya; Tsuladze, George; Shakhmatov, Mikhail; Hudman, Deborah; Kazachinskaia, Elena; Razumov, Ivan; Svyatchenko, Viktor; Loktev, Valery; Yamshchikov, Vladimir

    2007-01-01

    Gene-based delivery of recombinant antibody genes is a promising therapeutic strategy offering numerous advantages including sustained antibody levels, better safety profile and lower production cost. Here we describe generation of a recombinant antibody Fc-9E2 comprising a fusion protein between human Fc of IgG1 and a single-chain Fv derived from a hybridoma 9E2 secreting a mAb neutralizing West Nile virus (WNV). Fc-9E2 was shown to retain parental mAb's specificity and WNV-neutralizing capa...

  3. Feedback Enhancement of Antibody Responses via Complement and Fc Receptors

    OpenAIRE

    Dahlström, Jörgen

    2001-01-01

    IgG, IgM and IgE in complex with antigen have the capacity to regulate specific immune responses. In this investigation, the role of Fc receptors for IgG (FcγRI, FcγRII and FcγRIII) and complement receptors 1 and 2 (CR1/2) for antibody-mediated enhancement of antibody responses are investigated. IgM is known to efficiently activate complement and thereby enhance specific antibody responses but it is not known if this involves binding to CR1/2. Using CR1/2 deficient mice, immunized with sheep ...

  4. Immunity to rhabdoviruses in rainbow trout: the antibody response

    DEFF Research Database (Denmark)

    Lorenzen, Niels; Lapatra, S.E.

    1999-01-01

    occasional detrimental effect on rainbow trout farming. Research efforts have been focused on understanding the mechanisms involved in protective immunity. Several specific and nonspecific cellular and humoral parameters are believed to be involved, but only the antibody response has been characterised in......, have demonstrated that rainbow trout can produce specific and highly functional antibodies that are able to neutralise virus pathogenicity in vitro as well as in vivo. The apparently more restricted antibody response to IHNV and VHSV antigens in fish compared to mammals could possibly be explained by...

  5. Monoclonal antibodies and Fc fragments for treating solid tumors

    Directory of Open Access Journals (Sweden)

    Eisenbeis AM

    2012-01-01

    Full Text Available Andrea M Eisenbeis, Stefan J GrauDepartment of Neurosurgery, University Hospital of Cologne, Cologne, GermanyAbstract: Advances in biotechnology, better understanding of pathophysiological processes, as well as the identification of an increasing number of molecular markers have facilitated the use of monoclonal antibodies and Fc fragments in various fields in medicine. In this context, a rapidly growing number of these substances have also emerged in the field of oncology. This review will summarize the currently approved monoclonal antibodies used for the treatment of solid tumors with a focus on their clinical application, biological background, and currently ongoing trials.Keywords: targeted therapy, monoclonal antibodies, cancer, biological therapy

  6. Detection of antibodies to the 20s proteasome by ELISA

    DEFF Research Database (Denmark)

    Jørgensen, Karin Meinike; Frederiksen, Jette Lautrup; Nielsen, Christoffer Tandrup;

    2013-01-01

    The presence of antibodies against the 20S proteasome has been correlated with diseases like multiple sclerosis (MS) and systemic lupus erythematosus (SLE) but no definite association has been established. In order to investigate this further, we optimized an ELISA for proteasome antibodies...... and applied it to test a total of 324 serum and plasma samples from MS patients, SLE patients, and healthy controls. Our results yield a functional and reliable assay but no correlation between the amount of proteasome antibodies present and the development of MS or SLE could be established....

  7. Technological progresses in monoclonal antibody production systems.

    Science.gov (United States)

    Rodrigues, Maria Elisa; Costa, Ana Rita; Henriques, Mariana; Azeredo, Joana; Oliveira, Rosário

    2010-01-01

    Monoclonal antibodies (mAbs) have become vitally important to modern medicine and are currently one of the major biopharmaceutical products in development. However, the high clinical dose requirements of mAbs demand a greater biomanufacturing capacity, leading to the development of new technologies for their large-scale production, with mammalian cell culture dominating the scenario. Although some companies have tried to meet these demands by creating bioreactors of increased capacity, the optimization of cell culture productivity in normal bioreactors appears as a better strategy. This review describes the main technological progresses made with this intent, presenting the advantages and limitations of each production system, as well as suggestions for improvements. New and upgraded bioreactors have emerged both for adherent and suspension cell culture, with disposable reactors attracting increased interest in the last years. Furthermore, the strategies and technologies used to control culture parameters are in constant evolution, aiming at the on-line multiparameter monitoring and considering now parameters not seen as relevant for process optimization in the past. All progresses being made have as primary goal the development of highly productive and economic mAb manufacturing processes that will allow the rapid introduction of the product in the biopharmaceutical market at more accessible prices. PMID:20043321

  8. Antibody Production in Plants and Green Algae.

    Science.gov (United States)

    Yusibov, Vidadi; Kushnir, Natasha; Streatfield, Stephen J

    2016-04-29

    Monoclonal antibodies (mAbs) have a wide range of modern applications, including research, diagnostic, therapeutic, and industrial uses. Market demand for mAbs is high and continues to grow. Although mammalian systems, which currently dominate the biomanufacturing industry, produce effective and safe recombinant mAbs, they have a limited manufacturing capacity and high costs. Bacteria, yeast, and insect cell systems are highly scalable and cost effective but vary in their ability to produce appropriate posttranslationally modified mAbs. Plants and green algae are emerging as promising production platforms because of their time and cost efficiencies, scalability, lack of mammalian pathogens, and eukaryotic posttranslational protein modification machinery. So far, plant- and algae-derived mAbs have been produced predominantly as candidate therapeutics for infectious diseases and cancer. These candidates have been extensively evaluated in animal models, and some have shown efficacy in clinical trials. Here, we review ongoing efforts to advance the production of mAbs in plants and algae. PMID:26905655

  9. Monoclonal Antibodies for Systemic Lupus Erythematosus (SLE

    Directory of Open Access Journals (Sweden)

    Gabriella Moroni

    2010-01-01

    Full Text Available A number of monoclonal antibodies (mAb are now under investigation in clinical trials to assess their potential role in Systemic Lupus Erythematosus (SLE. The most frequently used mAb is rituximab, which is directed against CD20, a membrane protein expressed on B lymphocytes. Uncontrolled trials reported an improvement of SLE activity in non-renal patients and other studies even reported an improvement of severe lupus nephritis unresponsive to conventional treatments. However two randomized trials failed to show the superiority of rituximab over conventional treatment in non renal SLE and in lupus nephritis. Preliminary trials reported promising results with epratuzumab, a humanized mAb directed against CD22, and with belimumab, a human mAb that specifically recognizes and inhibits the biological activity of BLyS a cytokine of the tumornecrosis-factor (TNF ligand superfamily. Other clinical trials with mAb directed against TNF-alpha, interleukin-10 (Il-10, Il-6, CD154, CD40 ligand, IL-18 or complement component C5 are under way. At present, however, in spite of good results reported by some studies, no firm conclusion on the risk-benefit profile of these mAbs in patients with SLE can be drawn from the available studies.

