Immunospecific red cell binding of iodine 125-labeled immunoglobulin G erythrocyte autoantibodies

International Nuclear Information System (INIS)

Masouredis, S.P.; Branks, M.J.; Garratty, G.; Victoria, E.J.

1987-01-01

The primary interaction of autoantibodies with red cells has been studied by using labeled autoantibodies. Immunoglobulin G red cell autoantibodies obtained from IgG antiglobulin-positive normal blood donors were labeled with radioactive iodine and compared with alloanti-D with respect to their properties and binding behavior. Iodine 125 -labeled IgG autoantibody migrated as a single homogeneous peak with the same relative mobility as human IgG on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The isoelectric focusing pattern of labeled autoantibodies varied from donor to donor but was similar to that of alloanti-D, consisting of multiple IgG populations with isoelectric points in the neutral to alkaline range. 125 I-autoantibody bound to all human red cells of common Rh phenotypes. Evidence for immunospecific antibody binding of the labeled autoantibody was based on variation in equilibrium binding to nonhuman and human red cells of common and rare phenotypes, enhanced binding after red cell protease modification, antiglobulin reactivity of cell-bound IgG comparable to that of cell-bound anti-D, and saturation binding in autoantibody excess. Scatchard analysis of two 125 I-autoantibody preparations yielded site numbers of 41,500 and 53,300 with equilibrium constants of 3.7 and 2.1 X 10(8) L X mol-1. Dog, rabbit, rhesus monkey, and baboon red cells were antigen(s) negative by quantitative adsorption studies adsorbing less than 3% of the labeled autoantibody. Reduced ability of rare human D--red blood cells to adsorb the autoantibody and identification of donor autoantibodies that bind to Rh null red blood cells indicated that eluates contained multiple antibody populations of complex specificities in contrast to anti-D, which consists of a monospecific antibody population. Another difference is that less than 70% of the autoantibody IgG was adsorbed by maximum binding red blood cells as compared with greater than 85% for alloanti-D

Multifactorial aspects of antibody-mediated blood cell destruction

NARCIS (Netherlands)

Kapur, R.

2014-01-01

The research described in this thesis focuses on diseases of antibody-mediated blood cell destruction via FcγRs on phagocytes, in particular regarding platelets in fetal or neonatal alloimmune thrombocytopenia (FNAIT) and red blood cells (RBC) in hemolytic disease of the fetus and newborn (HDFN).

Enhanced neutralization potency of botulinum neurotoxin antibodies using a red blood cell-targeting fusion protein.

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Sharad P Adekar

2011-03-01

Full Text Available Botulinum neurotoxin (BoNT potently inhibits cholinergic signaling at the neuromuscular junction. The ideal countermeasures for BoNT exposure are monoclonal antibodies or BoNT antisera, which form BoNT-containing immune complexes that are rapidly cleared from the general circulation. Clearance of opsonized toxins may involve complement receptor-mediated immunoadherence to red blood cells (RBC in primates or to platelets in rodents. Methods of enhancing immunoadherence of BoNT-specific antibodies may increase their potency in vivo. We designed a novel fusion protein (FP to link biotinylated molecules to glycophorin A (GPA on the RBC surface. The FP consists of an scFv specific for murine GPA fused to streptavidin. FP:mAb:BoNT complexes bound specifically to the RBC surface in vitro. In a mouse model of BoNT neutralization, the FP increased the potency of single and double antibody combinations in BoNT neutralization. A combination of two antibodies with the FP gave complete neutralization of 5,000 LD50 BoNT in mice. Neutralization in vivo was dependent on biotinylation of both antibodies and correlated with a reduction of plasma BoNT levels. In a post-exposure model of intoxication, FP:mAb complexes gave complete protection from a lethal BoNT/A1 dose when administered within 2 hours of toxin exposure. In a pre-exposure prophylaxis model, mice were fully protected for 72 hours following administration of the FP:mAb complex. These results demonstrate that RBC-targeted immunoadherence through the FP is a potent enhancer of BoNT neutralization by antibodies in vivo.

Enhanced opsonisation of Rhesus D-positive human red blood cells by recombinant polymeric immunoglobulin G anti-G antibodies.

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Díaz-Solano, Dylana; Fuenmayor, Jaheli; Montaño, Ramon F

2018-02-01

Anti-RhD antibodies (anti-D) are important in the prophylaxis of haemolytic disease of the foetus and newborn (HDFN) due to RhD incompatibility. Current preparations of anti-D are sourced from hyperimmune human plasma, so its production carries a risk of disease and is dependent on donor availability. Despite the efforts to develop a monoclonal preparation with similar prophylactic properties to the plasma-derived anti-D, no such antibody is yet available. Here we studied the agglutinating, opsonic and haemolytic activities of two recombinant polymeric immunoglobulins (Ig) against the G antigen of the Rh complex. Recombinant polymeric anti-G IgG1 (IgG1μtp) and IgG3 (IgG3μtp) were produced in vitro, purified by protein G-affinity chromatography, and analysed by gel electrophoresis. Their agglutinating, opsonic and haemolytic activities were evaluated using haemagglutination, erythrophagocytosis, and complement activation assays. The recombinant IgG1μtp and IgG3μtp anti-G antibodies ranged from 150,000 to 1,000,000 Da in molecular weight, indicating the formation of polymeric IgG. No complement activation or haemolytic activity was detected upon incubation of RhD-positive red-blood cells with the polymeric anti-G IgG. Both polymers were better opsonins than a prophylactic preparation of plasma-derived anti-D. The enhanced opsonic properties of the polymeric anti-G IgG1μtp and IgG3μtp could allow them to mediate the clearance of RhD-positive red blood cells from circulation more efficiently than natural or other synthetic prophylactic anti-D options. Their inability to induce complement-mediated haemolysis would be prophylactically convenient and is comparable in vitro to that of the available plasma-derived polyclonal anti-D preparations. The described properties suggest that polymeric antibodies like these (but with anti-D specificity) may be testable candidates for prophylaxis of HDFN caused by anti-D.

Sperm, nuclear, phospholipid, and red blood cell antibodies and isotype RF in infertile couples and patients with autoimmune rheumatic diseases.

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Fichorova, R; Nakov, L; Baleva, M; Nikolov, K; Gegova, I

1996-12-01

To determine if measuring of nonorgan-specific autoantibodies is useful for better understanding and management of unexplained infertility. Sera were obtained from 70 infertile couples, 57 rheumatic patients, and 76 fertile donors. Sperm antibodies (SA) were detected by the tests of Kibrick and Friberg, anti-histones, anti-cardiolipin antibodies, and RF isotypes by ELISA, antinuclear antibodies by indirect immunofluorescence, and anti-red blood cell antibodies by Capture-R. Multiple autoimmune reactivity (both partners positive and/or more than one type of autoantibody involved), higher than naturally occurring in fertile individuals, was found in 55% of the idiopathically infertile couples. IgA-RF was the dominant autoimmune marker. SA revealed similar rates in patients with rheumatic diseases and in infertiles with or without other autoantibodies. Although no single autoimmunity marker could predict occurrence of SA, the coincidence of enhanced polyclonal autoimmunity in both partners of infertile couples might potentiate their negative effect on reproduction.

Clearance of Human IgG1-Sensitised Red Blood Cells In Vivo in Humans Relates to the In Vitro Properties of Antibodies from Alternative Cell Lines

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Armour, Kathryn L.; Smith, Cheryl S.; Ip, Natasha C. Y.; Ellison, Cara J.; Kirton, Christopher M.; Wilkes, Anthony M.; Williamson, Lorna M.; Clark, Michael R.

2014-01-01

We previously produced a recombinant version of the human anti-RhD antibody Fog-1 in the rat myeloma cell line, YB2/0. When human, autologous RhD-positive red blood cells (RBC) were sensitised with this IgG1 antibody and re-injected, they were cleared much more rapidly from the circulation than had been seen earlier with the original human-mouse heterohybridoma-produced Fog-1. Since the IgG have the same amino acid sequence, this disparity is likely to be due to alternative glycosylation that results from the rat and mouse cell lines. By comparing the in vitro properties of YB2/0-produced Fog-1 IgG1 and the same antibody produced in the mouse myeloma cell line NS0, we now have a unique opportunity to pinpoint the cause of the difference in ability to clear RBC in vivo. Using transfected cell lines that express single human FcγR, we showed that IgG1 made in YB2/0 and NS0 cell lines bound equally well to receptors of the FcγRI and FcγRII classes but that the YB2/0 antibody was superior in FcγRIII binding. When measuring complexed IgG binding, the difference was 45-fold for FcγRIIIa 158F, 20-fold for FcγRIIIa 158V and approximately 40-fold for FcγRIIIb. The dissimilarity was greater at 100-fold in monomeric IgG binding assays with FcγRIIIa. When used to sensitise RBC, the YB2/0 IgG1 generated 100-fold greater human NK cell antibody-dependent cell-mediated cytotoxicity and had a 103-fold advantage over the NS0 antibody in activating NK cells, as detected by CD54 levels. In assays of monocyte activation and macrophage adherence/phagocytosis, where FcγRI plays major roles, RBC sensitised with the two antibodies produced much more similar results. Thus, the alternative glycosylation profiles of the Fog-1 antibodies affect only FcγRIII binding and FcγRIII-mediated functions. Relating this to the in vivo studies confirms the importance of FcγRIII in RBC clearance. PMID:25302805

Clearance of human IgG1-sensitised red blood cells in vivo in humans relates to the in vitro properties of antibodies from alternative cell lines.

Directory of Open Access Journals (Sweden)

Kathryn L Armour

Full Text Available We previously produced a recombinant version of the human anti-RhD antibody Fog-1 in the rat myeloma cell line, YB2/0. When human, autologous RhD-positive red blood cells (RBC were sensitised with this IgG1 antibody and re-injected, they were cleared much more rapidly from the circulation than had been seen earlier with the original human-mouse heterohybridoma-produced Fog-1. Since the IgG have the same amino acid sequence, this disparity is likely to be due to alternative glycosylation that results from the rat and mouse cell lines. By comparing the in vitro properties of YB2/0-produced Fog-1 IgG1 and the same antibody produced in the mouse myeloma cell line NS0, we now have a unique opportunity to pinpoint the cause of the difference in ability to clear RBC in vivo. Using transfected cell lines that express single human FcγR, we showed that IgG1 made in YB2/0 and NS0 cell lines bound equally well to receptors of the FcγRI and FcγRII classes but that the YB2/0 antibody was superior in FcγRIII binding. When measuring complexed IgG binding, the difference was 45-fold for FcγRIIIa 158F, 20-fold for FcγRIIIa 158V and approximately 40-fold for FcγRIIIb. The dissimilarity was greater at 100-fold in monomeric IgG binding assays with FcγRIIIa. When used to sensitise RBC, the YB2/0 IgG1 generated 100-fold greater human NK cell antibody-dependent cell-mediated cytotoxicity and had a 103-fold advantage over the NS0 antibody in activating NK cells, as detected by CD54 levels. In assays of monocyte activation and macrophage adherence/phagocytosis, where FcγRI plays major roles, RBC sensitised with the two antibodies produced much more similar results. Thus, the alternative glycosylation profiles of the Fog-1 antibodies affect only FcγRIII binding and FcγRIII-mediated functions. Relating this to the in vivo studies confirms the importance of FcγRIII in RBC clearance.

Women's attitude towards prenatal screening for red blood cell antibodies, other than RhD

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van der Schoot CE

2008-11-01

Full Text Available Abstract Background Since July 1998 all Dutch women (± 200,000/y are screened for red cell antibodies, other than anti-RhesusD (RhD in the first trimester of pregnancy, to facilitate timely treatment of pregnancies at risk for hemolytic disease of the fetus and newborn (HDFN. Evidence for benefits, consequences and costs of screening for non-RhD antibodies is still under discussion. The screening program was evaluated in a nation-wide study. As a part of this evaluation study we investigated, according to the sixth criterium of Wilson and Jüngner, the acceptance by pregnant women of the screening program for non-RhD antibodies. Methods Controlled longitudinal survey, including a prenatal and a postnatal measurement by structured questionnaires. Main outcome measures: information satisfaction, anxiety during the screening process (a.o. STAI state inventory and specific questionnaire modules, overall attitude on the screening program. Univariate analysis was followed by standard multivariate analysis to identify significant predictors of the outcome measures. Participants: 233 pregnant women, distributed over five groups, according to the screening result. Results Satisfaction about the provided information was moderate in all groups. All screen- positive groups desired more supportive information. Anxiety increased in screen- positives during the screening process, but decreased to basic levels postnatally. All groups showed a strongly positive balance between perceived utility and burden of the screening program, independent on test results or background characteristics. Conclusion Women highly accept the non-RhD antibody screening program. However, satisfaction about provided information is moderate. Oral and written information should be provided by obstetric care workers themselves, especially to screen-positive women.

Avian influenza virus antibodies in Pacific Coast Red Knots (Calidris canutus rufa)

Science.gov (United States)

Johnson, James A.; DeCicco, Lucas H.; Ruthrauff, Daniel R.; Krauss, Scott; Hall, Jeffrey S.

2014-01-01

Prevalence of avian influenza virus (AIV) antibodies in the western Atlantic subspecies of Red Knot (Calidris canutus rufa) is among the highest for any shorebird. To assess whether the frequency of detection of AIV antibodies is high for the species in general or restricted only to C. c. rufa, we sampled the northeastern Pacific Coast subspecies of Red Knot (Calidris canutus roselaari) breeding in northwestern Alaska. Antibodies were detected in 90% of adults and none of the chicks sampled. Viral shedding was not detected in adults or chicks. These results suggest a predisposition of Red Knots to AIV infection. High antibody titers to subtypes H3 and H4 were detected, whereas low to intermediate antibody levels were found for subtypes H10 and H11. These four subtypes have previously been detected in shorebirds at Delaware Bay (at the border of New Jersey and Delaware) and in waterfowl along the Pacific Coast of North America.

Asymmetries in Chickens from Lines Selected and Relaxed for High or Low Antibody Titers to Sheep Red Blood Cells

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Yunjie Tu

2015-03-01

Full Text Available Wattle length, width, and area were measured to classify bilateral asymmetries in four lines of chickens. The lines were the S26 generation of White Leghorns selected for high (HAS or low (LAS response to sheep red blood cells and sublines in which selection had been relaxed for three generations (high antibody relaxed [HAR] and low antibody relaxed [LAR]. Antibody titers (AB were greater for HAS than for HAR with both greater than for LAS and LAR which while different for males did not differ for females. The low antibody lines were heavier and reached sexual maturity at younger age than the high antibody lines. In general, wattle length, width, and area were greater in the low than high antibody lines. In 24 comparisons for bilaterality 18 exhibited fluctuating asymmetry and 6 exhibited directional asymmetry with 5 of the 6 being for wattle length. There was not a clear pattern for changes in degree of asymmetry when selection was relaxed for 3 generations. For females, the relative asymmetry (RA of wattle area was larger (p≤0.05 for HAR than for LAR and not different from the selected lines and relaxed lines. There were no differences among lines for RA of wattle length and width of females and wattle length, width, and area of males.

Antibodies to Rickettsia spp. and Borrelia burgdorferi in Spanish Wild Red Foxes (Vulpes vulpes).

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Lledó, Lourdes; Serrano, José Luis; Isabel Gegúndez, María; Giménez-Pardo, Consuelo; Saz, José Vicente

2016-01-01

We examined 314 red foxes (Vulpes vulpes) from the province of Soria, Spain, for Rickettsia typhi, Rickettsia slovaca, and Borrelia burgdorferi infection. Immunofluorescence assays showed 1.9% had antibodies to R. typhi, 6.7% had antibodies to R. slovaca, and 8.3% had antibodies to B. burgdorferi. Serostatus was not correlated with sex or age. Because red foxes can be infected by Rickettsiae and B. burgdorferi, presence of red foxes may be and indicator for the presence of these pathogens.

Antibody recognition of Plasmodium falciparum infected red blood cells by symptomatic and asymptomatic individuals in the Brazilian Amazon

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Fratus, Alessandra Sampaio Bassi; Cabral, Fernanda Janku; Fotoran, Wesley Luzetti; Medeiros, Márcia Melo; Carlos, Bianca Cechetto; Martha, Rosimeire dalla; da Silva, Luiz Hildebrando Pereira; Lopes, Stefanie Costa Pinto; Costa, Fabio Trindade Maranhão; Wunderlich, Gerhard

2014-01-01

In the Amazon Region, there is a virtual absence of severe malaria and few fatal cases of naturally occurring Plasmodium falciparum infections; this presents an intriguing and underexplored area of research. In addition to the rapid access of infected persons to effective treatment, one cause of this phenomenon might be the recognition of cytoadherent variant proteins on the infected red blood cell (IRBC) surface, including the var gene encoded P. falciparum erythrocyte membrane protein 1. In order to establish a link between cytoadherence, IRBC surface antibody recognition and the presence or absence of malaria symptoms, we phenotype-selected four Amazonian P. falciparum isolates and the laboratory strain 3D7 for their cytoadherence to CD36 and ICAM1 expressed on CHO cells. We then mapped the dominantly expressed var transcripts and tested whether antibodies from symptomatic or asymptomatic infections showed a differential recognition of the IRBC surface. As controls, the 3D7 lineages expressing severe disease-associated phenotypes were used. We showed that there was no profound difference between the frequency and intensity of antibody recognition of the IRBC-exposed P. falciparum proteins in symptomatic vs. asymptomatic infections. The 3D7 lineages, which expressed severe malaria-associated phenotypes, were strongly recognised by most, but not all plasmas, meaning that the recognition of these phenotypes is frequent in asymptomatic carriers, but is not necessarily a prerequisite to staying free of symptoms. PMID:25099336

Hemolytic disease of the fetus and newborn due to multiple maternal antibodies.

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Markham, Kara Beth; Rossi, Karen Q; Nagaraja, Haikady N; O'Shaughnessy, Richard W

2015-07-01

The objective of the study was to determine whether women with combinations of red blood cell antibodies are more likely to develop significant hemolytic disease of the fetus and newborn than those with single antibodies. A retrospective exposure cohort study was conducted of pregnant women with red blood cell antibodies. The development of significant hemolytic disease of the fetus and newborn was then compared between patients with single antibodies and those with multiple antibodies. Data analysis was limited to pregnancies delivering since the year 2000. Thirteen percent of the patients referred to our program had multiple red blood cell antibodies. Odds of developing significant hemolytic disease of the fetus and newborn for patients with anti-Rh(D) combined with at least 1 additional red blood cell antibody were 3.65 times the odds for women with anti-Rh(D) antibodies in isolation (95% confidence interval, 1.84-7.33). In the setting of multiple antibodies including anti-Rh(D), Rh-positive fetuses/neonates have an increased odds of developing significant hemolytic disease even if the fetus is negative for the other corresponding red blood cell antigen. Women with multiple red blood cell antibodies are more likely to develop significant hemolytic disease of the fetus and newborn than those with a single antibody especially in the presence of anti-(Rh)D. This pathophysiology may suggest a more aggressive immune response in women who develop more than 1 red blood cell antibody. Copyright © 2015 Elsevier Inc. All rights reserved.

Red blood cell antibodies in pregnancy and their clinical consequences

DEFF Research Database (Denmark)

Nordvall, Maria; Dziegiel, Morten Hanefeld; Hegaard, Hanne Kristine

2009-01-01

The objective was to determine clinical consequences of various specificities for the infant/fetus. The population was patients referred between 1998 and 2005 to the tertiary center because of detected red blood cell (RBC) alloimmunization. Altogether 455 infants were delivered by 390 alloimmuniz...

Antiparietal cell antibody test

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APCA; Anti-gastric parietal cell antibody; Atrophic gastritis - anti-gastric parietal cell antibody; Gastric ulcer - anti-gastric parietal cell antibody; Pernicious anemia - anti-gastric parietal cell antibody; ...

Hyperreactive malarial splenomegaly is associated with low levels of antibodies against red blood cell and Plasmodium falciparum derived glycolipids in Yanomami Amerindians from Venezuela.

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Vivas, Livia; O'Dea, Kieran P; Noya, Oscar; Pabon, Rosalba; Magris, Magda; Botto, Carlos; Holder, Anthony A; Brown, K Neil

2008-03-01

The immunological basis of the aberrant immune response in hyperreactive malarial splenomegaly (HMS) is poorly understood, but believed to be associated with polyclonal B cell activation by an unidentified malaria mitogen, leading to unregulated immunoglobulin and autoantibody production. HMS has been previously reported in Yanomami communities in the Upper Orinoco region of the Venezuelan Amazon. To investigate a possible association between antibody responses against Plasmodium falciparum and uninfected red blood cell (URBC) glycolipids and splenomegaly, a direct comparison of the parasite versus host anti-glycolipid antibody responses was made in an isolated community of this area. The anti-P. falciparum glycolipid (Pfglp) response was IgG3 dominated, whereas the uninfected red blood cell glycolipid (URBCglp) response showed a predominance of IgG1. The levels of IgG1 against Pfglp, and of IgG4 and IgM against URBCglp were significantly higher in women, while the anti-Pfglp or URBCglp IgM levels were inversely correlated with the degree of splenomegaly. Overall, these results suggest differential regulation of anti-parasite and autoreactive responses and that these responses may be linked to the development and evolution of HMS in this population exposed to endemic malaria. The high mortality rates associated with HMS point out that its early diagnosis together with the implementation of malaria control measures in these isolated Amerindian communities are a priority.

Detection of adult T-cell leukemia virus (ATLV) bearing lymphocytes in concentrated red blood cells derived from ATL associated antibody (ATLA-Ab) positive donors.