  10. Antigliadin antibody in sporadic adult ataxia

    Directory of Open Access Journals (Sweden)

    Mahdi Aloosh

    2012-09-01

    Full Text Available Background: The most common neurologic manifestationof gluten sensitivity is ataxia, which accounts for up to 40%of idiopathic sporadic ataxia. Timing of diagnosis of glutenataxia is vital as it is one of the very few treatable causes ofsporadic ataxia and causes irreversible loss of Purkinje cells.Antigliadin antibody (AGA of the IgG type is the bestmarker for neurological manifestations of gluten sensitivity.This study was conducted to measure the prevalence ofgluten ataxia in a group of Iranian patients with idiopathicataxia.Methods: For 30 patients with idiopathic cerebellar ataxia, aquestionnaire about clinical and demographic data wascompleted. Serum AGA (IgA and IgG and antiendomysialantibody (AEA were assessed. Gluten ataxic patientsunderwent duodenal biopsy. Magnetic resonanceimaging was done for all patients to see if cerebellaratrophy is present.Results: Only 2 patients had a positive IgG AGA (6.7%who both had a positive AEA while none of themshowed changes of celiac disease in their duodenalbiopsies. Only presence of gastrointestinal symptomsand pursuit eye movement disorders were higher inpatients with gluten ataxia.Conclusion: Prevalence of gluten ataxia in Iranianpatients with idiopathic ataxia seems to be lower thanmost of other regions. This could be explained by smallsample size, differences in genetics and nutritionalhabits and also effect of serologic tests in clinical versusresearch setting. Further researches with larger samplesize are recommended.

  11. Missing Links in Antibody Assembly Control

    Directory of Open Access Journals (Sweden)

    Tiziana Anelli

    2013-01-01

    Full Text Available Fidelity of the humoral immune response requires that quiescent B lymphocytes display membrane bound immunoglobulin M (IgM on B lymphocytes surface as part of the B cell receptor, whose function is to recognize an antigen. At the same time B lymphocytes should not secrete IgM until recognition of the antigen has occurred. The heavy chains of the secretory IgM have a C-terminal tail with a cysteine instead of a membrane anchor, which serves to covalently link the IgM subunits by disulfide bonds to form “pentamers” or “hexamers.” By virtue of the same cysteine, unassembled secretory IgM subunits are recognized and retained (via mixed disulfide bonds by members of the protein disulfide isomerase family, in particular ERp44. This so-called “thiol-mediated retention” bars assembly intermediates from prematurely leaving the cell and thereby exerts quality control on the humoral immune response. In this essay we discuss recent findings on how ERp44 governs such assembly control in a pH-dependent manner, shuttling between the cisGolgi and endoplasmic reticulum, and finally on how pERp1/MZB1, possibly as a co-chaperone of GRP94, may help to overrule the thiol-mediated retention in the activated B cell to give way to antibody secretion.

  12. Occult choriocarcinoma: Detection using radiolabeled monoclonal antibodies

    International Nuclear Information System (INIS)

    Occult choriocarcinoma, manifested only by an elevated B-hCG level, can be a difficult management problem. The authors evaluated the ability of I-131-labeled 5F9.3, a murine monoclonal antibody reactive with choriocarcinomas but not hCG, to detect foci of choriocarcinoma in five patients referred with elevated B-hCG levels but in whom the location of residual disease was uncertain. I-131 5F9.3, 0.5-1.0 mCi, was injected intravenously in each patient and images with dynamic background subtraction of TcHSA were obtained at later time points. In four patients chest studies were true positive (confirmed surgically in all), the chest CT scans in these patients had been interpreted as not definitely showing active disease. In the fifth patient no abnormal focus of uptake was seen and subsequent B-hCG levels normalized. In two of the patients with chest lesions, foci of abdominal uptake were seen that were not due to tumor. One of these patients had a partial small bowel obstruction; the other appeared to have a false-positive study. I-131 5F9.3 is a promising agent for the detection of occult choriocarcinomas

  13. Antibody conjugated graphene nanocomposites for pathogen detection

    Science.gov (United States)

    Sign, Chandan; Sumana, Gajjala

    2016-04-01

    Graphene oxide (GO), due to its excellent electrochemical properties and large surface area, known to be highly suitable material for biosensing application. Here, we report in situ synthesis of silver nanopaticles (AgNPs) onto the GO sheets for the electrochemical detection of Salmonella typhimurium (S.typhimurium). The GO-AgNPs composites have been deposited onto the indium tin oxide (ITO) coated glass substrate by the electrophoretic deposition technique. Carbodiimide coupling (EDC-NHS) has been used for the immobilization of antibodies of Salmonella typhimurium (anti-S.typhimurium) for detection of S.typhimurium. The electron microscopy and UV-visible studies reveal successful synthesis GO-AgNPs composites while FT-IR studies suggest the proper immobilization of anti-S.typhi. The cyclic voltammetry (CV) has been utilized for detection using anti-S.typhi/GOAgNPs/ITO based immunoelectrode as a function of S.typhimurium concentration. The fabricated immunosensor shows improved sensitivity of 33.04 μACFU-1mLcm-2 in a wide detection range of 101 to 106 CFUmL-1. This immunosensor may be utilized for the detection of other food borne pathogens like aflatoxin and E.coli also.

  14. Preparation and Characterization of Olaquindox Polyclonal Antibody

    Institute of Scientific and Technical Information of China (English)

    CHE,Huilian; SHI Baoxia; CHEN,Jinlan; ZHU,Xuejia; HE,Jiguo

    2009-01-01

    2-[N-(2-Hydroxyethyl)-carbamoyl]-3-methyl-oxoquinoxaline-l,4-oxides (OLA) was first mono-esterificated by succinic acid anhydride, and was then turned into a derivative with carboxyl. The synthetic product was purified by recrystallization with a yield of 52.47%. Its chemical structure was determined by NMR, IR and MS spectra. The determined structure was used to synthese OLA hapten, which had the molecular structural characteristics of OLA. The OLA hapten and bovine serum albumin (BSA) were conjugated using the activated ester method, while the hapten and ovalbumin (OVA) were coupled by the mixed anhydride method. The UV scanning and infrared spectrum results showed that the hapten and the carrier protein were successfully coupled to BSA and OVA, with the combined ratios of 3.8 : 1 and 5 : 1, respectively. OLA-BSA was used as the immunogen to immunize four New Zealand white rabbits, and the high titer anti-serum produced relatively. The titers were 1 " 6400, 1 : 1600, 1 : 12800 and 1 : 6400, respectively, determined by indirect ELISA OLA-OVA as the coating antigen. The intermediate inhibition concentration (IC50) of OLA in the indirect ELISA test was 743.3 ng/mL. The lowest detection limit (IC20) was 5.71 ng/mL. High-purity OLA polyclonal antibody has been obtained by saturated ammonium sulfate and ion-exchange chromatography.

  15. Silanization and antibody immobilization on SU-8

    Science.gov (United States)

    Joshi, Manoj; Pinto, Richard; Rao, V. Ramgopal; Mukherji, Soumyo

    2007-01-01

    SU-8, an epoxy based negative photoresist, has emerged as a structural material for microfabricated sensors due to its attractive mechanical properties like low Young's modulus and chemical properties like inertness to various chemicals used in microfabrication. It can be used to fabricate MEMS structures of high aspect ratio. However, the use of SU-8 in BioMEMS application has been limited by the fact that immobilization of biomolecules on SU-8 surfaces has not been reported. In this study, the epoxy groups on the SU-8 surface were hydrolyzed in the presence of sulphochromic solution. Following this, the surface was treated with [3-(2-aminoethyl) aminopropyl]-trimethoxysilane (AEAPS). The silanized SU-8 surface was used to incubate human immunoglobulin (HIgG). The immobilization of HIgG was proved by allowing FITC tagged goat anti-human IgG to react with HIgG. This process of antibody immobilization was used to immobilize HIgG on microfabricated SU-8 cantilevers.

  16. Modelling Virus and Antibody Dynamics during Dengue Virus Infection Suggests a Role for Antibody in Virus Clearance.

    Directory of Open Access Journals (Sweden)

    Hannah E Clapham

    2016-05-01

    Full Text Available Dengue is an infection of increasing global importance, yet uncertainty remains regarding critical aspects of its virology, immunology and epidemiology. One unanswered question is how infection is controlled and cleared during a dengue infection. Antibody is thought to play a role, but little past work has examined the kinetics of both virus and antibody during natural infections. We present data on multiple virus and antibody titres measurements recorded sequentially during infection from 53 Vietnamese dengue patients. We fit mechanistic mathematical models of the dynamics of viral replication and the host immune response to these data. These models fit the data well. The model with antibody removing virus fits the data best, but with a role suggested for ADCC or other infected cell clearance mechanisms. Our analysis therefore shows that the observed viral and antibody kinetics are consistent with antibody playing a key role in controlling viral replication. This work gives quantitative insight into the relationship between antibody levels and the efficiency of viral clearance. It will inform the future development of mechanistic models of how vaccines and antivirals might modify the course of natural dengue infection.

  17. SINGLE CHAIN VARIABLE FRAGMENTS OF ANTIBODIES AGAINST DIPHTHERIA TOXIN B-SUBUNIT ISOLATED FROM PHAGE DISPLAY HUMAN ANTIBODY LIBRARY

    Directory of Open Access Journals (Sweden)

    Oliinyk O. S.

    2014-02-01

    Full Text Available Diphtheria toxin is an exoantigen of Corynebacterium diphtheriae that inhibits protein synthesis and kills sensitive cells. The aim of this study was to obtain human recombinant single-chain variable fragment (scFv antibodies against receptor-binding B subunit of diphtheria toxin. 12 specific clones were selected after three rounds of a phage display naїve (unimmunized human antibody library against recombinant B-subunit. scFv DNA inserts from these 12 clones were digested with MvaI, and 6 unique restriction patterns were found. Single-chain antibodies were expressed in Escherichia coli XL1-blue. The recombinant proteins were characterized by immunoblotting of bacterial extracts and detection with an anti-E-tag antibody. The toxin B-subunit-binding function of the single-chain antibody was shown by ELISA. The affinity constants for different clones were found to be from 106 to 108 М–1. Due to the fact, that these antibody fragments recognized epitopes in the receptor-binding Bsubunit of diphtheria toxin, further studies are interesting to evaluate their toxin neutralization properties and potential for therapeutic applications. Obtained scFv-antibodies can also be used for detection and investigation of biological properties of diphtheria toxin.

  18. Development and evaluation of an enzyme-labeled antibody test for the rapid detection of hog cholera antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Saunders, G.C.

    1977-01-01

    A rapid enzyme-labeled antibody (ELA) microtechnique for the screening of swine for hog cholera antibodies was developed and evaluated with a blind study, using a 640-sample hog cholera serum bank. The total time to run a group of 22 samples was approximately 1 hour. The ELA test results correlated >99% with hog cholera serum-neutralization test results on the same serums. Test results also indicated that the ELA test shares with the hog cholera serum-neutralization test the problem of cross reactions between the antibodies of hog cholera and bovine viral diarrhea.

  19. Optimization of AFP-radioimmunoassay using Antibody Capture Technique

    International Nuclear Information System (INIS)

    Alpha-fetoprotein (AFP) is a substance produced by the unborn baby. When the neural tube is not properly formed large amounts of AFP pass into the amniotic fluid and reach the mother's blood. By measuring AFP in the mother's blood and amniotic fluid, it is possible to tell whether or not there is a chance that the unborn baby has a neural tube defect. AFP also used as a tumor marker for hepatocellular carcinoma. There are many different techniques for measuring AFP in blood, but the most accurate one is the immunoassay technique. The immunoassays can be classified on the basis of methodology into three classes; (1) the antibody capture assays, (2) the antigen capture assay, (3)the two-antibody sandwich assays. In this present study, the antibody capture assay in which the antigen is attached to a solid support, and labeled antibody is allowed to bind, will be optimized

  20. Detection of serum antitrichomonal antibodies in urogenital trichomoniasis by immunofluorescence.

    Directory of Open Access Journals (Sweden)

    Bhatt R

    1992-04-01

    Full Text Available Trichomonas vaginalis is a frequently encountered genital pathogen in both males and females. In females, vaginitis due to this parasite is one of the most common manifestation. The indirect fluorescent technique (IFA test was carried out to detect antitrichomonal antibodies in 370 female patients using whole cell antigen. Seventy one (19.18% gave positive reaction for either of the class IgG, IgM and IgA antibodies. The level of the IgG class antibodies was found to be higher i.e. 58 (81.69% than IgM 11 (15.27% antibodies, which may be suggestive of past infection or a prolonged manifestation by the organisms.