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Morishima, Y; Ohya, K; Ueda, R; Fukuda, T

1986-01-01

Adult T cell leukemia associated antibody (ATLA-Ab) positive persons were screened by indirect immunofluorescence (IF) testing. Their lymphocytes were collected from concentrated red blood cells (CRC), and cultured in vitro with and without phytohemagglutinin (PHA) for 10 days. The expression of ATL virus (ATLV) positive lymphocytes during the in vitro culture was then analyzed by IF assay using mouse monoclonal antibody ATL-19 reactive to p19 core protein of ATLV. 97% of ATLA-Ab positive CRC (36 cases) demonstrated ATLV positive lymphocytes after being cultured for more than 10 days with PHA, whereas, none of ATLA-Ab negative CRC (22 cases) demonstrated ATLV positive lymphocytes. All of the 10 ATLA-Ab positive CRC that were stored for 2, 4, and 7 days contained lymphocytes which expressed ATLV after in vitro culture, while 7 of 10 CRC stored for 14 days and only 1 of 10 CRCs stored for 20 days, expressed ATLV positive lymphocytes. This data indicates that almost all of the ATLA-Ab positive blood contained ATLV positive lymphocytes, and that the in vitro appearance of these ATLV positive lymphocytes was reduced by storing the CRC for more than 14 days.

[Immunohematologic study and transfusion approach to patients with public antibodies].

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Solves, P; de la Rubia, J; Arriaga, F; Cervera, J; Arnao, M; Carpio, N; Marty, M L

1997-02-01

To analyze the different immunohematologic studies required to identify anti-red cell antibodies directed against high incidence antigens and comment the best tranfusion management. Five patients with suspected anti-red cell alloantibodies directed against high frequency antigens are reported. After a positive antibody screening test (AST), an agglutination test with a commercial panel of 24 red cells was performed. Red cells were treated with proteolytic enzymes and AET to try to identify the circulating antibody. However, it was necessary to send the samples to reference laboratories for definitive identification. In order to evaluate the haemolytic potential of the antibody serum samples were treated with DTT and immunoglobulin subtype was studied with the capillary agglutination test. Finally, we analyze the half life of Cr51 labelled red cells. To obtain compatible blood for transfusion, autologous transfusion and cross-match with blood from direct relatives were performed. AST was positive in every case. A decrease in the agglutination test was observed after ficin treatment in two patients, and an increase in the remaining. The treatment of red cells with ZZAP and AET resulted in a decrease of agglutination in three cases and an increase in the remaining two. Specificity of the antibodies was as follows: anti-Cellano (two cases), anti-Ku (one case) and anti-Yta (two cases). Anti-Kell antibodies were IgG1 and anti-Cartwright antibodies were IgG4. One patient was transfused with autologous blood alone, another patient received compatible blood from direct relatives. A third patient was transfused both with autologous and allogeneic compatible blood. The fourth patient did not need red cell transfusion and, finally the last patient had to be transfused with incompatible blood but no postransfusion haemolysis was observed. In patients with anti-red cell antibodies against high-frequency antigens, red blood cells treatment with proteolytic enzymes (ZZAP, ficin

Severe hemolytic disease of fetus and newborn caused by red blood cell antibodies undetected at first-trimester screening (CME).

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Dajak, Slavica; Stefanović, Vedran; Capkun, Vesna

2011-07-01

The objective was to determine clinical consequences of anti-D and non-D antibodies undetected at first-trimester screening for infant or fetus. This retrospective cohort study included all pregnant women with red blood cell (RBC) antibodies who were tested between 1993 and 2008. Data were obtained from the forms for tracking immunization at the transfusion department. Each form was analyzed for three data sets: the order of screening at which the antibodies were detected (initial or repeated screening), the order of pregnancy (first pregnancy or higher), and whether the antibodies caused severe hemolytic disease of fetus and newborn (HDFN). In D- women, anti-D was detected in 1.3% of cases. The anti-D was undetected in 72 (37%) cases on the first-trimester screening, of which eight cases were complicated by severe HDFN. In this group, three patients were primigravidae. An overall non-D incidence of 0.2% was observed. In 16 cases, non-D were undetected on the first-trimester screening (10 anti-c, two anti-E, two anti-C, one anti-S, and one case of anti-Rh17). Non-D antibodies undetected on initial screening caused 11 cases of severe HDFN (27% of all severe non-D HDFN). Ten of them were in multiparous women. Seven of 11 cases with severe HDFN that were missed were caused by anti-c. The third-trimester screening may detect RBC antibodies that were not present or detected on the first-trimester screening. Such screening may be especially relevant in D+ multiparous women due to the risk of HDFN. © 2010 American Association of Blood Banks.

Assessment of Red Blood Cell Parameters and Peripheral Smear at ...

African Journals Online (AJOL)

Cold agglutination disease (CAD) is characterized by an auto‑antibody which is able to agglutinate red blood cells (RBCs) at temperatures lower than that of the body, and subsequently to activate the complement system responsible for lysis of RBCs. Patients show hemolytic anemia of varying degrees of severity, which ...

Human monoclonal antibodies reactive with human myelomonocytic leukemia cells.

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Posner, M R; Santos, D J; Elboim, H S; Tumber, M B; Frackelton, A R

1989-04-01

Peripheral blood mononuclear cells from a patient with chronic myelogenous leukemia (CML), in remission, were depleted of CD8-positive T-cells and cultured with Epstein-Barr virus. Four of 20 cultures (20%) secreted human IgG antibodies selectively reactive with the cell surfaces of certain human leukemia cell lines. Three polyclonal, Epstein-Barr virus-transformed, B-cell lines were expanded and fused with the human-mouse myeloma analogue HMMA2.11TG/O. Antibody from secreting clones HL 1.2 (IgG1), HL 2.1 (IgG3), and HL 3.1 (IgG1) have been characterized. All three react with HL-60 (promyelocytic), RWLeu4 (CML promyelocytic), and U937 (monocytic), but not with KG-1 (myeloblastic) or K562 (CML erythroid). There is no reactivity with T-cell lines, Burkitt's cell lines, pre-B-leukemia cell lines, or an undifferentiated CML cell line, BV173. Leukemic cells from two of seven patients with acute myelogenous leukemia and one of five with acute lymphocytic leukemia react with all three antibodies. Normal lymphocytes, monocytes, polymorphonuclear cells, red blood cells, bone marrow cells, and platelets do not react. Samples from patients with other diverse hematopoietic malignancies showed no reactivity. Immunoprecipitations suggest that the reactive antigen(s) is a lactoperoxidase iodinatable series of cell surface proteins with molecular weights of 42,000-54,000 and a noniodinatable protein with a molecular weight of 82,000. Based on these data these human monoclonal antibodies appear to react with myelomonocytic leukemic cells and may detect a leukemia-specific antigen or a highly restricted differentiation antigen.

Detection and quantification of Duffy antigen on bovine red blood cell membranes using a polyclonal antibody

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Ana Teresa B.F. Antonangelo

2012-09-01

Full Text Available Babesiosis is one of the most important diseases affecting livestock agriculture worldwide. Animals from the subspecies Bos taurus indicus are more resistant to babesiosis than those from Bos taurus taurus. The genera Babesia and Plasmodium are Apicomplexa hemoparasites and share features such as invasion of red blood cells (RBC. The glycoprotein Duffy is the only human erythrocyte receptor for Pasmodium vivax and a mutation which abolishes expression of this glycoprotein on erythrocyte surfaces is responsible for making the majority of people originating from the indigenous populations of West Africa resistant to P. vivax. The current work detected and quantified the Duffy antigen on Bos taurus indicus and Bos taurus taurus erythrocyte surfaces using a polyclonal antibody in order to investigate if differences in susceptibility to Babesia are due to different levels of Duffy antigen expression on the RBCs of these animals, as is known to be the case in human beings for interactions of Plasmodium vivax-Duffy antigen. ELISA tests showed that the antibody that was raised against Duffy antigens detected the presence of Duffy antigen in both subspecies and that the amount of this antigen on those erythrocyte membranes was similar. These results indicate that the greater resistance of B. taurus indicus to babesiosis cannot be explained by the absence or lower expression of Duffy antigen on RBC surfaces.

Emergency transfusion of patients with unknown blood type with blood group O Rhesus D positive red blood cell concentrates: a prospective, single-centre, observational study.

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Selleng, Kathleen; Jenichen, Gregor; Denker, Kathrin; Selleng, Sixten; Müllejans, Bernd; Greinacher, Andreas

2017-05-01

Emergency patients with unknown blood type usually receive O Rhesus D negative (RhD-) red blood cell concentrates until their blood group is determined to prevent RhD+ related adverse transfusion reactions. As 85% of individuals are RhD+, this consumption of O RhD- red blood cell concentrates contributes to shortages of O RhD- red blood cell concentrates, sometimes forcing transfusion of known RhD- patients with RhD+ red blood cell concentrates. Here we report the outcome of this transfusion policy transfusing all emergency patients with unknown blood type with O RhD+ red blood cell concentrates. In this prospective single-centre observational study done between Jan 1, 2001, and Dec 31, 2015, we assessed all consecutive RhD- patients at the University Medicine Greifswald who received RhD+ red blood cell concentrates (emergency patients with unknown blood type; and RhD- patients receiving RhD+ red blood cell concentrates during RhD- red blood cell concentrate shortages). No patients were excluded. The primary endpoint was anti-D allo-immunisation at 2 months follow-up or later. Patients were followed up and tested for immunisation against red blood cell antigens using the direct antiglobulin test and an antibody screen every 3-5 days for 4 weeks or until death, or hospital discharge. Surviving patients were screened for development of anti-D antibodies for up to 12 months (at the predefined timepoints 2, 3, 6, and 12 months) after RhD+ red blood cell transfusion. 437 emergency patients, of whom 85 (20%) were RhD-, received 2836 RhD+ red blood cell concentrates. The overall risk of inducing anti-D antibodies (in all 437 recipients) was 17 (4%, 95% CI 2·44-6·14) of 437 (assuming all patients lost to follow-up developed anti-D allo-immunisation). During this period, 110 known RhD- patients received RhD+ red blood cell concentrates during RhD- red blood cell concentrate shortages. Of these, 29 (26%; 95% CI 19·0-35·3) developed anti-D allo-immunisation (assuming all

Red blood cell production

Science.gov (United States)

... bone marrow of bones. Stem cells in the red bone marrow called hemocytoblasts give rise to all of the formed elements in blood. If a hemocytoblast commits to becoming a cell called a proerythroblast, it will develop into a new red blood cell. The formation of a red blood ...

ANTI·RED CELL ACTIVITY OF LYMPHOCYTOTOXIC ANTIBODIES ...

African Journals Online (AJOL)

1971-05-08

May 8, 1971 ... it can be concluded that the Iymphocytes also share anti- gens with thymocytes .... antibodies, yet seem to lack the power to prolong graft survival, indicates ..... This limitation affects our knowledge of its in vivo activity when ...

Enzyme-linked immunosorbent assay (ELISA) using a specific monoclonal antibody as a new tool to detect Sudan dyes and Para red

International Nuclear Information System (INIS)

Ju Chunmei; Tang Yong; Fan Huiying; Chen Jinding

2008-01-01

To set up an immunoassay-based method to detect Sudan dyes and Para red, we generated a monoclonal antibody (Mab) using a specially designed carboxyl derivative of Sudan I (CSD I) as the immunogen. CSD I was synthesized by azocoupling reaction using 2-naphthol and diazotised 4-aminobenzoic acid. The antibody was obtained from a hybridoma, which was derived from the fusion of the mouse myeloma SP2/0 cells and the splenocytes from the mice immunized with the CSD I-bovine serum albumin (BSA) conjugate. In addition, we showed that the Mab was highly specific for Sudan I, III and Para red. The limit of detection was approximately 0.01 ng mL -1 in phosphate-buffered saline (PBS) buffer and 0.5 ng g -1 in chilli tomato sauce. The recoveries of Sudan I, III and Para red for the chilli tomato sauce were from 84% to 99% and coefficients of variation were from 14.9% to 33.3%. Thus, the enzyme-linked immunosorbent assay (ELISA) method is a rapid and high throughput screening tool to detect Sudan dyes and Para red in food products

Enzyme-linked immunosorbent assay (ELISA) using a specific monoclonal antibody as a new tool to detect Sudan dyes and Para red

Energy Technology Data Exchange (ETDEWEB)

Ju Chunmei [College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642 (China); Tang Yong [Center of Antibody Engineering, Department of Bioengineering, Jinan University, Guangzhou 510632 (China); Fan Huiying [College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642 (China); Chen Jinding [College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642 (China)], E-mail: jdchen@scau.edu.cn

2008-07-28

To set up an immunoassay-based method to detect Sudan dyes and Para red, we generated a monoclonal antibody (Mab) using a specially designed carboxyl derivative of Sudan I (CSD I) as the immunogen. CSD I was synthesized by azocoupling reaction using 2-naphthol and diazotised 4-aminobenzoic acid. The antibody was obtained from a hybridoma, which was derived from the fusion of the mouse myeloma SP2/0 cells and the splenocytes from the mice immunized with the CSD I-bovine serum albumin (BSA) conjugate. In addition, we showed that the Mab was highly specific for Sudan I, III and Para red. The limit of detection was approximately 0.01 ng mL{sup -1} in phosphate-buffered saline (PBS) buffer and 0.5 ng g{sup -1} in chilli tomato sauce. The recoveries of Sudan I, III and Para red for the chilli tomato sauce were from 84% to 99% and coefficients of variation were from 14.9% to 33.3%. Thus, the enzyme-linked immunosorbent assay (ELISA) method is a rapid and high throughput screening tool to detect Sudan dyes and Para red in food products.

Frequency and specificity of red blood cell alloimmunization in chilean transfused patients.

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Caamaño, José; Musante, Evangelina; Contreras, Margarita; Ulloa, Hernán; Reyes, Carolina; Inaipil, Verónica; Saavedra, Nicolás; Guzmán, Neftalí

2015-01-01

Alloimmunization is an adverse effect of blood transfusions. In Chile, alloimmunization frequency is not established, and for this reason the aim of this study was to investigate the prevalence and specificity of red blood cell (RBC) alloantibodies in Chilean transfused subjects. Records from 4,716 multi-transfused patients were analyzed. In these patients, antibody screening was carried out prior to cross-matching with a commercially available two-cell panel by the microcolum gel test, and samples with a positive screen were analyzed for the specificity of the alloantibody with a 16-cell identification panel. The incidence of RBC alloimmunization in transfused patients was 1.02% (48/4,716) with a higher prevalence in women (40/48). We detected 52 antibodies, the most frequent specificities identified were anti-E (30.8%), anti-K (26.9%), anti-D (7.7%), and anti-Fy(a) (5.8%). The highest incidence of alloantibodies was observed in cancer and gastroenterology patients. The data demonstrated a low alloimmunization frequency in Chilean transfused patients, principally associated with antibodies anti-E, anti-K, anti-D, and anti-Fy(a).

[Clinical case of the month. Mild hemolytic disease of the newborn due to an anti-Wr(a) antibody].

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Keutgens, A; Monfort, M; Wagemans, D; Van Cauwenberge, J-R; Gérard, C

2012-01-01

A Caucasian woman, with a A+ CCD.ee K neg erythrocyte phenotype and no history of blood transfusion, delivered a first child who developed mild anemia. The direct antiglobulin test performed on the newborn red blood cells belonging to the A+ CCD.ee K neg group, was strongly positive for IgG. During the pregnancy and after the delivery, the woman had a negative irregular antibody screening test, using standard red blood cells. However, at birth, using a collection of thawed red blood cells with rare phenotypes (private antigens), the lab showed an antibody anti-Wr(a) in the maternal serum. The activity of the maternal antibody, with a titer of 16, was completely inhibited by dithiothreitol, indicating the nature IgM of the circulating antibody. The presence of the antigen Wr(a) on the surface of the newborn and its biological father red blood cells was confirmed. The concentration of IgG anti-Wr(a) on baby erythrocytes was demonstrated by the presence of the antibody anti-Wr(a) in the eluate. This case illustrates the difficulties to detect antibodies against private antigens on baby erythrocytes, responsible of hemolytic diseases of newborn. Indeed, standard red blood cell panels used for irregular antibodies screening test do not express generally those private antigens.

Prevalence of antibodies against canine distemper virus among red foxes in Luxembourg

NARCIS (Netherlands)

B.C. Damien; S. Losch; J. Mossong; C.P. Muller (Claude); A.D.M.E. Osterhaus (Albert); B.E.E. Martina (Byron)

2002-01-01

textabstractCanine distemper virus (CDV) has a wide host spectrum, and during the past years, distemper has been observed in species that were previously not considered to be susceptible. In this study, we investigated the prevalence of CDV-specific antibodies in red foxes (Vulpes vulpes) sampled

Prevalence of antibodies against canine distemper virus among red foxes in Luxembourg.

NARCIS (Netherlands)

B.C. Damien; B.E.E. Martina (Byron); S. Losch; A.D.M.E. Osterhaus (Albert); C.P. Muller (Claude)

2002-01-01

textabstractCanine distemper virus (CDV) has a wide host spectrum, and during the past years, distemper has been observed in species that were previously not considered to be susceptible. In this study, we investigated the prevalence of CDV-specific antibodies in red foxes (Vulpes vulpes) sampled

21 CFR 640.10 - Red Blood Cells.

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2010-04-01

... 21 Food and Drugs 7 2010-04-01 2010-04-01 false Red Blood Cells. 640.10 Section 640.10 Food and... ADDITIONAL STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Red Blood Cells § 640.10 Red Blood Cells. The proper name of this product shall be Red Blood Cells. The product is defined as red blood cells remaining...

Frequency of red cell, leukocytic and platelet alloantibodies in patients with hematological diseases

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N. V. Mineeva

2014-07-01

Full Text Available History of multiple transfusions in patients with hematological diseases increases the likelihood of immunization to donor blood cells antigensand immunological complications development. Incidence of alloantibodies development in this patients was assessed in this work. Alloantibodies detection was performed in patients with aplastic anemia, acute leukemia, chronic lymphocytic leukemia, and autoimmune thrombocytopenia. 9696 patients were included in this study. Frequency of alloantibodies to red cell antigens was 3.8 %, with 0.9 % of the antibody belong to the immunoglobulin G, and 2.9 % of the cases – to immunoglobulin M. Most of the IgG antibodies had following specificity:monospecific anti-D antibody (21 cases, anti-DC and anti-DE antibodies (4 cases, anti-C (8 cases, anti-E (15, anti-c (13, and anti-K (11. Anti-e (1, anti-Fya (2, anti-Lea (4, anti-S (2, anti-s (2, anti-Jka (2 antibodies were less common. Granulocytes antibodies were found in 66.7 % of 384 patients, with results dependent on the detection method used. The presence of antiplatelet alloantibodies studied in 285 serum samples, of which antibodies were detected in 99 patients (34.7 %. Specificity of platelet antibodies was determined in three serum samples only: anti-2b, anti-1a, anti-1b. In other patients, probably present antibodies to several antigens simultaneously, and to identify them was not possible.

NMR water-proton spin-lattice relaxation time of human red blood cells and red blood cell suspensions

International Nuclear Information System (INIS)

Sullivan, S.G.; Rosenthal, J.S.; Winston, A.; Stern, A.

1988-01-01

NMR water-proton spin-lattice relaxation times were studied as probes of water structure in human red blood cells and red blood cell suspensions. Normal saline had a relaxation time of about 3000 ms while packed red blood cells had a relaxation time of about 500 ms. The relaxation time of a red blood cell suspension at 50% hematocrit was about 750 ms showing that surface charges and polar groups of the red cell membrane effectively structure extracellular water. Incubation of red cells in hypotonic saline increases relaxation time whereas hypertonic saline decreases relaxation time. Relaxation times varied independently of mean corpuscular volume and mean corpuscular hemoglobin concentration in a sample population. Studies with lysates and resealed membrane ghosts show that hemoglobin is very effective in lowering water-proton relaxation time whereas resealed membrane ghosts in the absence of hemoglobin are less effective than intact red cells. 9 refs.; 3 figs.; 1 table

Effect of screening for red cell antibodies, other than anti-D, to detect hemolytic disease of the fetus and newborn: a population study in the Netherlands.

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Koelewijn, J M; Vrijkotte, T G M; van der Schoot, C E; Bonsel, G J; de Haas, M

2008-05-01

Hemolytic disease of the fetus and newborn (HDFN) is a severe disease, resulting from maternal red cell (RBC) alloantibodies directed against fetal RBCs. The effect of a first-trimester antibody screening program on the timely detection of HDFN caused by antibodies other than anti-D was evaluated. Nationwide, all women (1,002 in 305,000 consecutive pregnancies during 18 months) with alloantibodies other than anti-D, detected by a first-trimester antibody screen, were included in a prospective index-cohort study. In a parallel-coverage validation study, patients with HDFN caused by antibodies other than anti-D, that were missed by the screening program, were retrospectively identified. The prevalence of positive antibody screens at first-trimester screening was 1,232 in 100,000; the prevalence of alloantibodies other than anti-D was 328 in 100,000, of which 191 of 100,000 implied a risk for occurrence of HDFN because the father carried the antigen. Overall, severe HDFN, requiring intrauterine or postnatal (exchange) transfusions, occurred in 3.7 percent of fetuses at risk: for anti-K in 11.6 percent; anti-c in 8.5 percent; anti-E in 1.1 percent; Rh antibodies other than anti-c, anti-D, or anti-E in 3.8 percent; and for antibodies other than Rh antibodies or anti-K, in none of the fetuses at risk. All affected children, where antibodies were detected, were promptly treated and healthy at the age of 1 year. The coverage validation study showed a sensitivity of the screening program of 75 percent. Five of 8 missed cases were caused by anti-c, with delay-induced permanent damage in at least 1. First-trimester screening enables timely treatment of HDFN caused by antibodies other than anti-D, however, with a sensitivity of only 75 percent. A second screening at Week 30 of c- women will enhance the screening program. Severe HDFN, caused by antibodies other than anti-D, is associated with anti-K, anti-c, and to a lesser extent with other Rh-alloantibodies.

Suppression of Red Blood Cell Autofluorescence for Immunocytochemistry on Fixed Embryonic Mouse Tissue.