  1. Generation and characterization of monoclonal antibodies specific to Coenzyme A

    Directory of Open Access Journals (Sweden)

    Malanchuk O. M.

    2015-06-01

    Full Text Available Aim. Generation of monoclonal antibodies specific to Coenzyme A. Methods. Hybridoma technique. KLH carrier protein conjugated with CoA was used for immunization. Screening of positive clones was performed with BSA conjugated to CoA. Results. Monoclonal antibody that specifically recognizes CoA and CoA derivatives, but not its precursors ATP and cysteine has been generated. Conclusion. In this study, we describe for the first time the production and characterization of monoclonal antibodies against CoA. The monoclonal antibody 1F10 was shown to recognize specifically CoA in Western blotting, ELISA and immunoprecipitation. These properties make this antiboby a particularly valuable reagent for elucidating CoA function in health and disease.

  2. [Nephrotoxicity of anti-CD3 monoclonal antibodies].

    Science.gov (United States)

    Alegre, M L; Depierreux, M; Florquin, S; Abramowicz, D; De Pauw, L; Kinnaert, P; Vereerstraeten, P; Goldman, M

    1991-01-01

    The OKT3 monoclonal antibody entails a transient acute nephrotoxicity when used either to prevent or to treat renal allograft rejection. This nephrotoxicity was reproduced experimentally in mice injected with 145-2C11, an anti-CD3 monoclonal antibody which shares many properties with OKT3. Both clinical and experimental data suggest that the renal lesions are due to the systemic release of cytokines that occurs prior to the immunosuppression. Pre-treatment with corticosteroids before the injection of anti-CD3 monoclonal antibody mitigates both the release of cytokines and the nephrotoxicity, in a dose-dependent manner. Experimental data suggest that very high doses of methylprednisolone (50 mg/kg) administered 2 to 3 hours before the monoclonal antibody are necessary to obtain an optimal protection. PMID:1827878

  3. Hypotheses of the origin of natural antibodies: a glycobiologist's opinion.

    Science.gov (United States)

    Khasbiullina, N R; Bovin, N V

    2015-07-01

    It is generally accepted that the generation of antibodies proceeds due to immunization of an organism by alien antigens, and the level and affinity of antibodies are directly correlated to the presence of immunogen. At the same time, vast experimental material has been obtained providing evidence of antibodies whose level remains unchanged and affinity is constant during a lifetime. In contrast to the first, adaptive immunoglobulins, the latter are named natural antibodies (nAbs). The nAbs are produced by B1 cells, whereas adaptive Abs are produced by B2. This review summarizes general data on nAbs and presents in more detail data on antigens of carbohydrate origin. Hypotheses on the origin of nAbs and their activation mechanisms are discussed. We present our thoughts on this matter supported by our experimental data on nAbs to glycans.

  4. The Clinical Proteomic Technologies for Cancer | Antibody Portal

    Science.gov (United States)

    An objective of the Reagents and Resources component of NCI's Clinical Proteomic Technologies for Cancer Initiative is to generate highly characterized monoclonal antibodies to human proteins associated with cancer.

  5. The Clinical Proteomic Technologies for Cancer | Antibody Scientific Committee

    Science.gov (United States)

    An objective of the Reagents and Resources component of NCI's Clinical Proteomic Technologies for Cancer Initiative is to generate highly characterized monoclonal antibodies to human proteins associated with cancer.

  6. Role of immunoglobulin G antibodies in diagnosis of food allergy.

    Science.gov (United States)

    Gocki, Jacek; Bartuzi, Zbigniew

    2016-08-01

    This paper presents current views on the role of immunoglobulin G (IgG) antibodies in the reactions with food antigens in the digestive tract and their role in the diagnosis of food allergy based on the assays of specific IgG class antibodies, with a special focus on contemporary practice guidelines. In the light of current scientific knowledge, the IgG-specific antibody-mediated reactions are a body's natural and normal defensive reactions to infiltrating food antigens, which are considered as pathogens. On the other hand, specific IgG antibodies against food allergens play a crucial role in the induction and maintaining of immunological tolerance to food antigens. The statements of many scientific societies stress that sIgG are of no significant importance in the diagnosis of food allergy since their presence is associated with a normal immune response to food allergens and attests to a protracted exposure to food antigens. PMID:27605894

  7. Radioimmunoimaging of experimental gliomas using radiolabelled monoclonal antibodies

    International Nuclear Information System (INIS)

    The biodistribution and tumour uptake of radiolabelled (131 I) glioma-seeking monoclonal antibodies (14 AC1) and their F(ab')2 fragments were investigated in nude mice having received glioma transplants. Radioimmunoimaging by external scintigraphy at 48 and 96 hours pointed to a superior tumour localisation by the fragments that was clearly related to the dose. Wholebody determinations of the biokinetic behaviour led to the following results: Faster clearance anc more ready elimination from the blood pool for the fragments, preferential uptake in the tumour; intact antibodies; binding in the liver, spleen and lungs. The study confirmed the value of fragments of monoclonal antibodies in the diagnosis of tumours and pointed to the possibility of using intact monoclonal antibodies as carriers of radioisotopes and cytotoxic drugs within the scope of therapeutic programmes. (TRV)

  8. Recent advances in the construction of antibody-drug conjugates

    Science.gov (United States)

    Chudasama, Vijay; Maruani, Antoine; Caddick, Stephen

    2016-02-01

    Antibody-drug conjugates (ADCs) comprise antibodies covalently attached to highly potent drugs using a variety of conjugation technologies. As therapeutics, they combine the exquisite specificity of antibodies, enabling discrimination between healthy and diseased tissue, with the cell-killing ability of cytotoxic drugs. This powerful and exciting class of targeted therapy has shown considerable promise in the treatment of various cancers with two US Food and Drug Administration approved ADCs currently on the market (Adcetris and Kadcyla) and approximately 40 currently undergoing clinical evaluation. However, most of these ADCs exist as heterogeneous mixtures, which can result in a narrow therapeutic window and have major pharmacokinetic implications. In order for ADCs to deliver their full potential, sophisticated site-specific conjugation technologies to connect the drug to the antibody are vital. This Perspective discusses the strategies currently used for the site-specific construction of ADCs and appraises their merits and disadvantages.

  9. Antibody structural modeling with prediction of immunoglobulin structure (PIGS)

    KAUST Repository

    Marcatili, Paolo

    2014-11-06

    © 2014 Nature America, Inc. All rights reserved. Antibodies (or immunoglobulins) are crucial for defending organisms from pathogens, but they are also key players in many medical, diagnostic and biotechnological applications. The ability to predict their structure and the specific residues involved in antigen recognition has several useful applications in all of these areas. Over the years, we have developed or collaborated in developing a strategy that enables researchers to predict the 3D structure of antibodies with a very satisfactory accuracy. The strategy is completely automated and extremely fast, requiring only a few minutes (~10 min on average) to build a structural model of an antibody. It is based on the concept of canonical structures of antibody loops and on our understanding of the way light and heavy chains pack together.

  10. Monoclonal antibodies against the human leukemia cell line K 562.

    Science.gov (United States)

    Böttger, V; Hering, S; Jantscheff, P; Micheel, B

    1985-01-01

    Three monoclonal antibodies raised against K 562, a cell line originally established from a patient with chronic myeloid leukemia (CML) in terminal blast crisis, were selected according to their distinct reaction pattern. Whereas two antibodies (ZIK-C1-A/C5 and ZIK-C1-A/H5 also designated C and H) recognized antigens, present on K 562 cells and other immature and mature hematopoietic cells (cell lines and normal blood and bone marrow cells), antibody ZIK-C1-A/D9 also designated Y showed an exclusive binding to K 562 cells. The results obtained (here and in the following paper) indicate, that antibody ZIK-C1-A/D9 defines an early differentiation antigen of hematopoiesis or a leukemia-associated antigen.

  11. Digital gangrene associated with anticentromere antibodies: a case report

    Directory of Open Access Journals (Sweden)

    Nair Bindu

    2010-06-01

    Full Text Available Abstract Introduction Anticentromere antibodies have been associated with peripheral vascular occlusive disease, most frequently accompanied by sclerodactyly in the context of a connective tissue disorder. We report a case of digital gangrene with no other clinical associations except positive anticentromere antibodies. Case presentation Our patient, a 53-year-old Caucasian woman, non-smoker, presented with progressive pain and blackening of the distal right third finger over the preceding five weeks. No sclerodactyly was evident. She was anticentromere antibody positive at greater than 100 U/mL. Angiography revealed diffuse distal vasculopathy in both upper extremities. Other investigations were unremarkable. Conclusions It is rare for anticentromere antibody-associated digital necrosis to develop without concomitant sclerodactyly. However, this patient's case illustrates the need to consider an autoimmune contribution to the pathogenesis of digital ischemia even in the absence of a recognizable connective tissue disease.

  12. [Continuous perfusion culture hybridoma cells for production of monoclonal antibody].

    Science.gov (United States)

    Mi, Li; Li, Ling; Feng, Qiang; Yu, Xiao-Ling; Chen, Zhi-Nan

    2002-05-01

    Hybridoma cells were cultured by continuous perfusion in Fibra-Cel of 5L packed-bed bioreactor for 22 days in low serum or serum-free media. The corresponded amino acids were fed and serum concentration was decreased by analyzing glucose concentration, oxygen uptake rate, secretary antibody amount and amino acids concentration in culture supernatant. Comparing with continuous perfusion culture that amino acids were not fed, antibody amount of production was increased about 2-3 times. The inoculated cell density was 2.5 x 10(5) cells/mL, while the final cell density was 8.79 x 10(8) cells/mL. Antibody production was reached 295 mg/L/d at average level, and the highest level was reached 532 mg/L/d. These results provided a primary mode of enlarge culture for monoclonal antibody industralization. PMID:12192875

  13. Prevalence of antileptospiral serum antibodies in dogs in Ireland

    Science.gov (United States)

    A total of 474 serum samples from client owned Irish dogs were tested for the presence of antibodies against serovars Canicola, Icterohaemorrhagiae, Bratislava, Autumnalis, Pomona, Altodouro, Grippotyphosa, Mozdok, Hardjobovis and Ballum. Six percent of dogs presented to veterinary practitioners for...

  14. The difference of ELISA and LABScreen in detecting HLA antibodies

    Institute of Scientific and Technical Information of China (English)

    吴萍萍

    2013-01-01

    Objective To compare the difference of ELISA and LABScreen in detecting HLA antibodies and evaluate their effects on allograft rejection.Methods Consecutive patients undergoing kidney transplantion from November,2008 to December,2009 in the First Affiliated Hospital

  15. Taking up Cancer Immunotherapy Challenges: Bispecific Antibodies, the Path Forward?

    Directory of Open Access Journals (Sweden)

    Joanie Del Bano

    2015-12-01

    Full Text Available As evidenced by the recent approvals of Removab (EU, Trion Pharma in 2009 and of Blincyto (US, Amgen in 2014, the high potential of bispecific antibodies in the field of immuno-oncology is eliciting a renewed interest from pharmaceutical companies. Supported by rapid advances in antibody engineering and the development of several technological platforms such as Triomab or bispecific T cell engagers (BiTEs, the “bispecifics” market has increased significantly over the past decade and may occupy a pivotal space in the future. Over 30 bispecific molecules are currently in different stages of clinical trials and more than 70 in preclinical phase. This review focuses on the clinical potential of bispecific antibodies as immune effector cell engagers in the onco-immunotherapy field. We summarize current strategies targeting various immune cells and their clinical interests. Furthermore, perspectives of bispecific antibodies in future clinical developments are addressed.

  16. Anti-collagen antibodies in sera from rheumatoid arthritis patients.

    Science.gov (United States)

    Beard, H K; Ryvar, R; Skingle, J; Greenbury, C L

    1980-11-01

    Anti-cartilage antibodies, demonstrable by immunofluorescence, were found in 3.3% of rheumatoid arthritis patients. In most of these patients antibodies to type II collagen were detected. In specificity studies on these anti-collagen antibodies, they appeared to be type specific, showing no reaction with collagen types I and III. Denatured type II collagen reacted much less well than native type II, but isolated peptides from different regions of the collagen molecule were differentiated by individual sera. Removal of the glycoside side chains from native type II collagen had no effect on its antigenicity. The findings suggest that these patients produce highly specific antibodies which react with the triple helix of type II collagen.

  17. Serum Vaccine Antibody Concentrations in Children Exposed to Perfluorinated Compounds

    DEFF Research Database (Denmark)

    Grandjean, P.; Andersen, E. W.; Budtz-Jorgensen, E.;

    2012-01-01

    whether PFC exposure is associated with antibody response to childhood vaccinations. Design, Setting, and Participants Prospective study of a birth cohort from the National Hospital in the Faroe Islands. A total of 656 consecutive singleton births were recruited during 1999-2001, and 587 participated...... in follow-up through 2008. Main Outcome Measures Serum antibody concentrations against tetanus and diphtheria toxoids at ages 5 and 7 years. Results Similar to results of prior studies in the United States, the PFCs with the highest serum concentrations were perfluorooctane sulfonic acid (PFOS......) and perfluorooctanoic acid (PFOA). Among PFCs in maternal pregnancy serum, PFOS showed the strongest negative correlations with antibody concentrations at age 5 years, for which a 2-fold greater concentration of exposure was associated with a difference of -39% (95% CI, -55% to -17%) in the diphtheria antibody...