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Whittington, Niteace C; Wray, Susan

2017-10-23

Autofluorescence is a problem that interferes with immunofluorescent staining and complicates data analysis. Throughout the mouse embryo, red blood cells naturally fluoresce across multiple wavelengths, spanning the emission and excitation spectra of many commonly used fluorescent reporters, including antibodies, dyes, stains, probes, and transgenic proteins, making it difficult to distinguish assay fluorescence from endogenous fluorescence. Several tissue treatment methods have been developed to bypass this issue with varying degrees of success. Sudan Black B dye has been commonly used to quench autofluorescence, but can also introduce background fluorescence. Here we present a protocol for an alternative called TrueBlack Lipofuscin Autofluorescence Quencher. The protocol described in this unit demonstrates how TrueBlack efficiently quenches red blood cell autofluorescence across red and green wavelengths in fixed embryonic tissue without interfering with immunofluorescent signal intensity or introducing background staining. We also identify optimal incubation, concentration, and multiple usage conditions for routine immunofluorescence microscopy. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

Engineering of red cells of Arabidopsis thaliana and comparative genome-wide gene expression analysis of red cells versus wild-type cells.

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Shi, Ming-Zhu; Xie, De-Yu

2011-04-01

We report metabolic engineering of Arabidopsis red cells and genome-wide gene expression analysis associated with anthocyanin biosynthesis and other metabolic pathways between red cells and wild-type (WT) cells. Red cells of A. thaliana were engineered for the first time from the leaves of production of anthocyanin pigment 1-Dominant (pap1-D). These red cells produced seven anthocyanin molecules including a new one that was characterized by LC-MS analysis. Wild-type cells established as a control did not produce anthocyanins. A genome-wide microarray analysis revealed that nearly 66 and 65% of genes in the genome were expressed in the red cells and wild-type cells, respectively. In comparison with the WT cells, 3.2% of expressed genes in the red cells were differentially expressed. The expression levels of 14 genes involved in the biosynthetic pathway of anthocyanin were significantly higher in the red cells than in the WT cells. Microarray and RT-PCR analyses demonstrated that the TTG1-GL3/TT8-PAP1 complex regulated the biosynthesis of anthocyanins. Furthermore, most of the genes with significant differential expression levels in the red cells versus the WT cells were characterized with diverse biochemical functions, many of which were mapped to different metabolic pathways (e.g., ribosomal protein biosynthesis, photosynthesis, glycolysis, glyoxylate metabolism, and plant secondary metabolisms) or organelles (e.g., chloroplast). We suggest that the difference in gene expression profiles between the two cell lines likely results from cell types, the overexpression of PAP1, and the high metabolic flux toward anthocyanins.

Red blood cell alloimmunization in transfused patients in sub-Saharan Africa: A systematic review and meta-analysis.

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Ngoma, Alain M; Mutombo, Paulin B; Ikeda, Kazuhiko; Nollet, Kenneth E; Natukunda, Bernard; Ohto, Hitoshi

2016-04-01

Previous studies of Sub-Saharan Africans show significant alloimmunization to red blood cell (RBC) antigens, but country-specific data are limited. Thus, the aim of this study was to estimate, by meta-analysis, the overall proportion of red blood cell alloantibodies among transfused patients. We systematically searched Medline, Embase, and the Africa-Wide Information database to identify relevant studies in any language. Case reports, comments, letters, conference abstracts, editorials, and review articles were excluded. Of the 269 potentially relevant articles, 11 studies fulfilled our selection criteria. Overall proportions of alloimmunization were 6.7 (95% CI: 5.7, 7.8) per 100 transfused patients. With regard to antibody specificity, among clinically significant antibodies, anti-E ranked as the most common, followed by anti-K, anti-C and anti-D. Meta-analysis of available literature quantifies and qualifies the clinical challenge of RBC alloimmunization among transfused patients in Sub-Saharan Africa. These results should drive policy decisions in favour of routine testing of RBC antigens and irregular antibodies for transfused patients as a standard of care throughout Sub-Saharan Africa. Copyright © 2015 Elsevier Ltd. All rights reserved.

Frequencies of maternal red blood cell alloantibodies in Port Harcourt, Nigeria

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Jeremiah Zaccheaus

2011-01-01

Full Text Available Background: Alloantibodies of clinical importance can cause transfusion reactions or hemolytic disease of the fetus and newborn (HDFN. The frequencies of these antibodies have not been reported in our locality. Aims: To determine the frequency of occurrence of alloantibodies among pregnant women in Port Harcourt, Nigeria. Settings and Design: This is a prospective study, which was carried out in the Braithwaite Memorial Specialist Hospital, Port Harcourt, Nigeria. Materials and Methods: Screening and identification of red blood cell alloantibodies was done on the sera of 500 pregnant women using the DiaMed, DiaCell, and DiaPanel reagents (Cressier, Switzerland. ABO and Rh blood groups were done using antisera bought from Biotec (Ipswich, UK. Results: Alloantibodies were identified in the serum of 17 of the 500 (3.4% pregnant women. The specificity of the antibodies was as follows: anti-C 6 (1.2%, anti-E 3 (0.6%, anti-Jsb 3 (0.6%, and anti-K 5 (1.0%. No anti-D was identified despite 8.6% of the study population being Rhesus D (Rh D negative. The distribution of the antibodies was found to be independent of the blood groups of the participants (x 2 = 4.050, P = 0.670. Blood group O constituted the highest percentage (48.0%. Conclusion: This study has identified the presence of non-Rh D antibodies to the proportion of 3.4%. Rh D antibody was absent in this population irrespective of the relatively high percentage of Rh D negative women. There is a need to determine the actual risk these antibodies may pose to the antenatal women and to include antibody screening and identification in routine antenatal care.

Antibody development in pediatric sickle cell patients undergoing erythrocytapheresis.

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Godfrey, Gwendolyn J; Lockwood, William; Kong, Maiying; Bertolone, Salvatore; Raj, Ashok

2010-12-01

Erythrocytapheresis, or red blood cell exchange transfusion (RBCX), with donor red blood cell (RBC) units is now increasingly used in the treatment of acute and chronic complications of sickle cell disease (SCD). As in all transfusions, RCBX carries a risk of immunization against foreign antigen on transfused cells. However, by selecting donor units with RBC phenotypes similar to the patient, the risk of allo- and autoimmunization can be reduced. The formation of RBC alloantibodies and/or autoantibodies in 32 multitransfused pediatric SCD patients undergoing monthly RBCX over a 11-year period (12/1998 to 12/2009) was evaluated utilizing a retrospective patient chart review at Kosair Children's Hospital, Louisville, Kentucky. After starting C, E, K antigen-matched RBCX, the rate of clinically significant allo-immunization decreased from 0.189/100 to 0.053/100 U, with a relative risk of 27.9%. Likewise, the rate of autoimmunization decreased from 0.063/100 to 0.035/100 U, with a relative risk of 55.9%. After controlling for clinically insignificant antibodies, our auto- and alloimmunization rate was much less than previously reported values. In addition, the incidence of clinically significant allo- and autoimmunization decreased in our patient population after starting minor antigen-matched RBCX. These results suggest that by matching selected RBC phenotypes, there may be an association in the risk of allo- and autoimmunization of multi-transfused SCD patients.

[Screening serum response special antibodies of U251 cell line from surface display phage antibody library].

Science.gov (United States)

Yu, Min; Tan, De-Yong; Qian, Wei; Lai, Jian-Hua; Sun, Gui-Lin

2004-05-01

U251 cell is a sensitive cell line to serum, which stops at G0 phase of cell cycle in no-serum medium, and recovers growth when the serum is added into no-serum medium. The cell can express corresponding proteins in different phase of cell cycle. Therefore it is very signification for the study of cell cycle regulation mechanism that explores these proteins. In this paper, the mouse antibody phage display library was added into the bottle in which the serum starvation U251 cells had been cultured, and the special antibody phages were absorbed. Then the absorbed antibody phages were amplified by adding E. coli TG1 and helper phage M13K07. Amplified antibody phages were added into bottle in which the serum cultured cell after serum starvation (follow named as serum recovered cells) were incubated, so that the cell absorbed the no-special antibody phages for the serum starvation cell and the special antibody phages were in supernatant. The remaining no-special antibody phages in the supernatant were discarded by repeating above program 3-4 times. The pure special antibody phages were gotten, and amplified by adding the host cell E. coli TG1 and helper phage M13K07. Then the host bacterium infected special antibody phage was spread on the plate medium with ampicillin, and the monoclonal antibody phages were gotten. Using same as above program, the monoclonal antibody phages absorbed specially for serum recovered U251 cells were obtained when the serum recovered cells instead of serum starvation cells and serum starvation cells instead of serum recovered cells. In this study, ninety-six positive monoclonal antibody phages that absorbed specially the serum starvation cells and eighty-two positive monoclonal antibody phages that absorbed specially the serum recovered cells were obtained. By using cell immunochemistry assay, two special signification antibodies were obtained. one (No.11) was the strong response in serum starvation cells, the other (No.2) was the strong

Efficient generation of monoclonal antibodies from single rhesus macaque antibody secreting cells.

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Meng, Weixu; Li, Leike; Xiong, Wei; Fan, Xuejun; Deng, Hui; Bett, Andrew J; Chen, Zhifeng; Tang, Aimin; Cox, Kara S; Joyce, Joseph G; Freed, Daniel C; Thoryk, Elizabeth; Fu, Tong-Ming; Casimiro, Danilo R; Zhang, Ningyan; A Vora, Kalpit; An, Zhiqiang

2015-01-01

Nonhuman primates (NHPs) are used as a preclinical model for vaccine development, and the antibody profiles to experimental vaccines in NHPs can provide critical information for both vaccine design and translation to clinical efficacy. However, an efficient protocol for generating monoclonal antibodies from single antibody secreting cells of NHPs is currently lacking. In this study we established a robust protocol for cloning immunoglobulin (IG) variable domain genes from single rhesus macaque (Macaca mulatta) antibody secreting cells. A sorting strategy was developed using a panel of molecular markers (CD3, CD19, CD20, surface IgG, intracellular IgG, CD27, Ki67 and CD38) to identify the kinetics of B cell response after vaccination. Specific primers for the rhesus macaque IG genes were designed and validated using cDNA isolated from macaque peripheral blood mononuclear cells. Cloning efficiency was averaged at 90% for variable heavy (VH) and light (VL) domains, and 78.5% of the clones (n = 335) were matched VH and VL pairs. Sequence analysis revealed that diverse IGHV subgroups (for VH) and IGKV and IGLV subgroups (for VL) were represented in the cloned antibodies. The protocol was tested in a study using an experimental dengue vaccine candidate. About 26.6% of the monoclonal antibodies cloned from the vaccinated rhesus macaques react with the dengue vaccine antigens. These results validate the protocol for cloning monoclonal antibodies in response to vaccination from single macaque antibody secreting cells, which have general applicability for determining monoclonal antibody profiles in response to other immunogens or vaccine studies of interest in NHPs.

A Case of Hemolytic Disease of the Newborn due to Dia Antibody

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Jethava, Ashif; Olivares, Esperanza; Shariatmadar, Sherry

2015-01-01

Anti-Dia is a clinically significant red cell antibody known to cause hemolytic disease of the newborn. Here, we report on a case of mild hemolytic disease of the newborn caused by Dia antibody. The mother had three prior pregnancies with no history of blood transfusion. She delivered a preterm 35-week-old female newborn by cesarean section. The neonate developed anemia and mild icterus on postnatal day five with hemoglobin of 9500 mg/dL and total bilirubin of 10 mg/dL. The direct antiglobulin test on the neonate's red blood cells was positive. The maternal serum and an eluate from the infant RBCs were negative in routine antibody detection tests but were positive using commercially prepared Di(a+) red cells. The neonate was discharged home in stable condition following treatment with erythropoietin and phototherapy. When a newborn has a positive DAT in the absence of major blood group incompatibility or commonly detected RBC antibodies, an antibody to a low frequency antigen such as Dia must be considered. Further immunohematology tests are required to determine presence of the antibody and the clinician must be alerted to closely monitor the infant for signs of anemia and hemolysis. PMID:26682081

A Case of Hemolytic Disease of the Newborn due to Dia Antibody

Directory of Open Access Journals (Sweden)

Ashif Jethava

2015-01-01

Full Text Available Anti-Dia is a clinically significant red cell antibody known to cause hemolytic disease of the newborn. Here, we report on a case of mild hemolytic disease of the newborn caused by Dia antibody. The mother had three prior pregnancies with no history of blood transfusion. She delivered a preterm 35-week-old female newborn by cesarean section. The neonate developed anemia and mild icterus on postnatal day five with hemoglobin of 9500 mg/dL and total bilirubin of 10 mg/dL. The direct antiglobulin test on the neonate’s red blood cells was positive. The maternal serum and an eluate from the infant RBCs were negative in routine antibody detection tests but were positive using commercially prepared Di(a+ red cells. The neonate was discharged home in stable condition following treatment with erythropoietin and phototherapy. When a newborn has a positive DAT in the absence of major blood group incompatibility or commonly detected RBC antibodies, an antibody to a low frequency antigen such as Dia must be considered. Further immunohematology tests are required to determine presence of the antibody and the clinician must be alerted to closely monitor the infant for signs of anemia and hemolysis.

Clinical significance of determination of changes of serum SOD and T-cell subsets distribution type after leukocyte-deduced red blood cell transfusion in patients with lung cancer

International Nuclear Information System (INIS)

Yu Zhengqin; Li Keqin; Xiang Hengquan

2006-01-01

Objective: To investigate the changes of serum SOD contents and T-cell subsets distribution type after leukocyte-deduced red blood cell transfusion in patients with lung cancer. Methods: Serum SOD levels was measured with RIA and T-cell subsets distribution type was detected with monoclonal antibody technic both before and after leukocyte-deduced red blood cell transfusion in 32 patients with lung cancer and 35 normal controls. Results: Before treatment, the serum levels of SOD and T-cell CIM/ CD8 value were significantly lower in the patients than those in controls (P 0.05). Conclusion: Determination of serum SOD level and T-cell subsets distribution type is clinically useful in the management of patients with lung cancer. (authors)

Uptake of carnitine by red blood cells

International Nuclear Information System (INIS)

Campa, M.; Borum, P.

1986-01-01

A significant amount of blood carnitine (70% of cord blood and 40% of blood from healthy adults) is partitioned into the red blood cell compartment of whole blood. Data indicate that the plasma compartment and the red blood cell compartment of whole blood represent different metabolic pools of carnitine. There are no data to indicate that red blood cells synthesize carnitine, but our understanding of the uptake of carnitine by red blood cells is negligible. Red blood cells were obtained from healthy adults, washed twice with normal saline, and used for uptake experiments. When the cells were incubated at 37 0 C in the presence of 14 C-carnitine, radioactivity was found both in the soluble cytosolic and membrane fractions of the cells following lysis. The uptake was dependent upon the time of incubation, temperature of incubation, and carnitine concentration in the incubation medium. Washed red blood cell membranes incubated with 14 C-carnitine showed specific binding of radioactivity. These data are consistent with the hypothesis that red blood cells have an uptake mechanism for L-carnitine

Shape-Shifted Red Blood Cells: A Novel Red Blood Cell Stage?

Science.gov (United States)

Chico, Verónica; Puente-Marin, Sara; Nombela, Iván; Ciordia, Sergio; Mena, María Carmen; Carracedo, Begoña; Villena, Alberto; Mercado, Luis; Coll, Julio; Ortega-Villaizan, María Del Mar

2018-04-19

Primitive nucleated erythroid cells in the bloodstream have long been suggested to be more similar to nucleated red cells of fish, amphibians, and birds than the red cells of fetal and adult mammals. Rainbow trout Ficoll-purified red blood cells (RBCs) cultured in vitro undergo morphological changes, especially when exposed to stress, and enter a new cell stage that we have coined shape-shifted RBCs (shRBCs). We have characterized these shRBCs using transmission electron microscopy (TEM) micrographs, Wright⁻Giemsa staining, cell marker immunostaining, and transcriptomic and proteomic evaluation. shRBCs showed reduced density of the cytoplasm, hemoglobin loss, decondensed chromatin in the nucleus, and striking expression of the B lymphocyte molecular marker IgM. In addition, shRBCs shared some features of mammalian primitive pyrenocytes (extruded nucleus surrounded by a thin rim of cytoplasm and phosphatidylserine (PS) exposure on cell surface). These shRBCs were transiently observed in heat-stressed rainbow trout bloodstream for three days. Functional network analysis of combined transcriptomic and proteomic studies resulted in the identification of proteins involved in pathways related to the regulation of cell morphogenesis involved in differentiation, cellular response to stress, and immune system process. In addition, shRBCs increased interleukin 8 (IL8), interleukin 1 β (IL1β), interferon ɣ (IFNɣ), and natural killer enhancing factor (NKEF) protein production in response to viral hemorrhagic septicemia virus (VHSV). In conclusion, shRBCs may represent a novel cell stage that participates in roles related to immune response mediation, homeostasis, and the differentiation and development of blood cells.

Shape-Shifted Red Blood Cells: A Novel Red Blood Cell Stage?

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Chico, Verónica; Puente-Marin, Sara; Ciordia, Sergio; Mena, María Carmen; Carracedo, Begoña; Mercado, Luis; Coll, Julio

2018-01-01

Primitive nucleated erythroid cells in the bloodstream have long been suggested to be more similar to nucleated red cells of fish, amphibians, and birds than the red cells of fetal and adult mammals. Rainbow trout Ficoll-purified red blood cells (RBCs) cultured in vitro undergo morphological changes, especially when exposed to stress, and enter a new cell stage that we have coined shape-shifted RBCs (shRBCs). We have characterized these shRBCs using transmission electron microscopy (TEM) micrographs, Wright–Giemsa staining, cell marker immunostaining, and transcriptomic and proteomic evaluation. shRBCs showed reduced density of the cytoplasm, hemoglobin loss, decondensed chromatin in the nucleus, and striking expression of the B lymphocyte molecular marker IgM. In addition, shRBCs shared some features of mammalian primitive pyrenocytes (extruded nucleus surrounded by a thin rim of cytoplasm and phosphatidylserine (PS) exposure on cell surface). These shRBCs were transiently observed in heat-stressed rainbow trout bloodstream for three days. Functional network analysis of combined transcriptomic and proteomic studies resulted in the identification of proteins involved in pathways related to the regulation of cell morphogenesis involved in differentiation, cellular response to stress, and immune system process. In addition, shRBCs increased interleukin 8 (IL8), interleukin 1 β (IL1β), interferon ɣ (IFNɣ), and natural killer enhancing factor (NKEF) protein production in response to viral hemorrhagic septicemia virus (VHSV). In conclusion, shRBCs may represent a novel cell stage that participates in roles related to immune response mediation, homeostasis, and the differentiation and development of blood cells. PMID:29671811

Long-term in vivo survival of Rh(D)-negative donor red cells in a patient with anti-LW

International Nuclear Information System (INIS)

Chaplin, H.; Hunter, V.L.; Rosche, M.E.; Shirey, R.S.

1985-01-01

The present study documents immediate and long-term survival of crossmatch-incompatible Rh(D)-negative donor red cells in a patient with anti-LW. A 67-year-old group A Rh(D)-positive man was admitted for urgent coronary artery bypass surgery. The direct antiglobulin test (DAT) was weakly positive in two of five laboratories. His serum contained anti-LW (two laboratories); his red cells were LW negative (three antisera). Two siblings were LW-positive. Surgery was delayed, and 3 ml Rh(D)-negative crossmatch-incompatible red cells stored in citrate-phosphate-dextrose-adenine-one were labeled with 25 microCi of 51 Cr and injected. Immediate survival was approximately 100 percent with 92 percent survival at 20 hours. Six daily blood samples showed a decreased red cell lifespan, (T 1/2 . 14 days). Because of medical complications, 4 units of Rh(D)-negative crossmatch-incompatible blood were then transfused without clinical or hemolytic reaction. The anti-IgG DAT became stronger. In vivo survival of the remaining 51 Cr-RBCs became normal (T 1/2 28 days over the succeeding 20 days). Following transfusion, no change in serum antibody strength was demonstrated by double-blind titration of seven coded samples. The observations support modest reduction of lifespan for 3 ml of LW-positive red cells, but normal survival following subsequent transfusion of approximately 700 ml of LW-positive red cells

A Case of Hemolytic Disease of the Newborn due to Di (a) Antibody.

Science.gov (United States)

Jethava, Ashif; Olivares, Esperanza; Shariatmadar, Sherry

2015-01-01

Anti-Di(a) is a clinically significant red cell antibody known to cause hemolytic disease of the newborn. Here, we report on a case of mild hemolytic disease of the newborn caused by Di(a) antibody. The mother had three prior pregnancies with no history of blood transfusion. She delivered a preterm 35-week-old female newborn by cesarean section. The neonate developed anemia and mild icterus on postnatal day five with hemoglobin of 9500 mg/dL and total bilirubin of 10 mg/dL. The direct antiglobulin test on the neonate's red blood cells was positive. The maternal serum and an eluate from the infant RBCs were negative in routine antibody detection tests but were positive using commercially prepared Di(a+) red cells. The neonate was discharged home in stable condition following treatment with erythropoietin and phototherapy. When a newborn has a positive DAT in the absence of major blood group incompatibility or commonly detected RBC antibodies, an antibody to a low frequency antigen such as Di(a) must be considered. Further immunohematology tests are required to determine presence of the antibody and the clinician must be alerted to closely monitor the infant for signs of anemia and hemolysis.