  18. A Cayley Tree Immune Network Model with Antibody Dynamics

    CERN Document Server

    Anderson, R W; Perelson, A S; Anderson, Russell W.; Neumann, Avidan U.; Perelson, Alan S.

    1993-01-01

    Abstract: A Cayley tree model of idiotypic networks that includes both B cell and antibody dynamics is formulated and analyzed. As in models with B cells only, localized states exist in the network with limited numbers of activated clones surrounded by virgin or near-virgin clones. The existence and stability of these localized network states are explored as a function of model parameters. As in previous models that have included antibody, the stability of immune and tolerant localized states are shown to depend on the ratio of antibody to B cell lifetimes as well as the rate of antibody complex removal. As model parameters are varied, localized steady-states can break down via two routes: dynamically, into chaotic attractors, or structurally into percolation attractors. For a given set of parameters, percolation and chaotic attractors can coexist with localized attractors, and thus there do not exist clear cut boundaries in parameter space that separate regions of localized attractors from regions of percola...

  19. Engineered Antibodies for Monitoring of Polynuclear Aromatic Hydrocarbons

    Energy Technology Data Exchange (ETDEWEB)

    Alexander E. Karu Ph.D; Victoria A. Roberts Ph.D.; Qing X. Li, Ph.D.

    2002-01-17

    This project was undertaken to fill needs in ODE's human and ecosystem health effects research, site remediation, rapid emergency response, and regulatory compliance monitoring programs. Doe has greatly stimulated development and validation of antibody-based, rapid, field-portable detection systems for small hazardous compounds. These range from simple dipsticks, microplate enzyme-linked immunosorbent assays (ELISAs), and hand-held colorimeters, to ultrasensitive microfluidic reactors, fiber-optic sensors and microarrays that can identify multiple analytes from patterns of cross-reactivity. Unfortunately, the technology to produce antibodies with the most desirable properties did not keep pace. Lack of antibodies remains a limiting factor in production and practical use of such devices. The goals of our project were to determine the chemical and structural bases for the antibody-analyte binding interactions using advanced computational chemistry, and to use this information to create useful new binding properties through in vitro genetic engineering and combinatorial library methods.

  20. Role of immunoglobulin G antibodies in diagnosis of food allergy

    Science.gov (United States)

    Bartuzi, Zbigniew

    2016-01-01

    This paper presents current views on the role of immunoglobulin G (IgG) antibodies in the reactions with food antigens in the digestive tract and their role in the diagnosis of food allergy based on the assays of specific IgG class antibodies, with a special focus on contemporary practice guidelines. In the light of current scientific knowledge, the IgG-specific antibody-mediated reactions are a body's natural and normal defensive reactions to infiltrating food antigens, which are considered as pathogens. On the other hand, specific IgG antibodies against food allergens play a crucial role in the induction and maintaining of immunological tolerance to food antigens. The statements of many scientific societies stress that sIgG are of no significant importance in the diagnosis of food allergy since their presence is associated with a normal immune response to food allergens and attests to a protracted exposure to food antigens. PMID:27605894

  1. Anti-B cell antibody therapies for inflammatory rheumatic diseases

    DEFF Research Database (Denmark)

    Faurschou, Mikkel; Jayne, David R W

    2014-01-01

    Several monoclonal antibodies targeting B cells have been tested as therapeutics for inflammatory rheumatic diseases. We review important observations from randomized clinical trials regarding the efficacy and safety of anti-B cell antibody-based therapies for rheumatoid arthritis, systemic lupus...... erythematosus, antineutrophil cytoplasmic antibody-associated vasculitis, polymyositis/dermatomyositis, and primary Sjögren's syndrome. For some anti-B cell agents, clinical benefits have been convincingly demonstrated, while other B cell-targeted therapies failed to improve outcomes when added to standard...... and functions in rheumatic disorders. Future studies should also evaluate how to maintain disease control by means of conventional and/or biologic immunosuppressants after remission-induction with anti-B cell antibodies....

  2. Characterization of single chain antibody targets through yeast two hybrid

    Directory of Open Access Journals (Sweden)

    Vielemeyer Ole

    2010-08-01

    Full Text Available Abstract Background Due to their unique ability to bind their targets with high fidelity, antibodies are used widely not only in biomedical research, but also in many clinical applications. Recombinant antibodies, including single chain variable fragments (scFv, are gaining momentum because they allow powerful in vitro selection and manipulation without loss of function. Regardless of the ultimate application or type of antibody used, precise understanding of the interaction between the antibody's binding site and its specific target epitope(s is of great importance. However, such data is frequently difficult to obtain. Results We describe an approach that allows detailed characterization of a given antibody's target(s using the yeast two-hybrid system. Several recombinant scFv were used as bait and screened against highly complex cDNA libraries. Systematic sequencing of all retained clones and statistical analysis allowed efficient ranking of the prey fragments. Multiple alignment of the obtained cDNA fragments provided a selected interacting domain (SID, efficiently narrowing the epitope-containing region. Interactions between antibodies and their respective targets were characterized for several scFv. For AA2 and ROF7, two conformation-specific sensors that exclusively bind the activated forms of the small GTPases Rab6 and Rab1 respectively, only fragments expressing the entire target protein's core region were retained. This strongly suggested interaction with a non-linear epitope. For two other scFv, TA10 and SF9, which recognize the large proteins giantin and non-muscle myosin IIA, respectively, precise antibody-binding regions within the target were defined. Finally, for some antibodies, secondary targets within and across species could be revealed. Conclusions Our method, utilizing the yeast two-hybrid technology and scFv as bait, is a simple yet powerful approach for the detailed characterization of antibody targets. It allows precise

  3. Mechanisms of Mitochondrial Damage in Keratinocytes by Pemphigus Vulgaris Antibodies*

    OpenAIRE

    Kalantari-Dehaghi, Mina; Chen, Yumay; Deng, Wu; Chernyavsky, Alex; Marchenko, Steve; Wang, Ping H.; Grando, Sergei A.

    2013-01-01

    Background: Previous studies suggested that mitochondrial antibodies contribute to pemphigus vulgaris (PV).Results: PV sera elicited mitochondrial damage, and mitochondria-protecting drugs exhibited protective effect in cell culture and mouse skin.Conclusion: PV antibodies altered O2 respiration, disrupted electron transfer chain, and increased reactive oxygen species.Significance: Results provide the mechanism of therapeutic action and justify the use of mitochondria-protecting drugs in PV.T...

  4. Characterization of human serum spreading factor with monoclonal antibody.

    OpenAIRE

    Barnes, D W; Silnutzer, J; See, C; Shaffer, M

    1983-01-01

    Serum spreading factor is a glycoprotein isolated from human serum that promotes spreading of a variety of cell types on culture dishes. We developed mouse hybridoma lines secreting monoclonal antibody to serum spreading factor that markedly inhibited the rate of serum spreading factor-promoted spreading of both fibroblastic and epithelial cells in culture. Fibronectin-promoted cell spreading was unaffected by monoclonal antibody to serum spreading factor, and the factor appeared to be distin...

  5. Antiglomerular basement membrane antibody-crescentic glomerulonephritis complicating chronic bronchiectasis.

    Science.gov (United States)

    Enríquez, R; Cabezuelo, J B; Sirvent, A E; Andrada, E; Amorós, F; Orti, C

    2001-04-01

    A 68-year-old woman with chronic bronchiectasis presented with haematuria and severe oligoanuric renal failure with no other serious systemic manifestation. Antiglomerular basement membrane (anti-GBM) antibodies and anti-myeloperoxidase antibodies were positive. Renal biopsy revealed anti-GBM crescentic glomerulonephritis. A conservative approach was followed and the patient is stable on chronic haemodialysis 6 months later. To the authors' knowledge, there has only been one previous report of anti-GBM disease complicating bronchiectasis.

  6. Interaction of campylobacter species with antibody, complement and phagocytes.

    OpenAIRE

    Bernatowska, E.; Jose, P; Davies, H; Stephenson, M.; Webster, D

    1989-01-01

    The opsonisation of four different campylobacter species for human neutrophils was studied using a chemiluminescence system and electron microscopy. Opsonisation of Campylobacter fetus, Campylobacter coli, and Campylobacter jejuni was mediated by antibody and enhanced by complement. Antibody was not, however, required for the phagocytosis of Campylobacter pylori because it activates the classical pathway of complement directly. This unusual property may be important in the pathogenesis of C p...

  7. Antibody responses in patients with invasive Staphylococcus aureus infections

    OpenAIRE

    Jacobsson, G; Colque-Navarro, P.; Gustafsson, E.; Andersson, R.; Möllby, R

    2010-01-01

    Abstract Correlation between antibody response and clinical outcome in Staphylococcus aureus bacteremia has yielded conflicting results. Immunization schedules have failed in clinical trials. Is the humoral response toward S. aureus of protective nature? A prospective study was performed in patients with invasive S. aureus (ISA) infections during the period 2003?2005. The antibody levels were determined at the beginning and at the end of treatment and one month later (n?=?96, n?=?7...

  8. Current research status of radioimmunotherapy monoclonal antibody drug

    International Nuclear Information System (INIS)

    Radioimmunotherapy (RIT) was one of the most important progresses in the field of cancer therapy over the past 20 years. It has been successfully applied in the treatment of blood system tumors such as NHL. For the utilization of RIT in therapy of solid tumors, however, development of more effective monoclonal antibodies, labeling methods and so on are needed. The current status of radionuclides, monoclonal antibodies and drugs commonly used in the RIT were briefly reviewed. (authors)

  9. Seroprevalence of Leptospira antibodies among populations at risk

    Directory of Open Access Journals (Sweden)

    Abdul-Baki Abdullah Al-Robasi

    2015-03-01

    Full Text Available Objective: This study was performed to assess the Leptospira IgG antibodies seroprevalence among populations at risk in Hodeida Governorate, Yemen. Methods: A total of 200 subjects (136 males and 64 females participated in this study during June and December 2012.They represented 10 sewage workers, 22 butchers, 16 construction workers, 108 agriculture workers, 20 hospital sanitary workers and 24 blood donors. Predesigned questionnaires and consent were taken from each individual. Blood samples were collected from subjects, and the sera were tested by ELISA to detect the presence of leptospira IgG antibodies. The possible related factors for seropositivity were evaluated. Results: Leptospira IgG antibodies were found positive in 42% of the participants. The highest seroprevalence level was detected in sewage workers (80%, followed by hospital sanitary workers (60%, construction workers (37.5% and farmers (37%. The lowest of antibodies was in butchers (36.4%. Seroprevalence among blood donors was 25% which was comparatively less than of the populations at risk. Seropositivity of Leptospira IgG antibodies was found higher among males than females (42.6% vs. 34.4%. The highest Leptospira antibodies seropositivity was among elderly participants (81.8%. The seropositivity of antibodies in population live in rural and urban areas was not significant differences. As for closely contacting with animals, the highest antibodies were discovered in people who had goats (80% and sheep (60.9%. Conclusion: Individuals engaged in risk activities are often exposed to leptospiral infection. Therefore, control and prevention policy toward these people are necessary. J Microbiol Infect Dis 2015;5(1: 1-4

  10. Development and characterization of antibody reagents for detecting nanoparticles

    OpenAIRE

    Ravichandran, Supriya; Sullivan, Mark A.; Callahan, Linda M.; Bentley, Karen L.; DeLouise, Lisa A.

    2015-01-01

    The increasing use of nanoparticles (NPs) in technological applications and in commercial products has escalated environmental health and safety concerns. The detection of NPs in the environment and in biological systems is challenged by limitations associated with commonly used analytical techniques. In this paper we report on the development and characterization of NP binding antibodies, termed NProbes. Phage display methodology was used to discover antibodies that bind NPs dispersed in sol...

  11. Transfusion management of patients with red blood cell antibodies

    OpenAIRE

    Bujandrić Nevenka B.; Grujić Jasmina N.; Krga-Milanović Mirjana M.

    2013-01-01

    Introduction. Red blood cell antibodies may cause a positive result of pre-transfusion blood compatibility testing (crossmatch test). It can be a problem to provide suitable blood units for patients with clinically significant antibodies to high-frequency antigens as well as for those with multiple alloantibody specificities. This study was aimed at identifying transfused patients in the population of South-Backa who had developed clinically significant red...