Decreased antibody formation in mice exposed to lead

Energy Technology Data Exchange (ETDEWEB)

Koller, L D; Kovacic, S

1974-07-12

Swiss Webster mice were given 1375, 137.5, or 13.75 ppM lead acetate in deionized water for 56 days. The control group was given deionized water orally. There were 120 mice in each group. The diet fed to all the mice was contaminated with 1.12 ppM lead. After 56 days, all mice were inoculated intraperitoneally with 0.2 ml of a 2% suspension of sheep red blood cells. Ten mice in each group were killed on days 3 to 7 to measure primary immune response (19S or IgM antibody) and on days 9 to 14 for the secondary response (7S or IgG antibody) after a second inoculation of sheep red blood cells while they remained on 137.5 ppM lead. The number of plaque forming cells was measured in the spleen. Erythrocytes were observed for basophilic stippling, packed cell volume was measured, serum was collected for hemolysin titration, and kidneys were examined for lead. Chronic exposure to lead produced a significant decrease in antibody synthesis, particularly IgG, indicating that the memory cell was involved. The results also indicated that the reduced antibody synthesis was responsible for the increased mortality from bacterial and viral diseases in animals that were chronically exposed to lead. Other environmental contaminants such as polychlorinated biphenyls, cadmium, mercury, DDT, and sulfur dioxide have also resulted in reduction of circulating antibodies in animals, in other experiments.

21 CFR 864.9175 - Automated blood grouping and antibody test system.

Science.gov (United States)

2010-04-01

...) Identification. An automated blood grouping and antibody test system is a device used to group erythrocytes (red blood cells) and to detect antibodies to blood group antigens. (b) Classification. Class II (performance... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Automated blood grouping and antibody test system...

Lymphocyte-dependent antibody-mediated cytotoxicity in Hashimoto thyroiditis

Science.gov (United States)

Calder, Elizabeth A.; Penhale, W. J.; McLeman, Dena; Barnes, E. W.; Irvine, W. J.

1973-01-01

In the presence of normal human lymphocytes, decomplemented sera from twentynine out of thirty-nine patients with Hashimoto thyroiditis caused significant lysis of thyroglobulin-coated chicken red blood cells, as estimated by the release of 51Cr; the mean% specific 51Cr release being 14·1 ± 1·9 (SEM). Serum from twenty-one control subjects studied concurrently caused no significant lysis of thyroglobulin-coated chicken red blood cells; the mean% specific 51Cr release being −1·6±0·7 (SEM). The degree of cytotoxicity correlated with the titre of thyroglobulin antibodies in the serum, determined by tanned red cell haemagglutination. The active component in the Hashimoto serum was localized in the 19S fraction, was unaffected by pre-absorption with anti-human IgM serum, but was neutralized by pre-absorption with anti-human IgG serum. These findings suggest that the cytotoxic activity of serum from patients with Hashimoto thyroiditis is due to the presence of thyroglobulin antibody of the IgG class in the form of complexes, either alone or with antigen. It is postulated that non-specific lymphocytes may play an important role in the pathogenesis of Hashimoto thyroiditis, being activated by the presence in the gland of thyroglobulin antibody, either alone or in the form of complexes attached to thyroid cells. PMID:4740445

Red blood cell alloimmunization among sickle cell Kuwaiti Arab patients who received red blood cell transfusion.

Science.gov (United States)

Ameen, Reem; Al Shemmari, Salem; Al-Bashir, Abdulaziz

2009-08-01

Sickle cell disease (SCD) is common in the Arabian Gulf region. Most cases require a red blood cell (RBC) transfusion, increasing the potential for RBC alloantibody development. The incidence of RBC alloimmunization among Kuwaiti Arab SCD patients is not yet known. This study retrospectively assessed the effect of using two different matching protocols on the incidence of alloimmunization among multiply transfused Kuwaiti Arab SCD patients. A total of 233 Kuwaiti Arab SCD patients were divided into two groups: Group 1 (n = 110) received RBC transfusion through standard ABO- and D-matched nonleukoreduced blood; Group 2 (n = 123) received RBCs matched for ABO, Rh, and K1 poststorage-leukoreduced blood. Multivariate analysis was performed on the factors associated with RBC alloimmunization and antibody specificity. Sixty-five percent of patients in Group 1 developed clinically significant RBC alloantibody with an increased prevalence in females; in patients in Group 2, 23.6% developed RBC alloantibodies (p = 0.01). In Group 1, 72 patients (65.5%) had alloantibodies directed against Rh and Kell systems (p = 0.01). Multivariate analysis further confirmed the results, showing that blood transfusion type and sex have significant effects on the rate of alloimmunizations. This study confirms the importance of selecting RBCs matched for Rh and Kell to reduce the risk of alloimmunizations among Kuwaiti Arab SCD patients.

Monoclonal antibodies reactive with hairy cell leukemia

NARCIS (Netherlands)

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

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.

In vivo red blood cell compatibility testing using indium-113m tropolone-labeled red blood cells

International Nuclear Information System (INIS)

Morrissey, G.J.; Gravelle, D.; Dietz, G.; Driedger, A.A.; King, M.; Cradduck, T.D.

1988-01-01

In vivo radionuclide crossmatch is a method for identifying compatible blood for transfusion when allo- or autoantibodies preclude the use of conventional crossmatching techniques. A technique for labeling small volumes of donor red blood cells with [/sup 113m/In]tropolone is reported. The use of /sup 113m/In minimizes the accumulation of background radioactivity and the radiation dose especially so when multiple crossmatches are performed. Labeling red cells with [/sup 113m/In]tropolone is faster and easier to perform than with other radionuclides. Consistently high labeling efficiencies are obtained and minimal /sup 113m/In activity elutes from the labeled red blood cells. A case study involving 22 crossmatches is presented to demonstrate the technique. The radiation dose equivalent from /sup 113m/In is significantly less than with other radionuclides that may be used to label red cells

Application of commercial enzyme linked immunosorbent assays (ELISA for the detection of antibodies for foot-and-mouth disease virus in wild boar and red deer

Directory of Open Access Journals (Sweden)

Terzić Svjetlana

2012-01-01

Full Text Available For detecting antibodies towards foot and mouth (FMD virus in sera collected from red deer hinds (Cervus elaphus and wild boars (Sus scrofa, three commercially available enzyme-linked immunosorbent assays (ELISA were used. Two ELISA kits (PrioCHECK FMDV NS and CHEKIT FMD-3ABC were used for the detection of antibodies towards non-structural proteins of FMD virus and one assay was based on the detection of antibodies for serotype O (PrioCHECK FMDV type O. All of the sera tested in our study were negative for antibodies against FMD virus. The aim of this study was to investigate the usefulness of commercially available ELISA kits given for marketing authorization in Croatia in testing the prevalence of FMD antibodies in wild boar and red deer populations. Since the producers of ELISA kits used in our study did not declare wild animals as a target species, we hypothesised that the same kits could be used for serological diagnosis of FMD in red deer and wild boars. Our study confirmed that the kits used are acceptable for detecting antibodies in both species tested, however, the investigation highlighted the problem of validating the kits due to the absence of available positive sera originating from red deer, as well as other susceptible species, especially artiodactyls.

Growth and replication of red rain cells at 121°C and their red fluorescence

Science.gov (United States)

Gangappa, Rajkumar; Wickramasinghe, Chandra; Wainwright, Milton; Kumar, A. Santhosh; Louis, Godfrey

2010-09-01

We have shown that the red cells found in the Red Rain (which fell on Kerala, India, in 2001) survive and grow after incubation for periods of up to two hours at 121°C . Under these conditions daughter cells appear within the original mother cells and the number of cells in the samples increases with length of exposure to 121°C. No such increase in cells occurs at room temperature, suggesting that the increase in daughter cells is brought about by exposure of the Red Rain cells to high temperatures. This is an independent confirmation of results reported earlier by two of the present authors, claiming that the cells can replicate under high pressure at temperatures upto 300°C. The flourescence behaviour of the red cells is shown to be in remarkable correspondence with the extended red emission observed in the Red Rectagle planetary nebula and other galactic and extragalactic dust clouds, suggesting, though not proving an extraterrestrial origin.

Affinity of antibody secreted by a single cell

International Nuclear Information System (INIS)

Doran, D.M.

1978-01-01

It was the intention of this research to measure the affinity of antibody secreted by a single cell, and to describe the spectrum of affinities displayed in response to antigenic stimulation. The single cell secreting specific antibody was isolated by means of the hemolytic plaque assay. The amount of antibody secreted by the cell was to be measured through the use of a solid phase radioimmunoassay. The affinity of the antibody would be estimated by comparing the diameter of the plaque, and the amount of antibody secreted, with a mathematical theory of the formation of a plaque in agar. As a test system, a solid phase radioimmunoassay was developed for human serum albumin using antibody coupled to Sephadex. A sensitivity of 1 nanogram was attained with this assay. A solid phase radioimmunoassay for mouse immunoglobulin M was developed, using antibody coupled to Sepharose. The sensitivity attained with this assay was only on the order of 10 micrograms. The mouse immunoglobulin M radioimmunoassay was not sensitive enough to measure the amount of antibody secreted by a single cell. From a theoretical equation, the relationship between antibody affinity, plaque diameter and antibody secretion rate was calculated for the experimental conditions used in this research. By assuming a constant antibody secretion rate, an effective binding constant for the antibody was estimated from the average plaque diameters. This effective binding constant was observed to increase during the immune response

Antibody titers in animal bite victims after post exposure vaccination with intradermally administered purified vero cell rabies vaccine using modified thai red cross regimen

International Nuclear Information System (INIS)

Hafeez, S.; Tahir, Z.

2014-01-01

To determine the seroconversion following rabies vaccination by intradermal route in cases of animal bite attending Anti rabies center, Lahore for post exposure prophylaxis. Study Design: Cross sectional descriptive study. Place and Duration: Antirabies center, Birdwood road Lahore, Microbiology laboratory, office of Bacteriologist, Government of Punjab, Lahore. Patients and Methods: Victims of all ages and both sexes having exposure with suspected rabid animal within 24 - 72 hours were included, fulfilling inclusion and exclusion criteria, over 3 months period from February to April 20. Patients of Category II and III wounds were included. Purified vero cell vaccine (PVR V) with antigenic content> 2.5 ml was used for intradermal vaccination according to modified Thai Red Cross regimen (2-2-2-0-2). Each victim received 0.1 ml intradermal dose on each deltoid on day 0, 3, 7 and 28th day of bite. Blood samples from victims were taken on day 0, 14 and 35. Antibody titers were estimated by ELISA kit. Results: Fifty cases were studied including 20 children. Male female ratio was 4:1. Optimum serocon version (> 0.5 IU/ml) was achieved in all cases by day 14. Antibody levels increased further (> 4 IV/ml) in 92% cases on day 35. Geometric mean titers were 3.2 IU/ml and 6.2 IU/ml on day 14 and 35 respectively. Conclusion: Intradermal route for cell culture rabies vaccine for postexposure prophylaxis in animal bite victims was efficacious and safe. The smaller dosage of vaccine was economically affordable by patients in referral centers. (author)

[A case of severe hemolytic disease of the newborn due to anti-Dia antibody].

Science.gov (United States)

Lee, Sun Min; Im, Sun Ju; Park, Su Eun; Lee, Eun Yup; Kim, Hyung Hoi

2007-10-01

Here we report a severe case of hemolytic anemia of the newborn with kernicterus caused by anti-Di(a) antibody. A full term male infant was transferred due to hyperbilirubinemia on the third day of life. Despite single phototherapy, the baby's total bilirubin had elevated to 30.1 mg/dL. After exchange transfusion, total bilirubin decreased to 11.45 mg/dL. The direct antiglobulin test on the infant's red cells was positive. The maternal and infant's sera showed a negative reaction in routine antibody detection tests, but were positive in Di(a) panel cells. The frequency of the Di(a) antigen among the Korean population is estimated to be 6.4-14.5%. Anti-Di(a) antibody could cause a hemolytic reaction against transfusion or hemolytic disease of the newborn. We suggest the need for reagent red blood cell panels to include Di(a) antigen positive cells in antibody identification test for Korean.

Abnormal red cell structure and function in neuroacanthocytosis.

Directory of Open Access Journals (Sweden)

Judith C A Cluitmans

Full Text Available Panthothenate kinase-associated neurodegeneration (PKAN belongs to a group of hereditary neurodegenerative disorders known as neuroacanthocytosis (NA. This genetically heterogeneous group of diseases is characterized by degeneration of neurons in the basal ganglia and by the presence of deformed red blood cells with thorny protrusions, acanthocytes, in the circulation.The goal of our study is to elucidate the molecular mechanisms underlying this aberrant red cell morphology and the corresponding functional consequences. This could shed light on the etiology of the neurodegeneration.We performed a qualitative and semi-quantitative morphological, immunofluorescent, biochemical and functional analysis of the red cells of several patients with PKAN and, for the first time, of the red cells of their family members.We show that the blood of patients with PKAN contains not only variable numbers of acanthocytes, but also a wide range of other misshapen red cells. Immunofluorescent and immunoblot analyses suggest an altered membrane organization, rather than quantitative changes in protein expression. Strikingly, these changes are not limited to the red blood cells of PKAN patients, but are also present in the red cells of heterozygous carriers without neurological problems. Furthermore, changes are not only present in acanthocytes, but also in other red cells, including discocytes. The patients' cells, however, are more fragile, as observed in a spleen-mimicking device.These morphological, molecular and functional characteristics of red cells in patients with PKAN and their family members offer new tools for diagnosis and present a window into the pathophysiology of neuroacanthocytosis.

Clinical evaluation of a 51Cr-labeled red blood cell survival test for in vivo blood compatibility testing

International Nuclear Information System (INIS)

Pineda, A.A.; Dharkar, D.D.; Wahner, H.W.

1984-01-01

Modified red blood cell survival studies with use of 51Cr were performed in three groups of subjects. Group 1 consisted of normal subjects who were given labeled autologous blood, group 2 were subjects in need of blood transfusions and given labeled ABO and Rh crossmatch-compatible blood, and group 3 were patients in need of blood transfusion but in whom problems arose in finding compatible blood. The results of the studies suggest that for patients with blood compatibility problems, normal red blood cell survival values at 1 hour do not exclude the possibility of severe hemolysis 24 hours later. Thus, if a 1-hour test result is normal, the procedure should be extended routinely to 24 hours. Moreover, the test can be used to evaluate the clinical importance of antibodies. We showed that anti-Yka and anti-Lan were clinically significant, but high-titer, low-avidity antibodies, anti-Kna, anti-I, and anti-HI were clinically insignificant in the cases studied. This finding emphasizes the importance of an in vivo test for the final compatibility evaluation in complicated blood replacement problems

Comparison of micro column technology with conventional tube methods for antibody detection

Directory of Open Access Journals (Sweden)

Sachin Garg

2017-01-01

Conclusion: MCT was found to be most efficacious when compared to CTT and tube LISS-IAT in detecting clinically significant red cell antibodies; although MCT missed 2 cases of Lea antibody which were detected by CTT and LISS-IAT.

RISK OF RED BLOOD CELL ALLOIMMUNISATION IN RWANDA ...

African Journals Online (AJOL)

2013-04-04

Apr 4, 2013 ... spin crossmatch (IS-XM) or indirect antiglobulin test crossmatch (IAT-XM). Objectives: ... with an odds ratio of 4.8; [95% CI=1.2-19.8]; and a p-value of 0.031. Conclusion: ... formation of single or multiple antibodies to red blood.

The antibody approach of labeling blood cells

International Nuclear Information System (INIS)

Srivastava, S.C.

1992-01-01

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 critically assessed and evaluated

The antibody approach of labeling blood cells

International Nuclear Information System (INIS)

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

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.

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.

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.

An enzyme-linked immunoabsorbent assay for estimating red cell survival of transfused red cells-validation using CR-51 labeling

International Nuclear Information System (INIS)

Drew, H.; Kickler, T.; Smith, B.; LaFrance, N.

1984-01-01

The survival time of transfused red cells antigenically distinct from the recipient's red cells was determined using an indirect enzyme linked antiglobulin test. These results were then compared to those determined by Cr-51 labeling. Three patients with hypoproliferative anemias and one patient (2 studies) with traumatic hemolytic anemia caused by a prosthetic heart valve were studied. Survival times were performed by transfusing a 5cc aliquot of Cr-51 labeled cells along with the remaining unit. One hour post transfusion, a blood sample was drawn and used as the 100% value. Subsequent samples drawn over a 2-3 week period were then compared to the initial sample to determine percent survival for both methods. The ELISA method for measuring red cell survival in antigenically distinct cells is in close agreement with the Cr-51 method. Although CR-51 labeling is the accepted method for red cell survival determination the ELISA method can be used when radioisotopes are unavailable or contraindicated or when the decision to estimate red cell survival is made after transfusion

Neutralisation of HIV-1 cell-cell spread by human and llama antibodies.

Science.gov (United States)

McCoy, Laura E; Groppelli, Elisabetta; Blanchetot, Christophe; de Haard, Hans; Verrips, Theo; Rutten, Lucy; Weiss, Robin A; Jolly, Clare

2014-10-02

Direct cell-cell spread of HIV-1 is a very efficient mode of viral dissemination, with increasing evidence suggesting that it may pose a considerable challenge to controlling viral replication in vivo. Much current vaccine research involves the study of broadly neutralising antibodies (bNabs) that arise during natural infection with the aims of eliciting such antibodies by vaccination or incorporating them into novel therapeutics. However, whether cell-cell spread of HIV-1 can be effectively targeted by bNabs remains unclear, and there is much interest in identifying antibodies capable of efficiently neutralising virus transmitted by cell-cell contact. In this study we have tested a panel of bNAbs for inhibition of cell-cell spread, including some not previously evaluated for inhibition of this mode of HIV-1 transmission. We found that three CD4 binding site antibodies, one from an immunised llama (J3) and two isolated from HIV-1-positive patients (VRC01 and HJ16) neutralised cell-cell spread between T cells, while antibodies specific for glycan moieties (2G12, PG9, PG16) and the MPER (2F5) displayed variable efficacy. Notably, while J3 displayed a high level of potency during cell-cell spread we found that the small size of the llama heavy chain-only variable region (VHH) J3 is not required for efficient neutralisation since recombinant J3 containing a full-length human heavy chain Fc domain was significantly more potent. J3 and J3-Fc also neutralised cell-cell spread of HIV-1 from primary macrophages to CD4+ T cells. In conclusion, while bNabs display variable efficacy at preventing cell-cell spread of HIV-1, we find that some CD4 binding site antibodies can inhibit this mode of HIV-1 dissemination and identify the recently described llama antibody J3 as a particularly potent inhibitor. Effective neutralisation of cell-cell spread between physiologically relevant cell types by J3 and J3-Fc supports the development of VHH J3 nanobodies for therapeutic or

Cell-free synthesis of functional antibody fragments to provide a structural basis for antibody-antigen interaction.

Directory of Open Access Journals (Sweden)

Takayoshi Matsuda

Full Text Available Growing numbers of therapeutic antibodies offer excellent treatment strategies for many diseases. Elucidation of the interaction between a potential therapeutic antibody and its target protein by structural analysis reveals the mechanism of action and offers useful information for developing rational antibody designs for improved affinity. Here, we developed a rapid, high-yield cell-free system using dialysis mode to synthesize antibody fragments for the structural analysis of antibody-antigen complexes. Optimal synthesis conditions of fragments (Fv and Fab of the anti-EGFR antibody 059-152 were rapidly determined in a day by using a 30-μl-scale unit. The concentration of supplemented disulfide isomerase, DsbC, was critical to obtaining soluble antibody fragments. The optimal conditions were directly applicable to a 9-ml-scale reaction, with linear scalable yields of more than 1 mg/ml. Analyses of purified 059-152-Fv and Fab showed that the cell-free synthesized antibody fragments were disulfide-bridged, with antigen binding activity comparable to that of clinical antibodies. Examination of the crystal structure of cell-free synthesized 059-152-Fv in complex with the extracellular domain of human EGFR revealed that the epitope of 059-152-Fv broadly covers the EGF binding surface on domain III, including residues that formed critical hydrogen bonds with EGF (Asp355EGFR, Gln384EGFR, H409EGFR, and Lys465EGFR, so that the antibody inhibited EGFR activation. We further demonstrated the application of the cell-free system to site-specific integration of non-natural amino acids for antibody engineering, which would expand the availability of therapeutic antibodies based on structural information and rational design. This cell-free system could be an ideal antibody-fragment production platform for functional and structural analysis of potential therapeutic antibodies and for engineered antibody development.

Antibody dependent cellular phagocytosis (ADCP) and antibody dependent cellular cytotoxicity (ADCC) of breast cancer cells mediated by bispecific antibody, MDX-210.

Science.gov (United States)

Watanabe, M; Wallace, P K; Keler, T; Deo, Y M; Akewanlop, C; Hayes, D F

1999-02-01

MDX-210 is a bispecific antibody (BsAb) with specificity for both the proto-oncogene product of HER-2/neu (c-erbB-2) and FcgammaRI (CD64). HER-2/neu is overexpressed in malignant tissue of approximately 30% of patients with breast cancer, and FcgammaRI is expressed on human monocytes, macrophages, and IFN-gamma activated granulocytes. We investigated phagocytosis and cytolysis of cultured human breast cancer cells by human monocyte-derived macrophages (MDM) mediated by BsAb MDX-210, its partially humanized derivative (MDX-H210), and its parent MoAb 520C9 (anti-HER-2/neu) under various conditions. Purified monocytes were cultured with GM-CSF, M-CSF, or no cytokine for five or six days. Antibody dependent cellular phagocytosis (ADCP) and cytolysis (ADCC) assays were performed with the MDM and HER-2/neu positive target cells (SK-BR-3). ADCP was measured by two-color fluorescence flow cytometry using PKH2 (green fluorescent dye) and phycoerythrin-conjugated (red) monoclonal antibodies (MoAb) against human CD14 and CD11b. ADCC was measured with a non-radioactive LDH detection kit. Both BsAb MDX-210 (via FcgammaRI) and MoAb 520C9 (mouse IgG1, via FcgammaRII) mediated similar levels of ADCP and ADCC. ADCP mediated by BsAb MDX-H210 was identical to that mediated by BsAb MDX-210. Confocal microscopy demonstrated that dual-labeled cells represented true phagocytosis. Both ADCP and ADCC were higher when MDM were pre-incubated with GM-CSF than when incubated with M-CSF. BsAb MDX-210 is as active in vitro as the parent MoAb 520C9 in inducing both phagocytosis and cytolysis of MDM. MDX-210 and its partially humanized derivative, MDX-H210, mediated similar levels of ADCP. GM-CSF appears to superior to M-CSF in inducing MDM-mediated ADCC and ADCP. These studies support the ongoing clinical investigations of BsAb MDX-210 and its partially humanized derivative.