  12. Monoclonal IgA Antibodies for Aflatoxin Immunoassays

    Science.gov (United States)

    Ertekin, Özlem; Pirinçci, Şerife Şeyda; Öztürk, Selma

    2016-01-01

    Antibody based techniques are widely used for the detection of aflatoxins which are potent toxins with a high rate of occurrence in many crops. We developed a murine monoclonal antibody of immunoglobulin A (IgA) isotype with a strong binding affinity to aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1), aflatoxin G2 (AFG2) and aflatoxin M1 (AFM1). The antibody was effectively used in immunoaffinity column (IAC) and ELISA kit development. The performance of the IACs was compatible with AOAC performance standards for affinity columns (Test Method: AOAC 991.31). The total binding capacity of the IACs containing our antibody was 111 ng, 70 ng, 114 ng and 73 ng for AFB1, AFB2, and AFG1 andAFG2, respectively. Furthermore, the recovery rates of 5 ng of each AF derivative loaded to the IACs were determined as 104.9%, 82.4%, 85.5% and 70.7% for AFB1, AFB2, AFG1 and AFG2, respectively. As for the ELISA kit developed using non-oriented, purified IgA antibody, we observed a detection range of 2–50 µg/L with 40 min total test time. The monoclonal antibody developed in this research is hitherto the first presentation of quadruple antigen binding IgA monoclonal antibodies in mycotoxin analysis and also the first study of their utilization in ELISA and IACs. IgA antibodies are valuable alternatives for immunoassay development, in terms of both sensitivity and ease of preparation, since they do not require any orientation effort. PMID:27187470

  13. Antibody therapeutics targeting ion channels: are we there yet?

    Science.gov (United States)

    Sun, Han; Li, Min

    2013-01-01

    The combination of technological advances, genomic sequences and market success is catalyzing rapid development of antibody-based therapeutics. Cell surface receptors and ion channel proteins are well known drug targets, but the latter has seen less success. The availability of crystal structures, better understanding of gating biophysics and validation of physiological roles now form an excellent foundation to pursue antibody-based therapeutics targeting ion channels to treat a variety of diseases. PMID:23381110

  14. Integrated Design of Antibodies for Systems Biology Using Ab Designer

    OpenAIRE

    Pisitkun, Trairak; Dummer, Patrick; Somparn, Poorichaya; Hirankarn, Nattiya; Kopp, Jeffrey B.; Knepper, Mark A.

    2014-01-01

    In the current era of large-scale biology, systems biology has evolved as a powerful approach to identify complex interactions within biological systems. In addition to high throughput identification and quantification techniques, methods based on high-quality mono-specific antibodies remain an essential element of the approach. To assist the large-scale design and production of peptide-directed antibodies for systems biology studies, we developed a fully integrated online application, AbDesi...

  15. Antierythropoietin Antibodies in Hemodialysis Patients Treated with Recombinant Erythropoietin

    OpenAIRE

    Savaş ÖZTÜRK; Alper GÜMÜŞ; Vecihi MEMİLİ; Muhammet Emin DÜZ; Egemen CEBECİ; Macit KOLDAŞ; Rümeyza KAZANCIOĞLU

    2014-01-01

    OBJECTIVE: Erythropoietin resistance is a serious problem in patients treated with recombinant erythropoietin. Antierythropoietin antibodies are considered to be one of the causes of this resistance. MATERIAL and ME THODS: We investigated antierythropoietin antibodies in chronic hemodialysis patients and compared the results with healthy controls by means of establishing an ELISA method. A total of 121 chronic hemodialysis patients receiving recombinant erythropoietin were included in the ...

  16. Production and characterization of yeast killer toxin monoclonal antibodies

    OpenAIRE

    Polonelli, L; Morace, G

    1987-01-01

    Monoclonal antibodies were obtained after fusion of mouse myeloma cells with spleen cells isolated from mice primed with a crude extract of yeast killer toxin produced by a strain of Hansenula anomala. Hybridomas were selected by specific immunoassay reaction of their fluid with crude yeast killer toxin extract. Among the monoclonal antibodies, which were characterized by the Western blot technique, one (designated KT4) proved to have precipitating properties, thus permitting the neutralizati...

  17. Antibody-mediated Prevention of Fusarium Mycotoxins in the Field

    OpenAIRE

    Yu-Cai Liao; Elena Glinka; Jing-Bo Zhang; He-Ping Li; Zu-Quan Hu

    2008-01-01

    Fusarium mycotoxins directly accumulated in grains during the infection of wheat and other cereal crops by Fusarium head blight (FHB) pathogens are detrimental to humans and domesticated animals. Prevention of the mycotoxins via the development of FHB-resistant varieties has been a challenge due to the scarcity of natural resistance against FHB pathogens. Various antibodies specific to Fusarium fungi and mycotoxins are widely used in immunoassays and antibody-mediated resistance in planta aga...

  18. Changes of antithroglobulin antibody in children with congenital hypothyroidism

    OpenAIRE

    Cho, Eun Mi; Kim, Uk Hyun; Choi, Byung Ho; Ko, Cheol Woo

    2013-01-01

    Purpose It has been reported that antithroglobulin (anti-TG) antibody is increased in the sera of both children with transient congenital hypothyroidism and their mothers. And transplacental transport of thyroid autoantibody was proposed as the pathogenesis of transient congenital hypothyroidism. However this is not known in nontransient congenital hypothyroidism. This study was done to see changes of anti-TG antibody in children with nontransient congenital hypothyroidism. Methods Study pati...

  19. Characterization of a monoclonal antibody to human erythropoietin.

    OpenAIRE

    Weiss, T L; Kavinsky, C J; Goldwasser, E

    1982-01-01

    Hybrid cells that synthesize a monospecific antibody directed toward erythropoietin have been produced by the fusion of mouse plasmacytoma cells with spleen cells from rats immunized against human erythropoietin. The antibody binds the alpha and beta forms and the asialo alpha form of erythropoietin to the same extent. It is an immunoglobulin of the IgG class and binds only erythropoietin in an impure preparation of the hormone. Biologically active unlabeled erythropoietin competes with biolo...

  20. Production and Purification of Rabbit's Polyclonal Antibody Against Factor VIII.

    OpenAIRE

    Sohrabi, Simin; Akbarzadeh, Azim; Norouzian, Dariush; Farhangi, Ali; Mortazavi, Mehri; Mehrabi, Mohammad Reza; Chiani, Mohsen; Saffari, Zahra; Ghassemi, Soheil

    2011-01-01

    The attempt is made to produce recombinant factor VIII but the first step in producing such product is production and purification of rabbit's polyclonal antibody against factor VIII. The second and third steps involve monoclonal antibody and recombinant factor VIII production. Factor VIII is one of the most important coagulating factor where its deficiency leads to diseases like hemophilia type A or classic. It is an inherited disease. Previously, it was obtained through fractionation of blo...