Shape memory of human red blood cells.

Science.gov (United States)

Fischer, Thomas M

2004-05-01

The human red cell can be deformed by external forces but returns to the biconcave resting shape after removal of the forces. If after such shape excursions the rim is always formed by the same part of the membrane, the cell is said to have a memory of its biconcave shape. If the rim can form anywhere on the membrane, the cell would have no shape memory. The shape memory was probed by an experiment called go-and-stop. Locations on the membrane were marked by spontaneously adhering latex spheres. Shape excursions were induced by shear flow. In virtually all red cells, a shape memory was found. After stop of flow and during the return of the latex spheres to the original location, the red cell shape was biconcave. The return occurred by a tank-tread motion of the membrane. The memory could not be eliminated by deforming the red cells in shear flow up to 4 h at room temperature as well as at 37 degrees C. It is suggested that 1). the characteristic time of stress relaxation is >80 min and 2). red cells in vivo also have a shape memory.

Noncytotoxic orange and red/green derivatives of DsRed-Express2 for whole-cell labeling

Directory of Open Access Journals (Sweden)

Glick Benjamin S

2009-04-01

Full Text Available Abstract Background Whole-cell labeling is a common application of fluorescent proteins (FPs, but many red and orange FPs exhibit cytotoxicity that limits their use as whole-cell labels. Recently, a tetrameric red FP called DsRed-Express2 was engineered for enhanced solubility and was shown to be noncytotoxic in bacterial and mammalian cells. Our goal was to create derivatives of this protein with different spectral properties. Results Building on previous studies of DsRed mutants, we created two DsRed-Express2 derivatives: E2-Orange, an orange FP, and E2-Red/Green, a dual-color FP with both red and green emission. We show that these new FPs retain the low cytotoxicity of DsRed-Express2. In addition, we show that these new FPs are useful as second or third colors for flow cytometry and fluorescence microscopy. Conclusion E2-Orange and E2-Red/Green will facilitate the production of healthy, stably fluorescent cell lines and transgenic organisms for multi-color labeling studies.

Single-cell measurement of red blood cell oxygen affinity

OpenAIRE

Caprio, Di; Stokes, Chris; Higgins, John M.; Schonbrun, Ethan

2015-01-01

Oxygen is transported throughout the body by hemoglobin in red blood cells. While the oxygen affinity of blood is well understood and is routinely assessed in patients by pulse oximetry, variability at the single-cell level has not been previously measured. In contrast, single-cell measurements of red blood cell volume and hemoglobin concentration are taken millions of times per day by clinical hematology analyzers and are important factors in determining the health of the hematologic system....

21 CFR 660.30 - Reagent Red Blood Cells.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 7 2010-04-01 2010-04-01 false Reagent Red Blood Cells. 660.30 Section 660.30...) BIOLOGICS ADDITIONAL STANDARDS FOR DIAGNOSTIC SUBSTANCES FOR LABORATORY TESTS Reagent Red Blood Cells § 660.30 Reagent Red Blood Cells. (a) Proper name and definition. The proper name of the product shall be...

Separation of breast cancer cells from peripherally circulating blood using antibodies fixed in microchannels

Science.gov (United States)

Feng, Juan; Soper, Steven A.; McCarley, Robin L.; Murphy, Michael C.

2004-07-01

Bio-Micro Electro Mechanical System (Bio-MEMS) technology was applied to the problem of early breast cancer detection and diagnosis. A micro-device is being developed to identify and specifically collect tumor cells of low abundance (1 tumor cell among 107 normal blood cells) from circulating whole blood. By immobilizing anti-EpCAM (Epithelial Cell Adhesion Molecule) antibodies on polymer micro-channel walls by chemically modifying the surface of the PMMA, breast cancer cells from the MCF-7 cell line, which over-express EpCAM, were selected from a sample volume by the strong binding affinity between the antibody and antigen. To validate the capture of the breast cancer cells, three fluorochrome markers, each identified by a separate color, were used to reliably identify the cancer cells. The cancer cells were defined by DAPI+ (blue), CD45- and the FITC-cell membrane linker+ (green). White blood cells, which may interfere in the detection of the cancer cells, were identified by DAPI+ (blue), CD45+ (red), and the FITC-cell membrane linker+ (green). EpCAM/anti-EpCAM binding models from the literature were used to estimate an optimal velocity, 2mm/sec, for maximizing the number of cells binding and the critical binding force. At higher velocities, shear forces (> 0.48 dyne) will break existing bonds and prevent the formation of new ones. This detection micro-device can be assembled with other lab-on-a-chip components for follow-up gene and protein analysis.

Mycoplasma infection of cell lines can simulate the expression of Fc receptors by binding of the carbohydrate moiety of antibodies.

Science.gov (United States)

Lemke, H; Krausse, R; Lorenzen, J; Havsteen, B

1985-05-01

During the production of Fc receptor (FcR)-bearing hybridomas it was observed with a particular monoclonal anti-sheep red blood cell antibody (anti-SRBC 1/5, IgG1) that the contamination with Mycoplasma arginini of in vitro cultured cell lines leads to an apparent FcR activity. This property did not correspond with the serological typing since other antibodies of the same isotype could not support FcR rosette formation. Another mycoplasma strain M. orale lacked this property. Analysis of the binding reaction revealed that M. arginini contains a lectin which binds the carbohydrate moiety of the anti-SRBC 1/5 antibody, i.e. anti-SRBC 1/5 synthesized under the influence of tunicamycin or deglycosylated by NaIO4 oxidation did not support rosette formation. These data suggest that binding of antibodies to certain mycoplasma strains may be a pathogenic factor during mycoplasma infections by masking the microorganisms with the host's own defense molecules. The experiments with M. arginini-infected cell lines gain immunological importance since we obtained identical results with staphylococcal protein A, as another bacteriological FcR, and cell lines expressing intrinsic membrane FcR. Although it is an open question whether the glycoconjugates are directly bound by the FcR or else by influencing the three-dimensional structure of the antibodies, it seems possible that FcR in general may be lectins.

Antithyroglobulin Antibodies and Antimicrosomal Antibodies in Various Thyroid Diseases

International Nuclear Information System (INIS)

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

1979-01-01

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.

Human Red Cells With Paroxysmal Nocturnal Haemoglobinuria ...

African Journals Online (AJOL)

The purified cells were used as hosts for the culture of P.falciparum in vitro. Results show that GPI-linked molecules on the red cell surface are not required for the efficient entry of the parasites, and that the PNH red cells are competent to sustain the growth of P.falciparum. Nigerian Quarterly Journal of Hospital Medicine Vol ...

The homeostasis of Plasmodium falciparum-infected red blood cells.

Directory of Open Access Journals (Sweden)

Jakob M A Mauritz

2009-04-01

Full Text Available The asexual reproduction cycle of Plasmodium falciparum, the parasite responsible for severe malaria, occurs within red blood cells. A merozoite invades a red cell in the circulation, develops and multiplies, and after about 48 hours ruptures the host cell, releasing 15-32 merozoites ready to invade new red blood cells. During this cycle, the parasite increases the host cell permeability so much that when similar permeabilization was simulated on uninfected red cells, lysis occurred before approximately 48 h. So how could infected cells, with a growing parasite inside, prevent lysis before the parasite has completed its developmental cycle? A mathematical model of the homeostasis of infected red cells suggested that it is the wasteful consumption of host cell hemoglobin that prevents early lysis by the progressive reduction in the colloid-osmotic pressure within the host (the colloid-osmotic hypothesis. However, two critical model predictions, that infected cells would swell to near prelytic sphericity and that the hemoglobin concentration would become progressively reduced, remained controversial. In this paper, we are able for the first time to correlate model predictions with recent experimental data in the literature and explore the fine details of the homeostasis of infected red blood cells during five model-defined periods of parasite development. The conclusions suggest that infected red cells do reach proximity to lytic rupture regardless of their actual volume, thus requiring a progressive reduction in their hemoglobin concentration to prevent premature lysis.

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...... 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....

B Cells and B Cell Blasts Withstand Cryopreservation While Retaining Their Functionality for Producing Antibody.

Science.gov (United States)

Fecher, Philipp; Caspell, Richard; Naeem, Villian; Karulin, Alexey Y; Kuerten, Stefanie; Lehmann, Paul V

2018-05-31

In individuals who have once developed humoral immunity to an infectious/foreign antigen, the antibodies present in their body can mediate instant protection when the antigen re-enters. Such antigen-specific antibodies can be readily detected in the serum. Long term humoral immunity is, however, also critically dependent on the ability of memory B cells to engage in a secondary antibody response upon re-exposure to the antigen. Antibody molecules in the body are short lived, having a half-life of weeks, while memory B cells have a life span of decades. Therefore, the presence of serum antibodies is not always a reliable indicator of B cell memory and comprehensive monitoring of humoral immunity requires that both serum antibodies and memory B cells be assessed. The prevailing view is that resting memory B cells and B cell blasts in peripheral blood mononuclear cells (PBMC) cannot be cryopreserved without losing their antibody secreting function, and regulated high throughput immune monitoring of B cell immunity is therefore confined to-and largely limited by-the need to test freshly isolated PBMC. Using optimized protocols for freezing and thawing of PBMC, and four color ImmunoSpot ® analysis for the simultaneous detection of all immunoglobulin classes/subclasses we show here that both resting memory B cells and B cell blasts retain their ability to secrete antibody after thawing, and thus demonstrate the feasibility of B cell immune monitoring using cryopreserved PBMC.

B Cells and B Cell Blasts Withstand Cryopreservation While Retaining Their Functionality for Producing Antibody

Directory of Open Access Journals (Sweden)

Philipp Fecher

2018-05-01

Full Text Available In individuals who have once developed humoral immunity to an infectious/foreign antigen, the antibodies present in their body can mediate instant protection when the antigen re-enters. Such antigen-specific antibodies can be readily detected in the serum. Long term humoral immunity is, however, also critically dependent on the ability of memory B cells to engage in a secondary antibody response upon re-exposure to the antigen. Antibody molecules in the body are short lived, having a half-life of weeks, while memory B cells have a life span of decades. Therefore, the presence of serum antibodies is not always a reliable indicator of B cell memory and comprehensive monitoring of humoral immunity requires that both serum antibodies and memory B cells be assessed. The prevailing view is that resting memory B cells and B cell blasts in peripheral blood mononuclear cells (PBMC cannot be cryopreserved without losing their antibody secreting function, and regulated high throughput immune monitoring of B cell immunity is therefore confined to—and largely limited by—the need to test freshly isolated PBMC. Using optimized protocols for freezing and thawing of PBMC, and four color ImmunoSpot® analysis for the simultaneous detection of all immunoglobulin classes/subclasses we show here that both resting memory B cells and B cell blasts retain their ability to secrete antibody after thawing, and thus demonstrate the feasibility of B cell immune monitoring using cryopreserved PBMC.

Effects of helicopter transport on red blood cell components.

Science.gov (United States)

Otani, Taiichi; Oki, Ken-ichi; Akino, Mitsuaki; Tamura, Satoru; Naito, Yuki; Homma, Chihiro; Ikeda, Hisami; Sumita, Shinzou

2012-01-01

There are no reported studies on whether a helicopter flight affects the quality and shelf-life of red blood cells stored in mannitol-adenine-phosphate. Seven days after donation, five aliquots of red blood cells from five donors were packed into an SS-BOX-110 container which can maintain the temperature inside the container between 2 °C and 6 °C with two frozen coolants. The temperature of an included dummy blood bag was monitored. After the box had been transported in a helicopter for 4 hours, the red blood cells were stored again and their quality evaluated at day 7 (just after the flight), 14, 21 and 42 after donation. Red blood cell quality was evaluated by measuring adenosine triphosphate, 2,3-diphosphoglycerate, and supernatant potassium, as well as haematocrit, intracellular pH, glucose, supernatant haemoglobin, and haemolysis rate at the various time points. During the experiment the recorded temperature remained between 2 and 6 °C. All data from the red blood cells that had undergone helicopter transportation were the same as those from a control group of red blood cell samples 7 (just after the flight), 14, 21, and 42 days after the donation. Only supernatant Hb and haemolysis rate 42 days after the donation were slightly increased in the helicopter-transported group of red blood cell samples. All other parameters at 42 days after donation were the same in the two groups of red blood cells. These results suggest that red blood cells stored in mannitol-adenine-phosphate are not significantly affected by helicopter transportation. The differences in haemolysis by the end of storage were small and probably not of clinical significance.

Efficacy of Wnt-1 monoclonal antibody in sarcoma cells

International Nuclear Information System (INIS)

Mikami, Iwao; Koizumi, Kiyoshi; Jablons, David M; You, Liang; He, Biao; Xu, Zhidong; Batra, Sonny; Lee, Amie Y; Mazieres, Julien; Reguart, Noemi; Uematsu, Kazutsugu

2005-01-01

Sarcomas are one of the most refractory diseases among malignant tumors. More effective therapies based on an increased understanding of the molecular biology of sarcomas are needed as current forms of therapy remain inadequate. Recently, it has been reported that Wnt-1/β-catenin signaling inhibits apoptosis in several cancers. In this study, we investigated the efficacy of a monoclonal anti-Wnt-1 antibody in sarcoma cells. We treated cell lines A-204, SJSA-1, and fresh primary cultures of lung metastasis of sarcoma with a monoclonal anti-Wnt-1 antibody. Wnt-1 siRNA treatment was carried out in A-204. We assessed cell death using Crystal Violet staining. Apoptosis induction was estimated by flow cytometry analysis (Annexin V and PI staining). Cell signaling changes were determined by western blotting analysis. We detected Wnt-1 expression in all tissue samples and cell lines. Significant apoptosis induction was found in monoclonal anti-Wnt-1 antibody treated cells compared to control monoclonal antibody treated cells (p < 0.02). Similarly, we observed increased apoptosis in Wnt-1 siRNA treated cells. Blockade of Wnt-1 signaling in both experiments was confirmed by analyzing intracellular levels of Dishevelled-3 and of cytosolic β-catenin. Furthermore, the monoclonal anti-Wnt-1 antibody also induced cell death in fresh primary cultures of metastatic sarcoma in which Wnt-1 signaling was active. Our results indicate that Wnt-1 blockade by either monoclonal antibody or siRNA induces cell death in sarcoma cells. These data suggest that Wnt-1 may be a novel therapeutic target for the treatment of a subset of sarcoma cells in which Wnt-1/β-catenin signaling is active

Red cell ferritin and iron stores in chronic granulocytic leukemia

International Nuclear Information System (INIS)

Cermak, J.; Neuwirth, J.; Voglova, J.; Brabec, V.; Chrobak, L.

1994-01-01

Basic red cell ferritin was investigated in 28 patients with different phases of chronic granulocytic leukemia (GCL). Red cell ferritin was significantly decreased in remission after busulphan treatment and significantly elevated in the blast crisis as compared to healthy controls. Bone marrow stainable iron was decreased or absent in 86% of patients in the initial phase at the time of diagnosis and in 92% of those in remission. Red cell ferritin correlated with serum ferritin, however, serum ferritin level remained above normal range during all phases of the disease. A negative correlation between red cell ferritin and hemoglobin (Hb) (r = -0.605, p < 0.001) suggested that red cell ferritin level reflected the rate of iron utilization for heme synthesis. Decrease red cell iron observed in the remission may be explained by regression of dyserythropoiesis and by restoration of normal Hb synthesis after busulphan treatment. A progressive dyserythropoiesis in the blast crisis may lead to an increased red cell ferritin level. (author)

Effects of helicopter transport on red blood cell components

Science.gov (United States)

Otani, Taiichi; Oki, Ken-ichi; Akino, Mitsuaki; Tamura, Satoru; Naito, Yuki; Homma, Chihiro; Ikeda, Hisami; Sumita, Shinzou

2012-01-01

Background There are no reported studies on whether a helicopter flight affects the quality and shelf-life of red blood cells stored in mannitol-adenine-phosphate. Materials and methods Seven days after donation, five aliquots of red blood cells from five donors were packed into an SS-BOX-110 container which can maintain the temperature inside the container between 2 °C and 6 °C with two frozen coolants. The temperature of an included dummy blood bag was monitored. After the box had been transported in a helicopter for 4 hours, the red blood cells were stored again and their quality evaluated at day 7 (just after the flight), 14, 21 and 42 after donation. Red blood cell quality was evaluated by measuring adenosine triphosphate, 2,3-diphosphoglycerate, and supernatant potassium, as well as haematocrit, intracellular pH, glucose, supernatant haemoglobin, and haemolysis rate at the various time points. Results During the experiment the recorded temperature remained between 2 and 6 °C. All data from the red blood cells that had undergone helicopter transportation were the same as those from a control group of red blood cell samples 7 (just after the flight), 14, 21, and 42 days after the donation. Only supernatant Hb and haemolysis rate 42 days after the donation were slightly increased in the helicopter-transported group of red blood cell samples. All other parameters at 42 days after donation were the same in the two groups of red blood cells. Discussion These results suggest that red blood cells stored in mannitol-adenine-phosphate are not significantly affected by helicopter transportation. The differences in haemolysis by the end of storage were small and probably not of clinical significance. PMID:22153688

A noncognate interaction with anti-receptor antibody-activated helper T cells induces small resting murine B cells to proliferate and to secrete antibody

DEFF Research Database (Denmark)

Owens, T

1988-01-01

on resting B cells (even in the presence of intact F23.1 antibody), but could induce antibody secretion by anti-Ig-preactivated B cells. Both F23.1+ clones (E9.D4 and 4.35F2) and one F23.1- clone (D2.2) could synergize with supernatants from activated E9.D4 T cells to induce B cell activation. F(ab')2......Culture of small resting allogeneic B cells (of an irrelevant haplotype) with two clones of T helper (Th) cells that were activated by the F23.1 anti-T cell receptor antibody led to the activation of B cells to proliferate and to secrete antibody. Th cell supernatants by themselves had no effect...... fragments of F23.1 induced E9.D4 to activate B cells as efficiently as intact F23.1 and B cell populations that had been incubated with F23.1 were not activated when cultured with E9.D4, although T cells recognized cell-presented F23.1 and were weakly activated. Reduction of the density of F23.1 adsorbed...

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.

Study of red blood cell alloimmunization in multitransfused thalassemic children of Jammu region

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Ashu Dogra

2015-01-01

Full Text Available Introduction: Thalassemia is one of the most common genetic disorder of hemoglobin synthesis in Jammu region. Although RBC transfusion is life saving for these patients, it may be associated with some complications like RBC alloimmunization. Thus, the aim of this study was to determine the frequency of alloimmunization and the most common alloantibodies involved. Material and Methods: This was a descriptive study involving a total of 70 thalassemic patients in the age range of 2-17 years receiving regular blood transfusions, registered at SMGS Blood Bank, Jammu. Relevant clinical and laboratory data was collected with reference to age at the start of transfusions, total number of transfusions received and splenectomy status. Antibodies screening, antibody identification, and cross matching was done on allpatient samples included in the study, during the period between November 2009 and October 2010. Results: In this study, a total of six alloantibodies six patients (8.5% and one autoantibody (1.42% was detected. All identified alloantibodies belonged to Rh system (i.e. anti-E, in 3 patients (50%, anti D, in one patient (16.66% and Kell system (anti-K, in two patients (33.34%. Higher frequency of alloimmunization was found, with increase in number of transfusions and in those who received transfusions after 1 year of age. Alloimmunization was not significantly associated with gender and splenectomy status (P-value > 0.05. Conclusion: Red cell alloantibodies developed in 8.5% of thalassemic patients and 1.42% had autoantibodies. The most common alloantibodies identified were anti Rh system antibodies (anti-E and anti-D present in 50% and 16.66% of patients respectively. Alloimmunization is not an uncommon problem faced by blood banks and finding compatible units for regularly transfused thalassemic patients may become very difficult. In order to reduce alloimmunization, a policy for performing extended red cell phenotyping of these patients is

Trypanosoma cruzi infection induces a massive extrafollicular and follicular splenic B-cell response which is a high source of non-parasite-specific antibodies.

Science.gov (United States)

Bermejo, Daniela A; Amezcua Vesely, María C; Khan, Mahmood; Acosta Rodríguez, Eva V; Montes, Carolina L; Merino, Maria C; Toellner, Kai Michael; Mohr, Elodie; Taylor, Dale; Cunningham, Adam F; Gruppi, Adriana

2011-01-01

Acute infection with Trypanosoma cruzi, the aetiological agent of Chagas' disease, results in parasitaemia and polyclonal lymphocyte activation. It has been reported that polyclonal B-cell activation is associated with hypergammaglobulinaemia and delayed parasite-specific antibody response. In the present study we analysed the development of a B-cell response within the different microenvironments of the spleen during acute T. cruzi infection. We observed massive germinal centre (GC) and extrafollicular (EF) responses at the peak of infection. However, the EF foci were evident since day 3 post-infection (p.i.), and, early in the infection, they mainly provided IgM. The EF foci response reached its peak at 11 days p.i. and extended from the red pulp into the periarteriolar lymphatic sheath. The GCs were detected from day 8 p.i. At the peak of parasitaemia, CD138(+) B220(+) plasma cells in EF foci, red pulp and T-cell zone expressed IgM and all the IgG isotypes. Instead of the substantial B-cell response, most of the antibodies produced by splenic cells did not target the parasite, and parasite-specific IgG isotypes could be detected in sera only after 18 days p.i. We also observed that the bone marrow of infected mice presented a strong reduction in CD138(+) B220(+) cells compared with that of normal mice. Hence, in acute infection with T. cruzi, the spleen appears to be the most important lymphoid organ that lodges plasma cells and the main producer of antibodies. The development of a B-cell response during T. cruzi infection shows features that are particular to T. cruzi and other protozoan infection but different to other infections or immunization with model antigens.

Antibodies against alpha-synuclein reduce oligomerization in living cells.

Directory of Open Access Journals (Sweden)

Thomas Näsström

Full Text Available Recent research implicates soluble aggregated forms of α-synuclein as neurotoxic species with a central role in the pathogenesis of Parkinson's disease and related disorders. The pathway by which α-synuclein aggregates is believed to follow a step-wise pattern, in which dimers and smaller oligomers are initially formed. Here, we used H4 neuroglioma cells expressing α-synuclein fused to hemi:GFP constructs to study the effects of α-synuclein monoclonal antibodies on the early stages of aggregation, as quantified by Bimolecular Fluorescence Complementation assay. Widefield and confocal microscopy revealed that cells treated for 48 h with monoclonal antibodies internalized antibodies to various degrees. C-terminal and oligomer-selective α-synuclein antibodies reduced the extent of α-synuclein dimerization/oligomerization, as indicated by decreased GFP fluorescence signal. Furthermore, ELISA measurements on lysates and conditioned media from antibody treated cells displayed lower α-synuclein levels compared to untreated cells, suggesting increased protein turnover. Taken together, our results propose that extracellular administration of monoclonal antibodies can modify or inhibit early steps in the aggregation process of α-synuclein, thus providing further support for passive immunization against diseases with α-synuclein pathology.

Radiolabeled red blood cells: status, problems, and prospects

Energy Technology Data Exchange (ETDEWEB)

Srivastava, S.C.

1983-01-01

Radionuclidic labels for red cells can be divided into two main categories - cohort or pulse labels, and random labels. The random labels are incorporated into circulating cells of all ages and the labeling process is usually carried out in vitro. The red cell labels in predominant use involve random labeling and employ technetium-99m, chromium-51, indium-111, and gallium-68, roughly in that order. The extent of usefulness depends on the properties of the label such as the half-life, decay mode, and in-vivo stability, etc. Labeled cells can be used for red cell survival measurements when the half-life of the radionuclide is sufficiently long. The major portion of this article deals with random labels.

Radiolabeled red blood cells: status, problems, and prospects

International Nuclear Information System (INIS)

Srivastava, S.C.

1983-01-01

Radionuclidic labels for red cells can be divided into two main categories - cohort or pulse labels, and random labels. The random labels are incorporated into circulating cells of all ages and the labeling process is usually carried out in vitro. The red cell labels in predominant use involve random labeling and employ technetium-99m, chromium-51, indium-111, and gallium-68, roughly in that order. The extent of usefulness depends on the properties of the label such as the half-life, decay mode, and in-vivo stability, etc. Labeled cells can be used for red cell survival measurements when the half-life of the radionuclide is sufficiently long. The major portion of this article deals with random labels

Clinically significant anti M antibodies--a report of two cases.

Science.gov (United States)

Kaur, Gagandeep; Basu, Sabita; Kaur, Paramjit; Kaur, Ravneet

2012-12-01

Most anti-M antibodies are not active at 37°C and are thus of no clinical significance. Occasionally these antibodies have a wide thermal range and can lead to hemolytic transfusion reactions or hemolytic disease of the new born. We describe two cases of anti-M antibodies, both of which were clinically significant. The first case was detected due to crossmatch incompatibility and the second presented as a blood group discrepancy. When the antibody is active at 37°C, M antigen negative red cell units should be issued. Copyright © 2012 Elsevier Ltd. All rights reserved.

Hemolytic disease of the fetus and newborn due to anti-Ge3: combined antibody-dependent hemolysis and erythroid precursor cell growth inhibition.

Science.gov (United States)

Blackall, Douglas P; Pesek, Gina D; Montgomery, Matthew M; Oza, Krishna K; Arndt, Patricia A; Garratty, George; Shahcheraghi, Ali; Denomme, Gregory A

2008-10-01

The Gerbich (Ge) antigens are a collection of high-incidence antigens carried on the red blood cell membrane glycoproteins, glycophorins C and D. Antibodies against these antigens are uncommon, and there have been only rare case reports of hemolytic disease of the fetus and newborn due to anti-Ge. In this case report, we present a neonate with severe anemia and hyperbilirubinemia due to anti-Ge3. Routine and special laboratory studies undertaken in this case suggested two mechanisms for the patient's hemolysis and persistent anemia. Antibody-dependent hemolysis was associated with early-onset hyperbilirubinemia, anemia, and a mild reticulocytosis, and inhibition of erythroid progenitor cell growth was associated with late anemia and normal bilirubin and reticulocyte values. Though rare, anti-Ge3 can be a dangerous antibody in pregnancy. Affected neonates may require intensive initial therapy and close follow-up for at least several weeks after delivery.

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.

Rapid isolation of antibody from a synthetic human antibody library by repeated fluorescence-activated cell sorting (FACS.

Directory of Open Access Journals (Sweden)

Sung Sun Yim

Full Text Available Antibodies and their derivatives are the most important agents in therapeutics and diagnostics. Even after the significant progress in the technology for antibody screening from huge libraries, it takes a long time to isolate an antibody, which prevents a prompt action against the spread of a disease. Here, we report a new strategy for isolating desired antibodies from a combinatorial library in one day by repeated fluorescence-activated cell sorting (FACS. First, we constructed a library of synthetic human antibody in which single-chain variable fragment (scFv was expressed in the periplasm of Escherichia coli. After labeling the cells with fluorescent antigen probes, the highly fluorescent cells were sorted by using a high-speed cell sorter, and these cells were reused without regeneration in the next round of sorting. After repeating this sorting, the positive clones were completely enriched in several hours. Thus, we screened the library against three viral antigens, including the H1N1 influenza virus, Hepatitis B virus, and Foot-and-mouth disease virus. Finally, the potential antibody candidates, which show K(D values between 10 and 100 nM against the target antigens, could be successfully isolated even though the library was relatively small (∼ 10(6. These results show that repeated FACS screening without regeneration of the sorted cells can be a powerful method when a rapid response to a spreading disease is required.

How immunoglobulin G antibodies kill target cells: revisiting an old paradigm.

Science.gov (United States)

Biburger, Markus; Lux, Anja; Nimmerjahn, Falk

2014-01-01

The capacity of immunoglobulin G (IgG) antibodies to eliminate virtually any target cell has resulted in the widespread introduction of cytotoxic antibodies into the clinic in settings of cancer therapy, autoimmunity, and transplantation, for example. More recently, it has become apparent that also the protection from viral infection via IgG antibodies may require cytotoxic effector functions, suggesting that antibody-dependent cellular cytotoxicity (ADCC) directed against malignant or virally infected cells is one of the most essential effector mechanisms triggered by IgG antibodies to protect the host. A detailed understanding of the underlying molecular and cellular pathways is critical, therefore, to make full use of this antibody effector function. Several studies over the last years have provided novel insights into the effector pathways and innate immune effector cells responsible for ADCC reactions. One of the most notable outcomes of many of these reports is that cells of the mononuclear phagocytic system rather than natural killer cells are critical for removal of IgG opsonized target cells in vivo. © 2014 Elsevier Inc. All rights reserved.

Radioimmunodetection of tumor with Ga-67 labeled antibodies

International Nuclear Information System (INIS)

Furukawa, Takako; Endo, Keigo; Ohmomo, Yoshiro

1986-01-01

Antibodies against tumor associated antigen; anti-AFP polyclonal antibody, anti-thyroglobulin monoclonal antibody and anti-hCG monoclonal antibody, were labeled with Ga-67, using deferoxamine (DF) as a bifunctional chelating agent. The immunoreactivity and in vivo stability of the Ga-67 labeled antibodies were examined. The effect of DF conjugation to antibodies on the antigen-binding activity was evaluated by RIA and Scatchard analysis or tanned sheep red blood cell hemagglutination technique. When DF was conjugated to antibody at the molar ratio of 1 : 1, the antibody activity of the DF-conjugated antibodies was fully retained. Whereas, in heavily conjugated antibodies, the maximum antigen binding capacity was reduced. Biodistribution study in normal mice demonstrated the high in vivo stability of Ga-67 labeled antibodies. The labeling of DF-antibody conjugated with Ga-67 was performed easily and quickly, with a high labeling efficiency, requiring no further purification. Thus, this labeling method, providing in vivo stability of Ga-67 labeled antibody and full retention of immunoreactivity, would be useful for the radioimmunodetection of various cancers. (author)

In vivo regeneration of red cell 2,3-diphosphoglycerate following transfusion of DPG-depleted AS-1, AS-3 and CPDA-1 red cells.

Science.gov (United States)

Heaton, A; Keegan, T; Holme, S

1989-01-01

Regeneration of 2,3-diphosphoglycerate (DPG) was determined following transfusion of DPG-depleted group O red cells into group A recipients. Blood from five donors was stored in the adenine-containing solutions CPDA-1, AS-1 or AS-3 for 35 d at 4 degrees C. Post-transfusion red cell DPG and ATP were measured in separated group O red cells over a 7 d period. The studies confirmed rapid in vivo DPG regeneration with greater than or equal to 50% of the maximum level being achieved within 7 h. An average of 95% of the recipients' pre-transfusion DPG level was achieved by 72 h and by 7 d mean (+/- SEM) DPG levels relative to recipient's pre-transfusion DPG averaged 84% (+/- 13%), 92% (+/- 17%) and 84% (+/- 21%) for CPDA-1, AS-1 and AS-3 red cells, respectively. Results were comparable to those previously reported for blood stored in ACD for 15-20 d (Valeri & Hirsch, 1969; Beutler & Wood, 1969). The immediate regeneration rate, V, closely approximated first order regeneration kinetics with AS-3 red cells exhibiting double the rate of CPDA-1 red cells (P less than 0.001). AS-1 red cells exhibited an intermediate rate of regeneration which was not significantly different compared to either CPDA-1 or AS-3 (P greater than 0.05). V exhibited a significant (P less than 0.05) positive correlation with ATP levels 5-7 h post-infusion. ATP regeneration of the infused cells was rapid with a mean increase of 1.2 mumol/g Hb above post-storage levels being achieved 1 h following transfusion.

Red blood-cell alloantibodies in multiply transfused patients in the occupied Palestinian territory: a pilot study.

Science.gov (United States)

Yaseen, Ahmad; Suleiman, Sa'd; Zenah, Omar Abu; Abu Taha, Adham

2018-02-21

Red blood-cell transfusion has greatly reduced the mortality and morbidity in multiply transfused patients with thalassaemia and sickle cell disease. However, this can result in red blood-cell isoimmunisation with autoantibodies and alloantibodies, which can lead to serious complications such as delayed haemolytic transfusion reaction. The aim of this study was to assess the frequency and types of alloantibodies in multiply transfused patients living in the north of the West Bank. This pilot study was done at three thalassaemia centres in Nablus, Jenin, and Tulkarm in the occupied Palestinian territory where 300 patients with thalassaemia and sickle cell anaemia regularly receive blood transfusions. Alloantibody screening and identification were done using three-cell and eleven-cell panels (DiaPanel, Bio-rad, Switzerland) respectively. Ethical approval was obtained from Institutional Review Board Centre at Najah University. Written consent was obtained from participants. 131 patients were enrolled. Of the 20 (15%) patients with alloantibodies, 14 (70%) were diagnosed with β-thalassaemia major, three (15%) were diagnosed with sickle cell anaemia, two (10%) were diagnosed with thalassaemia intermedia, and one (5%) was diagnosed with sickle cell thalassaemia. 13 (65%) patients had alloantibodies that belonged to the Rh blood group system (nine [45%] patients had anti-D; two [10%] had anti-E; one [5%] had anti Rh-C; and one [5%] had anti-c). Anti-Kell was found in seven (35%) patients. Our data showed a quite high prevalence of alloimmunisation in multiply transfused patients. Rh and Kell blood group system antibodies were the only alloantibodies identified in this study. To reduce alloimmunisation, it will be essential to introduce a policy for extended red blood-cell phenotyping of these patients and for the issuing of antigen-matched blood (at least for Rh and Kell antigen). Najah National University. Copyright © 2018 Elsevier Ltd. All rights reserved.

Evidence that cell surface charge reduction modifes capillary red cell velocity-flux relationships in hamster cremaster muscle

NARCIS (Netherlands)

Vink, H.; Wieringa, P. A.; Spaan, J. A.

1995-01-01

1. From capillary red cell velocity (V)-flux (F) relationships of hamster cremaster muscle a yield velocity (VF = 0) can be derived at which red cell flux is zero. Red cell velocity becomes intermittent and/or red blood cells come to a complete standstill for velocities close to this yield velocity,

Evaluation of radioimmunoassay of anti-thyroglobulin antibodies

International Nuclear Information System (INIS)

Lourme, J.; Dessaint, J.P.; Capron, A.

1985-01-01

A statistical analysis is performed on the results of 881 determinations of thyroglobulin antibodies in humans. Antibodies were assayed comparatively by radioimmunoassay using a sandwich method and by tanned red cell haemagglutination. A very good concordance was found between the two techniques, apart from the low titer zone. A significant correlation was observed between on the one side, the radioactivity index of the diluted serum, defined as the increment of radioactivity bound by undiluted patient serum over the positive threshold, divided by this threshold, and, on the other side, the antibody titer, i.e. the reciprocal of the highest serum dilution superior to the positive threshold by radioimmunoassay. The corresponding linear regression allows to define a arbitrary unit system which associates values of the radioactivity index with an average antibody titer [fr

21 CFR 864.8540 - Red cell lysing reagent.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Red cell lysing reagent. 864.8540 Section 864.8540 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Reagents § 864.8540 Red cell lysing reagent...

A simple assay for the detection of antibodies to endocrine islet cell surface antigens

International Nuclear Information System (INIS)

Contreas, G.; Madsen, O.D.; Vissing, H.; Lernmark, Aa.

1986-01-01

A simple and sensitive immunoradiometric assay for the detection of islet cell surface antibodies (CIRMA) has been developed. Live, transformed islet cells derived from a liver metastasis of a transplantable islet cell tumor were grown in removable microtiter wells and incubated with antibody. Cell-bound antibodies were quantitated using 125 I-labelled second antibodies. The assay was used to detect islet cell antibodies present in sera from non-diabetic and diabetic BB rats and proved to be particularly effective for screening hybridoma supernatants in order to identify monoclonal antibodies recognizing islet cell surface antigens. (Auth.)

Hyperemic peripheral red marrow in a patient with sickle cell anemia demonstrated on Tc-99m labeled red blood cell venography

International Nuclear Information System (INIS)

Heiden, R.A.; Locko, R.C.; Stent, T.R.

1991-01-01

A 25-year-old gravid woman, homozygous for sickle cell anemia, with a history of recent deep venous thrombosis, was examined using Tc-99m labeled red blood cell venography for recurrent thrombosis. Although negative for thrombus, the study presented an unusual incidental finding: the patient's peripheral bone marrow was hyperemic in a distribution consistent with peripheral red bone marrow expansion. Such a pattern has not been documented before using this technique. This report supports other literature that has demonstrated hyperemia of peripheral red bone marrow in other hemolytic anemias. This finding may ultimately define an additional role of scintigraphy in assessing the pathophysiologic status of the sickle cell patient

The restoration in vivo of 2,3-diphosphoglycerate (2,3-DPG) in stored red cells, after transfusion. The levels of red cells 2,3-DPG.

Science.gov (United States)

Stan, Ana; Zsigmond, Eva

2009-01-01

Since the main reason for transfusing preserved red cells is to increase the oxygen carrying capacity of the recipient, the circulating preserved red cells should have at the time of transfusion normal oxygen uptake and normal oxyhemoglobin dissociation characteristics. We evaluated the effectiveness of transfused red cells, through periodical determination of erythrocyte components, during 72 hours after transfusions of large quantities (3,000 mL) of blood. Three patients with massive hemorrhages, two after amputation and one after nephrectomy were given each 3,000 mL preserved blood (in ACD, 10 days, at 4 degrees C). Red cell 2,3-DPG and serum inorganic phosphorus were determined prior to transfusion and after, periodically, for three days. Red cell 2,3-DPG was determined by Krimsky's method and inorganic phosphorus by Kuttner and Lichtenstein's method. The in vivo restoration of 2,3-DPG--of transfused red cells is shown as a percentage of recipient's final 2,3-DPG level, and was calculated in each of the three patients. The level of erythrocyte 2,3-DPG was greater than 60% of the final level within 24 hours, after the end of transfusion. The in vivo rates of restoration of 2,3-DPG in transfused red cells for periods of 0-6, 6-24, 24-48 and 48-72 hours are 0.251, 0.238, 0.133, 0.120 mM/L cells/hour. The therapeutic significance of the increased oxygen affinity of stored blood becomes very important in clinical conditions, when large volumes of red cells are urgently needed. After massive transfusions, the restoration of 2,3-DPG in red cells produces a decrease of serum inorganic phosphorus through its consumption. The stored blood with low values of erythrocyte 2,3-DPG can be used without hesitation when correcting a chronic anemia for instance, but in acute situation, when the organism needs restoration of the oxygen releasing capacity within minutes, the resynthesis is obviously insufficient. In such situations, fresh blood or blood with a near normal 2,3-DPG

A novel bispecific antibody, S-Fab, induces potent cancer cell killing.

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

2015-01-01

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

Red Blood Cell Count Automation Using Microscopic Hyperspectral Imaging Technology.

Science.gov (United States)

Li, Qingli; Zhou, Mei; Liu, Hongying; Wang, Yiting; Guo, Fangmin

2015-12-01

Red blood cell counts have been proven to be one of the most frequently performed blood tests and are valuable for early diagnosis of some diseases. This paper describes an automated red blood cell counting method based on microscopic hyperspectral imaging technology. Unlike the light microscopy-based red blood count methods, a combined spatial and spectral algorithm is proposed to identify red blood cells by integrating active contour models and automated two-dimensional k-means with spectral angle mapper algorithm. Experimental results show that the proposed algorithm has better performance than spatial based algorithm because the new algorithm can jointly use the spatial and spectral information of blood cells.

B7h-expressing dendritic cells and plasma B cells mediate distinct outcomes of ICOS costimulation in T cell-dependent antibody responses

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Larimore Kevin

2012-06-01

Full Text Available Abstract Background The ICOS-B7h costimulatory receptor-ligand pair is required for germinal center formation, the production of isotype-switched antibodies, and antibody affinity maturation in response to T cell-dependent antigens. However, the potentially distinct roles of regulated B7h expression on B cells and dendritic cells in T cell-dependent antibody responses have not been defined. Results We generated transgenic mice with lineage-restricted B7h expression to assess the cell-type specific roles of B7h expression on B cells and dendritic cells in regulating T cell-dependent antibody responses. Our results show that endogenous B7h expression is reduced on B cells after activation in vitro and is also reduced in vivo on antibody-secreting plasma B cells in comparison to both naïve and germinal center B cells from which they are derived. Increasing the level of B7h expression on activated and plasma B cells in B-B7hTg mice led to an increase in the number of antibody-secreting plasma cells generated after immunization and a corresponding increase in the concentration of antigen-specific high affinity serum IgG antibodies of all isotypes, without affecting the number of responding germinal center B cells. In contrast, ICOS costimulation mediated by dendritic cells in DC-B7hTg mice contributed to germinal center formation and selectively increased IgG2a production without affecting the overall magnitude of antibody responses. Conclusions Using transgenic mice with lineage-restricted B7h expression, we have revealed distinct roles of ICOS costimulation mediated by dendritic cells and B cells in the regulation of T cell-dependent antibody responses.

Utilization and quality of cryopreserved red blood cells in transfusion medicine

NARCIS (Netherlands)

Henkelman, S.; Noorman, F.; Badloe, J. F.; Lagerberg, J. W. M.

Cryopreserved (frozen) red blood cells have been used in transfusion medicine since the Vietnam war. The main method to freeze the red blood cells is by usage of glycerol. Although the usage of cryopreserved red blood cells was promising due to the prolonged storage time and the limited cellular

Seventy-five genetic loci influencing the human red blood cell.

Science.gov (United States)

van der Harst, Pim; Zhang, Weihua; Mateo Leach, Irene; Rendon, Augusto; Verweij, Niek; Sehmi, Joban; Paul, Dirk S; Elling, Ulrich; Allayee, Hooman; Li, Xinzhong; Radhakrishnan, Aparna; Tan, Sian-Tsung; Voss, Katrin; Weichenberger, Christian X; Albers, Cornelis A; Al-Hussani, Abtehale; Asselbergs, Folkert W; Ciullo, Marina; Danjou, Fabrice; Dina, Christian; Esko, Tõnu; Evans, David M; Franke, Lude; Gögele, Martin; Hartiala, Jaana; Hersch, Micha; Holm, Hilma; Hottenga, Jouke-Jan; Kanoni, Stavroula; Kleber, Marcus E; Lagou, Vasiliki; Langenberg, Claudia; Lopez, Lorna M; Lyytikäinen, Leo-Pekka; Melander, Olle; Murgia, Federico; Nolte, Ilja M; O'Reilly, Paul F; Padmanabhan, Sandosh; Parsa, Afshin; Pirastu, Nicola; Porcu, Eleonora; Portas, Laura; Prokopenko, Inga; Ried, Janina S; Shin, So-Youn; Tang, Clara S; Teumer, Alexander; Traglia, Michela; Ulivi, Sheila; Westra, Harm-Jan; Yang, Jian; Zhao, Jing Hua; Anni, Franco; Abdellaoui, Abdel; Attwood, Antony; Balkau, Beverley; Bandinelli, Stefania; Bastardot, François; Benyamin, Beben; Boehm, Bernhard O; Cookson, William O; Das, Debashish; de Bakker, Paul I W; de Boer, Rudolf A; de Geus, Eco J C; de Moor, Marleen H; Dimitriou, Maria; Domingues, Francisco S; Döring, Angela; Engström, Gunnar; Eyjolfsson, Gudmundur Ingi; Ferrucci, Luigi; Fischer, Krista; Galanello, Renzo; Garner, Stephen F; Genser, Bernd; Gibson, Quince D; Girotto, Giorgia; Gudbjartsson, Daniel Fannar; Harris, Sarah E; Hartikainen, Anna-Liisa; Hastie, Claire E; Hedblad, Bo; Illig, Thomas; Jolley, Jennifer; Kähönen, Mika; Kema, Ido P; Kemp, John P; Liang, Liming; Lloyd-Jones, Heather; Loos, Ruth J F; Meacham, Stuart; Medland, Sarah E; Meisinger, Christa; Memari, Yasin; Mihailov, Evelin; Miller, Kathy; Moffatt, Miriam F; Nauck, Matthias; Novatchkova, Maria; Nutile, Teresa; Olafsson, Isleifur; Onundarson, Pall T; Parracciani, Debora; Penninx, Brenda W; Perseu, Lucia; Piga, Antonio; Pistis, Giorgio; Pouta, Anneli; Puc, Ursula; Raitakari, Olli; Ring, Susan M; Robino, Antonietta; Ruggiero, Daniela; Ruokonen, Aimo; Saint-Pierre, Aude; Sala, Cinzia; Salumets, Andres; Sambrook, Jennifer; Schepers, Hein; Schmidt, Carsten Oliver; Silljé, Herman H W; Sladek, Rob; Smit, Johannes H; Starr, John M; Stephens, Jonathan; Sulem, Patrick; Tanaka, Toshiko; Thorsteinsdottir, Unnur; Tragante, Vinicius; van Gilst, Wiek H; van Pelt, L Joost; van Veldhuisen, Dirk J; Völker, Uwe; Whitfield, John B; Willemsen, Gonneke; Winkelmann, Bernhard R; Wirnsberger, Gerald; Algra, Ale; Cucca, Francesco; d'Adamo, Adamo Pio; Danesh, John; Deary, Ian J; Dominiczak, Anna F; Elliott, Paul; Fortina, Paolo; Froguel, Philippe; Gasparini, Paolo; Greinacher, Andreas; Hazen, Stanley L; Jarvelin, Marjo-Riitta; Khaw, Kay Tee; Lehtimäki, Terho; Maerz, Winfried; Martin, Nicholas G; Metspalu, Andres; Mitchell, Braxton D; Montgomery, Grant W; Moore, Carmel; Navis, Gerjan; Pirastu, Mario; Pramstaller, Peter P; Ramirez-Solis, Ramiro; Schadt, Eric; Scott, James; Shuldiner, Alan R; Smith, George Davey; Smith, J Gustav; Snieder, Harold; Sorice, Rossella; Spector, Tim D; Stefansson, Kari; Stumvoll, Michael; Tang, W H Wilson; Toniolo, Daniela; Tönjes, Anke; Visscher, Peter M; Vollenweider, Peter; Wareham, Nicholas J; Wolffenbuttel, Bruce H R; Boomsma, Dorret I; Beckmann, Jacques S; Dedoussis, George V; Deloukas, Panos; Ferreira, Manuel A; Sanna, Serena; Uda, Manuela; Hicks, Andrew A; Penninger, Josef Martin; Gieger, Christian; Kooner, Jaspal S; Ouwehand, Willem H; Soranzo, Nicole; Chambers, John C

2012-12-20

Anaemia is a chief determinant of global ill health, contributing to cognitive impairment, growth retardation and impaired physical capacity. To understand further the genetic factors influencing red blood cells, we carried out a genome-wide association study of haemoglobin concentration and related parameters in up to 135,367 individuals. Here we identify 75 independent genetic loci associated with one or more red blood cell phenotypes at P < 10(-8), which together explain 4-9% of the phenotypic variance per trait. Using expression quantitative trait loci and bioinformatic strategies, we identify 121 candidate genes enriched in functions relevant to red blood cell biology. The candidate genes are expressed preferentially in red blood cell precursors, and 43 have haematopoietic phenotypes in Mus musculus or Drosophila melanogaster. Through open-chromatin and coding-variant analyses we identify potential causal genetic variants at 41 loci. Our findings provide extensive new insights into genetic mechanisms and biological pathways controlling red blood cell formation and function.

Red cell survival and sequestration in acute intermittent porphyria

International Nuclear Information System (INIS)

Nawalkha, P.L.; Soni, S.G.; Agrawal, V.K.; Misra, S.N.

1980-01-01

Life span and sequestration of red cells have been studied in twenty one proved cases of acute intermittent porphyria of different age and sex group from Bikaner District, Rajasthan State (India). Chromium-51 labelled red cells were used in the study and the excess count method of Bughe Jones and Szur was used to calculate the index of sequestration. The mean apparent half survival time of erythrocytes in the control subjects was 25.9 +- 2.9 (S.D.) days and the same in the prophyria patients was 27.0 +- 3.8 days. This shows that the life span of red cells is normal in both the patient and the control. Excess destruction of red blood cells was found to take place in either spleen or liver in the disease and no excess accumulation of erythrocytes occurred over spleen as compared to liver. (M.G.B.)

Modeling single cell antibody excretion on a biosensor

NARCIS (Netherlands)

Stojanovic, Ivan; Baumgartner, W.; van der Velden, T.J.G.; Terstappen, Leonardus Wendelinus Mathias Marie; Schasfoort, Richardus B.M.

2016-01-01

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

Red Cell Alloimmunization to Rhesus Antigen Among Pregnant Women Attending a Tertiary Care Hospital in Oman

Directory of Open Access Journals (Sweden)

Tamima Al-Dughaishi

2016-01-01

Full Text Available Objectives: The detection of maternal alloimmunization against red cell antigens is vital in the management of hemolytic disease of the fetus and newborn. We sought to measure the presence of allosensitization to Rhesus D (RhD antibodies in antenatal women attending a tertiary care hospital and assess the fetal outcome in sensitized women. Methods: We conducted a retrospective review of pregnant Omani women who registered at the Sultan Qaboos University Hospital between June 2011 and June 2013. Pregnant women were tested for ABO blood type and were screened for RhD antigen and antibodies at their first antenatal clinic visit. In women who tested positive for the RhD antibodies, an antibody titer was performed to evaluate the severity of their case. Results: Data was available on 1,251 pregnant women who were managed and delivered at Sultan Qaboos University Hospital. The prevalence of RhD negative pregnant women was 7.3%. Blood group O was the most common followed by A, B, and AB. The rate of RhD negative alloimmunization was 10%, and anti-D was the most common antibody detected. There were no stillbirths or neonatal deaths. Postnatal transfusion was necessary for only one baby. Conclusions: The prevalence of RhD negativity was comparable to other Asian countries. Previous RhD alloimmunization and history of miscarriages were the most common maternal medical history.

Induction of CD4 suppressor T cells with anti-Leu-8 antibody

International Nuclear Information System (INIS)

Kanof, M.E.; Strober, W.; James, S.P.

1987-01-01

To characterize the conditions under which CD4 T cells suppress polyclonal immunoglobulin synthesis, we investigated the capacity of CD4 T cells that coexpress the surface antigen recognized by the monoclonal antibody anti-Leu-8 to mediate suppression. In an in vitro system devoid of CD8 T cells, CD4, Leu-8+ T cells suppressed pokeweed mitogen-induced immunoglobulin synthesis. Similarly, suppressor function was induced in unfractionated CD4 T cell populations after incubation with anti-Leu-8 antibody under cross-linking conditions. This induction of suppressor function by anti-Leu-8 antibody was not due to expansion of the CD4, Leu-8+ T cell population because CD4 T cells did not proliferate in response to anti-Leu-8 antibody. However, CD4, Leu-8+ T cell-mediated suppression was radiosensitive. Finally, CD4, Leu-8+ T cells do not inhibit immunoglobulin synthesis when T cell lymphokines were used in place of helper CD4 T cells (CD4, Leu-8- T cells), suggesting that CD4 T cell-mediated suppression occurs at the T cell level. We conclude that CD4 T cells can be induced to suppress immunoglobulin synthesis by modulation of the membrane antigen recognized by anti-Leu-8 antibody

Radioimmunoassay for antibodies against surface membrane antigens using adhering cells

Energy Technology Data Exchange (ETDEWEB)

Tax, A; Manson, L A [Wistar Inst. of Anatomy and Biology, Philadelphia, Pa. (USA)

1976-07-01

A radioimmunoassay using cells adhering to plastic is described. In this assay, A-10 mammary carcinoma attached to the surface of plastic in microtiter plates were permitted to bind antibody and the bound antibody was detected with purified rabbit /sup 125/I-antimouse-Fab. The bound radioactive material was eluted with glycine-HCl buffer (pH 2.5), and the acid eluates were counted in a gamma counter. This assay can be used to detect cytolic or noncytolic antibody to cell surface antigens in studies with any tumor or normal cell that will adhere to a solid surface.

Erythrokinetics, ferrokinetics and red cell survival in sickle cell anaemia under subtropical climatic conditions

International Nuclear Information System (INIS)

Cardenas, R.

1975-01-01

Ferrokinetic parameters were evaluated with 59 Fe and red-cell survival with 51 Cr by classical techniques in a total of 17 patients with sickle-cell disease. The mean plasma 59 Fe half-disappearance time in these patients was 29.5 min as compared with a normal value of 92 min, and the t1/2 51 Cr 8.0 days as compared with a normal value of 26.0 days. The mean red-cell iron turnover rate was elevated to 9 times normal. The increased destruction of red cells appeared to take place predominantly, though not entirely, in the spleen. Eight of the 17 patients studied were identified as having intercurrent complications, but these did not significantly affect the results of the investigations. A group of 5 boys in whom the red-cell iron turnover rate was elevated to a lesser degree than in the other patients were subjected to more detailed studies of plasma 59 Fe clearance with particular reference to ineffective erythropoiesis. In these patients, the plasma 59 Fe clearance curves showed precocious humps characteristic of ineffective erythropoiesis. Detailed analysis of the results indicated ineffective erythropoiesis corresponding to 3.6, 16.0, 22.6, 32.0 and 50.0 % of the iron initially taken up by the bone marrow. It is concluded that while the anaemia in most patients with sickle-cell disease is mainly due to shortened survival of the circulating red cells, with increased destruction of red cells in the spleen, ineffective erythropoiesis may none the less be an important factor determining the actual degree of this anaemia

CD47 limits antibody dependent phagocytosis against non-malignant B cells.

Science.gov (United States)

Gallagher, Sandra; Turman, Sean; Lekstrom, Kristen; Wilson, Susan; Herbst, Ronald; Wang, Yue

2017-05-01

Recent studies have demonstrated the importance of CD47 in protecting malignant B cells from antibody dependent cellular phagocytosis (ADCP). Combined treatment of anti-CD47 and -CD20 antibodies synergistically augment elimination of tumor B cells in xenograft mouse models. This has led to the development of novel reagents that can potentially enhance killing of malignant B cells in patients. B cell depleting therapy is also a promising treatment for autoimmune patients. In the current study, we aimed to investigate whether or not CD47 protects non-malignant B cells from ADCP. We show that CD47 is expressed on all B cells in mice, with the highest level on plasma cells in bone marrow and spleen. Although its expression is dispensable for B cell development in mice, CD47 on B cells limits antibody mediated phagocytosis. B cell depletion following in vivo anti-CD19 treatment is more efficient in CD47-/- mice than in wild type mice. In vitro, both naïve and activated B cells from CD47-/- mice are more sensitive to ADCP than wild type B cells. Lastly, we show in an ADCP assay that blocking CD47 can enhance anti-CD19 antibody mediated phagocytosis of wild type B cells. These results suggest that in addition to its already demonstrated benefit in cancer, targeting CD47 may be used as an adjunct in combination with B cell depletion antibodies for treatment of autoimmune diseases. Copyright © 2017 Elsevier Ltd. All rights reserved.

Red Dot Basal Cell Carcinoma: An Unusual Variant of a Common Malignancy.

Science.gov (United States)

Loh, Tiffany Y; Cohen, Philip R

2016-05-01

Red dot basal cell carcinoma is a distinct but rare subtype of basal cell carcinoma (BCC). It presents as a red macule or papule; therefore, in most cases, it may easily be mistaken for a benign vascular lesion, such as a telangiectasia or angioma. A red dot BCC in an older woman is described. Clinical and histological differences between red dot BCCs and telangiectasias are described. A 72-year-old woman initially presented with a painless red macule on her nose. Biopsy of the lesion established the diagnosis of a red dot BCC. Pubmed was searched for the following terms: angioma, basal cell carcinoma, dermoscope, diascopy, red dot, non-melanoma skin cancer, telangiectasia, and vascular. The papers were reviewed for cases of red dot basal cell carcinoma. Clinical and histological characteristics of red dot basal cell carcinoma and telangiectasias were compared. Red dot BCC is an extremely rare variant of BCC that may be confused with benign vascular lesions. Although BCCs rarely metastasize and are associated with low mortality, they have the potential to become locally invasive and destructive if left untreated. Thus, a high index of suspicion for red dot BCC is necessary. J Drugs Dermatol. 2016;15(5):645-647.

Reflection coefficients of permeant molecules in human red cell suspensions.

Science.gov (United States)

Owen, J D; Eyring, E M

1975-08-01

The Staverman reflection coefficient, sigma for several permeant molecules was determined in human red cell suspensions with a Durrum stopped-flow spectrophotometer. This procedure was first used with dog, cat, and beef red cells and with human red cells. The stopped-flow technique used was similar to the rapid-flow method used by those who originally reported sigma measurements in human red cells for molecules which rapidly penetrate the red cell membrane. The sigma values we obtained agreed with those previously reported for most of the slow penetrants, except malonamide, but disagreed with all the sigma values previously reported for the rapid penetrants. We were unable to calculate an "equivalent pore radius" with our sigma data. The advantages of our equipment and our experimental procedure are discussed. Our sigma data suggest that sigma is indirectly proportional to the log of the nonelectrolyte permeability coefficient, omega. Since a similar trend has been previously shown for log omega and molar volume of the permeant molecules, a correlatioo was shown between sigma and molar volume suggesting the membrane acts as a sieve.

Direct binding of radioiodinated monoclonal antibody to tumor cells: significance of antibody purity and affinity for drug targeting or tumor imaging

International Nuclear Information System (INIS)

Kennel, S.J.; Foote, L.J.; Lankford, P.K.; Johnson, M.; Mitchell, T.; Braslawsky, G.R.

1983-01-01

For MoAb to be used efficiently for drug targeting and tumor imaging, the fraction of antibody binding to tumor cells must be maximized. The authors have studied the binding of 125 I MoAb in three different tumor systems. The fraction of antibody that could be bound to the cell surface was directly proportional to the antibody purity. The affinity constant also limits the fraction of antibody that can bind to cells at a given antigen concentration. Rearrangement of the standard expression for univalent equilibrium binding between two reactants shows that in antigen excess, the maximum fraction of antibody that can bind =Ka[Ag total]/1 + Ka[Ag total]. Binding data using four different MoAb with three cell systems confirm this relationship. Estimates for reasonable concentrations of tumor antigens in vivo indicate that antibodies with binding constants less than 10 8 M -1 are not likely to be useful for drug targeting or tumor imaging

The absolute counting of red cell-derived microparticles with red cell bead by flow rate based assay.

Science.gov (United States)

Nantakomol, Duangdao; Imwong, Malika; Soontarawirat, Ingfar; Kotjanya, Duangporn; Khakhai, Chulalak; Ohashi, Jun; Nuchnoi, Pornlada

2009-05-01

Activation of red blood cell is associated with the formation of red cell-derived microparticles (RMPs). Analysis of circulating RMPs is becoming more refined and clinically useful. A quantitative Trucount tube method is the conventional method uses for quantitating RMPs. In this study, we validated a quantitative method called "flow rate based assay using red cell bead (FCB)" to measure circulating RMPs in the peripheral blood of healthy subjects. Citrated blood samples collected from 30 cases of healthy subjects were determined the RMPs count by using double labeling of annexin V-FITC and anti-glycophorin A-PE. The absolute RMPs numbers were measured by FCB, and the results were compared with the Trucount or with flow rate based calibration (FR). Statistical correlation and agreement were analyzed using linear regression and Bland-Altman analysis. There was no significant difference in the absolute number of RMPs quantitated by FCB when compared with those two reference methods including the Trucount tube and FR method. The absolute RMPs count obtained from FCB method was highly correlated with those obtained from Trucount tube (r(2) = 0.98, mean bias 4 cell/microl, limit of agreement [LOA] -20.3 to 28.3 cell/microl), and FR method (r(2) = 1, mean bias 10.3 cell/microl, and LOA -5.5 to 26.2 cell/microl). This study demonstrates that FCB is suitable and more affordable for RMPs quantitation in the clinical samples. This method is a low cost and interchangeable to latex bead-based method for generating the absolute counts in the resource-limited areas. (c) 2008 Clinical Cytometry Society.

Antibodies and Selection of Monoclonal Antibodies.

Science.gov (United States)

Hanack, Katja; Messerschmidt, Katrin; Listek, Martin

Monoclonal antibodies are universal binding molecules with a high specificity for their target and are indispensable tools in research, diagnostics and therapy. The biotechnological generation of monoclonal antibodies was enabled by the hybridoma technology published in 1975 by Köhler and Milstein. Today monoclonal antibodies are used in a variety of applications as flow cytometry, magnetic cell sorting, immunoassays or therapeutic approaches. First step of the generation process is the immunization of the organism with appropriate antigen. After a positive immune response the spleen cells are isolated and fused with myeloma cells in order to generate stable, long-living antibody-producing cell lines - hybridoma cells. In the subsequent identification step the culture supernatants of all hybridoma cells are screened weekly for the production of the antibody of interest. Hybridoma cells producing the antibody of interest are cloned by limited dilution till a monoclonal hybridoma is found. This is a very time-consuming and laborious process and therefore different selection strategies were developed since 1975 in order to facilitate the generation of monoclonal antibodies. Apart from common automation of pipetting processes and ELISA testing there are some promising approaches to select the right monoclonal antibody very early in the process to reduce time and effort of the generation. In this chapter different selection strategies for antibody-producing hybridoma cells are presented and analysed regarding to their benefits compared to conventional limited dilution technology.

Restrictive versus liberal transfusion strategy for red blood cell transfusion

DEFF Research Database (Denmark)

Holst, Lars B; Petersen, Marie W; Haase, Nicolai

2015-01-01

OBJECTIVE: To compare the benefit and harm of restrictive versus liberal transfusion strategies to guide red blood cell transfusions. DESIGN: Systematic review with meta-analyses and trial sequential analyses of randomised clinical trials. DATA SOURCES: Cochrane central register of controlled...... differences with 95% confidence intervals. RESULTS: 31 trials totalling 9813 randomised patients were included. The proportion of patients receiving red blood cells (relative risk 0.54, 95% confidence interval 0.47 to 0.63, 8923 patients, 24 trials) and the number of red blood cell units transfused (mean...... were associated with a reduction in the number of red blood cell units transfused and number of patients being transfused, but mortality, overall morbidity, and myocardial infarction seemed to be unaltered. Restrictive transfusion strategies are safe in most clinical settings. Liberal transfusion...

Detection of microparticles from human red blood cells by multiparametric flow cytometry

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Grisendi, Giulia; Finetti, Elena; Manganaro, Daniele; Cordova, Nicoletta; Montagnani, Giuliano; Spano, Carlotta; Prapa, Malvina; Guarneri, Valentina; Otsuru, Satoru; Horwitz, Edwin M.; Mari, Giorgio; Dominici, Massimo

2015-01-01

Background During storage, red blood cells (RBC) undergo chemical and biochemical changes referred to as “storage lesions”. These events determine the loss of RBC integrity, resulting in lysis and release of microparticles. There is growing evidence of the clinical importance of microparticles and their role in blood transfusion-related side effects and pathogen transmission. Flow cytometry is currently one of the most common techniques used to quantify and characterise microparticles. Here we propose multiparametric staining to monitor and quantify the dynamic release of microparticles by stored human RBC. Material and methods RBC units (n=10) were stored under blood bank conditions for up to 42 days. Samples were tested at different time points to detect microparticles and determine the haemolysis rate (HR%). Microparticles were identified by flow cytometry combining carboxyfluorescein diacetate succinimidyl ester (CFSE) dye, annexin V and anti-glycophorin A antibody. Results We demonstrated that CFSE can be successfully used to label closed vesicles with an intact membrane. The combination of CFSE and glycophorin A antibody was effective for monitoring and quantifying the dynamic release of microparticles from RBC during storage. Double staining with CFSE/glycophorin A was a more precise approach, increasing vesicle detection up to 4.7-fold vs the use of glycophorin A/annexin V alone. Moreover, at all the time points tested, we found a robust correlation (R=0.625; p=0.0001) between HR% and number of microparticles detected. Discussion Multiparametric staining, based on a combination of CFSE, glycophorin A antibody and annexin V, was able to detect, characterise and monitor the release of microparticles from RBC units during storage, providing a sensitive approach to labelling and identifying microparticles for transfusion medicine and, more broadly, for cell-based therapies. PMID:25369588

Synthesis and Characterization of Anti-HER2 Antibody Conjugated CdSe/CdZnS Quantum Dots for Fluorescence Imaging of Breast Cancer Cells

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Takashi Jin

2009-11-01

Full Text Available The early detection of HER2 (human epidermal growth factor receptor 2 status in breast cancer patients is very important for the effective implementation of anti-HER2 antibody therapy. Recently, HER2 detections using antibody conjugated quantum dots (QDs have attracted much attention. QDs are a new class of fluorescent materials that have superior properties such as high brightness, high resistance to photo-bleaching, and multi-colored emission by a single-light source excitation. In this study, we synthesized three types of anti-HER2 antibody conjugated QDs (HER2Ab-QDs using different coupling agents (EDC/sulfo-NHS, iminothiolane/sulfo-SMCC, and sulfo-SMCC. As water-soluble QDs for the conjugation of antibody, we used glutathione coated CdSe/CdZnS QDs (GSH-QDs with fluorescence quantum yields of 0.23~0.39 in aqueous solution. Dispersibility, hydrodynamic size, and apparent molecular weights of the GSH-QDs and HER2Ab-QDs were characterized by using dynamic light scattering, fluorescence correlation spectroscopy, atomic force microscope, and size-exclusion HPLC. Fluorescence imaging of HER2 overexpressing cells (KPL-4 human breast cancer cell line was performed by using HER2Ab-QDs as fluorescent probes. We found that the HER2Ab-QD prepared by using SMCC coupling with partially reduced antibody is a most effective probe for the detection of HER2 expression in KPL-4 cells. We have also studied the size dependency of HER2Ab-QDs (with green, orange, and red emission on the fluorescence image of KPL-4 cells.

[Establishment and identification of mouse lymphoma cell line EL4 expressing red fluorescent protein].

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Li, Yan-Jie; Cao, Jiang; Chen, Chong; Wang, Dong-Yang; Zeng, Ling-Yu; Pan, Xiu-Ying; Xu, Kai-Lin

2010-02-01

This study was purposed to construct a lentiviral vector encoding red fluorescent protein (DsRed) and transfect DsRed into EL4 cells for establishing mouse leukemia/lymphoma model expressing DsRed. The bicistronic SIN lentiviral transfer plasmid containing the genes encoding neo and internal ribosomal entry site-red fluorescent protein (IRES-DsRed) was constructed. Human embryonic kidney 293FT cells were co-transfected with the three plasmids by liposome method. The viral particles were collected and used to transfect EL4 cells, then the cells were selected by G418. The results showed that the plasmid pXZ208-neo-IRES-DsRed was constructed successfully, and the viral titer reached to 10(6) U/ml. EL4 cells were transfected by the viral solution efficiently. The transfected EL4 cells expressing DsRed survived in the final concentration 600 microg/ml of G418. The expression of DsRed in the transfected EL4 cells was demonstrated by fluorescence microscopy and flow cytometry. In conclusion, the EL4/DsRed cell line was established successfully.

21 CFR 864.7100 - Red blood cell enzyme assay.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Red blood cell enzyme assay. 864.7100 Section 864.7100 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7100 Red blood cell...

Comparative study on antibody immobilization strategies for efficient circulating tumor cell capture.

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Ates, Hatice Ceren; Ozgur, Ebru; Kulah, Haluk

2018-03-23

Methods for isolation and quantification of circulating tumor cells (CTCs) are attracting more attention every day, as the data for their unprecedented clinical utility continue to grow. However, the challenge is that CTCs are extremely rare (as low as 1 in a billion of blood cells) and a highly sensitive and specific technology is required to isolate CTCs from blood cells. Methods utilizing microfluidic systems for immunoaffinity-based CTC capture are preferred, especially when purity is the prime requirement. However, antibody immobilization strategy significantly affects the efficiency of such systems. In this study, two covalent and two bioaffinity antibody immobilization methods were assessed with respect to their CTC capture efficiency and selectivity, using an anti-epithelial cell adhesion molecule (EpCAM) as the capture antibody. Surface functionalization was realized on plain SiO 2 surfaces, as well as in microfluidic channels. Surfaces functionalized with different antibody immobilization methods are physically and chemically characterized at each step of functionalization. MCF-7 breast cancer and CCRF-CEM acute lymphoblastic leukemia cell lines were used as EpCAM positive and negative cell models, respectively, to assess CTC capture efficiency and selectivity. Comparisons reveal that bioaffinity based antibody immobilization involving streptavidin attachment with glutaraldehyde linker gave the highest cell capture efficiency. On the other hand, a covalent antibody immobilization method involving direct antibody binding by N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC)-N-hydroxysuccinimide (NHS) reaction was found to be more time and cost efficient with a similar cell capture efficiency. All methods provided very high selectivity for CTCs with EpCAM expression. It was also demonstrated that antibody immobilization via EDC-NHS reaction in a microfluidic channel leads to high capture efficiency and selectivity.

Antibody and B cell responses to Plasmodium sporozoites

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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.

Enzymes and membrane proteins of ADSOL-preserved red blood cells

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Maria Sueli Soares Leonart

2000-03-01

Full Text Available CONTEXT: The preservative solution ADSOL (adenine, dextrose, sorbitol, sodium chloride and mannitol maintains red cell viability for blood trans-fusion for 6 weeks. It would be useful to know about its preservation qualities over longer periods. OBJECTIVE: To determine some red cell biochemical parameters for peri-ods of up to 14 weeks in order to determine whether the red cell metabo-lism integrity would justify further studies aiming at increasing red cell preservation and viability. DESIGN: Biochemical evaluation designed to study red cell preservation. SETTING: São Paulo University erythrocyte metabolism referral center. SAMPLE: Six normal blood donors from the University Hospital of the Universidade Federal do Paraná, Curitiba, Brazil. MAIN MEASUREMENTS: Weekly assay of erythrocyte adenosine-5´-triphosphate (ATP, 2,3-diphosphoglycerate (2,3DPG, hexokinase (HX, phosphofructokinase (PFK, pyruvate kinase (PK, glucose-6-phosphate dehydrogenase (G-6-PD, 6-phosphogluconic dehydrogenase (6-PGD, glyceraldehyde-3-phosphate dehydrogenase (GAPD, glutathione reduc-tase (GR, glutathione peroxidase (GSHPx, plasma sodium and potas-sium, blood pH, and membrane proteins of red cells preserved in ADSOL were studied during storage for 14 weeks storage. RESULTS: During ADSOL preservation, erythrocyte ATP concentration decreased 60% after 5 weeks, and 90% after 10 weeks; the pH fell from 6.8 to 6.4 by the 14th week. 2,3-DPG concentration was stable during the first week, but fell 90% after 3 weeks and was exhausted after 5 weeks. By the end of the 5th week, an activity decrease of 16-30% for Hx, GAPD, GR, G-6-PD and 6-PGD, 35% for PFK and GSHPx, and 45% for PK were observed. Thereafter, a uniform 10% decay was observed for all enzymes up to the 14th week. The red blood cell membrane pro-teins did not show significant alterations in polyacrylamide gel electro-phoresis (SDS-PAGE during the 14 weeks. CONCLUSION: Although the blood viability was shown to be poor

[Red Blood Cells Raman Spectroscopy Comparison of Type Two Diabetes Patients and Rats].

Science.gov (United States)

Wang, Lei; Liu, Gui-dong; Mu, Xin; Xiao, Hong-bin; Qi, Chao; Zhang, Si-qi; Niu Wen-ying; Jiang, Guang-kun; Feng, Yue-nan; Bian, Jing-qi

2015-10-01

By using confocal Raman spectroscopy, Raman spectra were measured in normal rat red blood cells, normal human red blood cells, STZ induced diabetetic rats red blood cells, Alloxan induced diabetetic rats red blood cells and human type 2 diabetes red blood cells. Then principal component analysis (PCA) with support vector machine (SVM) classifier was used for data analysis, and then the distance between classes was used to judge the degree of close to two kinds of rat model with type 2 diabetes. The results found significant differences in the Raman spectra of red blood cell in diabetic and normal red blood cells. To diabetic red blood cells, the peak in the amide VI C=O deformation vibration band is obvious, and amide V N-H deformation vibration band spectral lines appear deviation. Belong to phospholipid fatty acyl C-C skeleton, the 1 130 cm(-1) spectral line is enhanced and the 1 088 cm(-1) spectral line is abated, which show diabetes red cell membrane permeability increased. Raman spectra of PCA combined with SVM can well separate 5 types of red blood cells. Classifier test results show that the classification accuracy is up to 100%. Through the class distance between the two induced method and human type 2 diabetes, it is found that STZ induced model is more close to human type 2 diabetes. In conclusion, Raman spectroscopy can be used for diagnosis of diabetes and rats STZ induced diabetes method is closer to human type 2 diabetes.

SMIM1 underlies the Vel blood group and influences red blood cell traits

DEFF Research Database (Denmark)

Cvejic, Ana; Haer-Wigman, Lonneke; Stephens, Jonathan C

2013-01-01

The blood group Vel was discovered 60 years ago, but the underlying gene is unknown. Individuals negative for the Vel antigen are rare and are required for the safe transfusion of patients with antibodies to Vel. To identify the responsible gene, we sequenced the exomes of five individuals negative...... and expression of the Vel antigen on SMIM1-transfected cells confirm SMIM1 as the gene underlying the Vel blood group. An expression quantitative trait locus (eQTL), the common SNP rs1175550 contributes to variable expression of the Vel antigen (P = 0.003) and influences the mean hemoglobin concentration of red...... blood cells (RBCs; P = 8.6 × 10(-15)). In vivo, zebrafish with smim1 knockdown showed a mild reduction in the number of RBCs, identifying SMIM1 as a new regulator of RBC formation. Our findings are of immediate relevance, as the homozygous presence of the deletion allows the unequivocal identification...

Avoiding Anemia: Boost Your Red Blood Cells

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... Issues Subscribe January 2014 Print this issue Avoiding Anemia Boost Your Red Blood Cells En español Send ... Disease When Blood Cells Bend Wise Choices Preventing Anemia To prevent or treat iron-deficiency anemia: Eat ...

Study on the Measurement of 51Cr-tagged Red Cell Survival: Reevaluation of its method and the effect of Blood loss on red cell survival with 51Cr

International Nuclear Information System (INIS)

Choi, Hak Yong; Koh, Chang Soon; Lee, Moon Ho

1970-01-01

Reappraisal measurement of apparent half survival time of red cell by 51 Cr method was made and effects of blood-letting over red cell survival were observed. The study was performed on 53 normal male subjects under three different experimental conditions. 1) Group 1: Mean 51 Cr red cell half survival by ACD wash method was 29.7 days. T 1 /2 of Ascorbic acid method was 29.0 days in group with 100 mg dose and 29.1 days in group with 50 mg dose respectively. There was no difference between these two methods in regards to red cell half survival. No difference were noted in amount of ascorbic acid administered. 2) Group 2: As daily amount of blood loss in increased the shortening of red cell half survival was noted. Rapid phase was seen when blood loss ranged 10 to 25 ml per day, while slow phase noted when more loss amounted 25 ml more daily. Thus, it was clear that there was more than an exponential relation between T 1 /2 and the amount of blood loss. 3) Group 3: T 1 /2 measured cpm per whole blood was within normal range and T 1 /2 measured by cpm per red mass showed shortening tendency when compared with the former in the group measured after blood loss (from 25 ml daily up to 100 ml daily in 10 days). In the group with rather constant blood loss of 100 ml daily for 10 consecutive days revealed the significant difference in two measurement (P 1 /2 in non-steady state. When red cell production is increased compared with red cell destruction, T 1 /2 measured by cpm per red cell mass shorter than that by cpm per whole blood. Shortening of T 1 /2 measured by cpm per whole blood is more prominent, if red destruction is enhanced and exceeds production. 5) It is clear that when expressing red cell destruction rate, T 1 /2 measured by cpm per whole blood is more adequate and production more consistent with cpm red cell mass. 6) T 1 /2 measured during blood-letting, when corrected by amount of blood loss, it remains normal. It is erroneous to use conventional equational

Volume-dependent K+ transport in rabbit red blood cells comparison with oxygenated human SS cells

Energy Technology Data Exchange (ETDEWEB)

Al-Rohil, N.; Jennings, M.L.

1989-07-01

In this study the volume-dependent or N-ethylmaleimide (NEM)-stimulated, ouabain-insensitive K+ influx and efflux were measured with the tracer 86Rb+ in rabbit red blood cells. The purpose of the work was to examine the rabbit as a potential model for cell volume regulation in human SS red blood cells and also to investigate the relationship between the NEM-reactive sulfhydryl group(s) and the signal by which cell swelling activates the transport. Ouabain-resistant K+ efflux and influx increase nearly threefold in cells swollen hypotonically by 15%. Pretreatment with 2 mM NEM stimulates efflux 5-fold and influx 10-fold (each measured in an isotonic medium). The ouabain-resistant K+ efflux was dependent on the major anion in the medium. The anion dependence of K+ efflux in swollen or NEM-stimulated cells was as follows: Br- greater than Cl- much greater than NO3- = acetate. The magnitudes of both the swelling- and the NEM-stimulated fluxes are much higher in young cells (density separated but excluding reticulocytes) than in older cells. Swelling- or NEM-stimulated K+ efflux in rabbit red blood cells was inhibited 50% by 1 mM furosemide, and the inhibitory potency of furosemide was enhanced by extracellular K+, as is known to be true for human AA and low-K+ sheep red blood cells. The swelling-stimulated flux in both rabbit and human SS cells has a pH optimum at approximately 7.4. We conclude that rabbit red blood cells are a good model for swelling-stimulated K+ transport in human SS cells.

Regulation of antibody synthesis in the X-irradiated Mexican axolotl

International Nuclear Information System (INIS)

Charlemagne, J.

1981-01-01

The effects of X-irradiation were studied on the Mexican axolotl antibody synthesis. To reduce the anti-horse red blood cell (HRBC) antibody titers, 150 rd and smaller doses are ineffective, 200-450 rd are increasingly effective, and 700 rd are maximally effective (and lethal). A significant enhancement of the anti-HRBC titers was observed in low doses (50-150 rd X-irradiated animals). This enhancement was also observed when a low X-ray dose was applied only on the thymic areas. In whole body, but thymus area-shielded, 100 rd X-irradiated animals, the anti-HRBC titers were similar to those of the nonirradiated, HRBC-immunized control group. To explain these phenomena, it is suggested that a radiosensitive, adult thymectomy-sensitive and hydrocortisone-sensitive suppressor T cell subpopulation regulates the antibody synthesis in the axolotl. (orig.) [de

Regulation of antibody synthesis in the X-irradiated Mexican axolotl

Energy Technology Data Exchange (ETDEWEB)

Charlemagne, J.

1981-09-01

The effects of X-irradiation were studied on the Mexican axolotl antibody synthesis. To reduce the anti-horse red blood cell (HRBC) antibody titers, 150 rd and smaller doses are ineffective, 200-450 rd are increasingly effective, and 700 rd are maximally effective (and lethal). A significant enhancement of the anti-HRBC titers was observed in low doses (50-150 rd X-irradiated animals). This enhancement was also observed when a low X-ray dose was applied only on the thymic areas. In whole body, but thymus area-shielded, 100 rd X-irradiated animals, the anti-HRBC titers were similar to those of the nonirradiated, HRBC-immunized control group. To explain these phenomena, it is suggested that a radiosensitive, adult thymectomy-sensitive and hydrocortisone-sensitive suppressor T cell subpopulation regulates the antibody synthesis in the axolotl.

Density increment and decreased survival of rat red blood cells induced by cadmium

International Nuclear Information System (INIS)

Kunimoto, M.; Miura, T.

1986-01-01

Male Wistar rats were injected with CdCl 2 subcutaneously to examine in vivo effects of Cd on density and survival of red blood cells. During the 7 days after administration of 1.0 mg Cd/kg, the following sequence of events occurred: (1) a progressive increase in the amount of more dense red blood cells concomitant with a decrease in that of light red blood cells from the first to the third day; (2) an increase in the spleen weight at the third day; (3) a decrease in the hematocrit value and an increase in the amount of light red blood cells at the fifth day; and (4) a recovery of the hematocrit value at the seventh day. Five days after administration, the hematocrit value decreased in a dose-dependent mode and the decrease was significant at the 1% level at 1.0 and 1.5 mg Cd/kg. A highly significant splenomegaly was also observed at 0.5 to 1.5 mg Cd/kg. In order to label red blood cells in vivo, [ 3 H] diisopropylfluorophosphate ([ 3 H]DFP) was injected into rats. At Day 11, Cd at either 0.5 or 1.0 mg/kg was administered to [ 3 H]DFP-prelabeled animals. Cd administration accelerated 3 H-labeled red cell clearance from the blood. Six days after Cd administration, the radioactivity of red blood cells was 76 and 68% of the control at 0.5 and 1.0 mg Cd/kg, respectively. In vitro treatment of rat red density and accelerated in vivo clearance of red blood cells from the recipient circulation. These results show that Cd at low dose can cause anemia by increasing red cell density and by accelerating red cell sequestration, presumably in the spleen

A radioimmunoassay for antibodies against surface membrane antigens using adhering cells

International Nuclear Information System (INIS)

Tax, A.; Manson, L.A.

1976-01-01

A radioimmunoassay using cells adhering to plastic is described. In this assay, A-10 mammary carcinoma attached to the surface of plastic in microtiter plates were permitted to bind antibody and the bound antibody was detected with purified rabbit 125 I-antimouse-Fab. The bound radioactive material was eluted with glycine-HCl buffer (pH 2.5), and the acid eluates were counted in a gamma counter. This assay can be used to detect cytolic or noncytolic antibody to cell surface antigens in studies with any tumor or normal cell that will adhere to a solid surface

Biosignatures of Kerala red rain cells: Implications in understanding their origin

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Gangappa, R.; Thomas, M.; Hogg, S.

2013-09-01

The red rain that fell over Kerala, southern India (2001-2012) was characterised by the red pigmented particles. Earlier proposal claiming that these are known algal bloom blown from trees (Sampath et al, 2001; DiGregorio, 2007) has been studied by us and disproved. Also, further investigation reporting their extraordinary properties including a suggestion that they lack DNA (Louis and Kumar 2003; 2006; 2008) has been invalidated (Gangappa and Hogg, 2013). However, their claim regarding the growth and replication of these cells at 300ºC needs more investigation if it is to gain acceptance. Current study provide evidences regarding the biological properties of Kerala red rain cells to gain insights into environmental conditions from which they may have originated. Combined with various research strategies and high resolution instruments, we have demonstrated the following interesting properties of Kerala red rain cells: (1) unusually thick external envelope enclosing the central core; (2)stability of red pigment at temperatures about 100ºC and pH variations; (3) absence of eukaryotic ultrastructures; (4) possible replication at 121ºC with nanostructures (possible daughter cells) having similar morphological features inside the large mother cells at such high temperature. They contain high percentage of carbon, iron, silicon and aluminum and often enclosed in a silicon rich biofilms. Further investigation shows that the positive detection of DNA in these cells was possible only after the complete removal of red pigment, thereby providing an explanation for the negative outcome of earlier studies in this regard. Moreover, evidences are shown to support that these cells contain high amounts of UV absorbing compounds, porphyrin complexes and possible scytonemin. Kerala red rain cells may prove to be polyextermophiles belonging to prokaryotes and may have possibly originated from the environment containing above mentioned chemical elements, high energy UV exposure and

Red Dot Basal Cell Carcinoma: Report of Cases and Review of This Unique Presentation of Basal Cell Carcinoma.

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Cohen, Philip R

2017-03-22

Red dot basal cell carcinoma is a unique variant of basal cell carcinoma. Including the three patients described in this report, red dot basal cell carcinoma has only been described in seven individuals. This paper describes the features of two males and one female with red dot basal cell carcinoma and reviews the characteristics of other patients with this clinical subtype of basal cell carcinoma. A 70-year-old male developed a pearly-colored papule with a red dot in the center on his nasal tip. A 71-year-old male developed a red dot surrounded by a flesh-colored papule on his left nostril. Lastly, a 74-year-old female developed a red dot within an area of erythema on her left mid back. Biopsy of the lesions all showed nodular and/or superficial basal cell carcinoma. Correlation of the clinical presentation and pathology established the diagnosis of red dot basal cell carcinoma. The tumors were treated by excision using the Mohs surgical technique. Pubmed was searched with the keyword: basal, cell, cancer, carcinoma, dot, red, and skin. The papers generated by the search and their references were reviewed. Red dot basal cell carcinoma has been described in three females and two males; the gender was not reported in two patients. The tumor was located on the nose (five patients), back (one patient) and thigh (one patient). Cancer presented as a solitary small red macule or papule; often, the carcinoma was surrounded by erythema or a flesh-colored papule. Although basal cell carcinomas usually do not blanch after a glass microscope slide is pressed against them, the red dot basal cell carcinoma blanched after diascopy in two of the patients, resulting in a delay of diagnosis in one of these individuals. Dermoscopy may be a useful non-invasive modality for evaluating skin lesions when the diagnosis of red dot basal cell carcinoma is considered. Mohs surgery is the treatment of choice; in some of the patients, the ratio of the area of the postoperative wound to that

Circulating red cell-derived microparticles in human malaria.

Science.gov (United States)

Nantakomol, Duangdao; Dondorp, Arjen M; Krudsood, Srivicha; Udomsangpetch, Rachanee; Pattanapanyasat, Kovit; Combes, Valery; Grau, Georges E; White, Nicholas J; Viriyavejakul, Parnpen; Day, Nicholas P J; Chotivanich, Kesinee

2011-03-01

In patients with falciparum malaria, plasma concentrations of cell-derived microparticles correlate with disease severity. Using flow cytometry, we quantified red blood cell-derived microparticles (RMPs) in patients with malaria and identified the source and the factors associated with production. RMP concentrations were increased in patients with Plasmodium falciparum (n = 29; median, 457 RMPs/μL [range, 13-4,342 RMPs/μL]), Plasmodium vivax (n = 5; median, 409 RMPs/μL [range, 281-503/μL]), and Plasmodium malariae (n = 2; median, 163 RMPs/μL [range, 127-200 RMPs/μL]) compared with those in healthy subjects (n = 11; median, 8 RMPs/μL [range, 3-166 RMPs/μL]; P = .01). RMP concentrations were highest in patients with severe falciparum malaria (P = .01). Parasitized red cells produced >10 times more RMPs than did unparasitized cells, but the overall majority of RMPs still derived from uninfected red blood cells (URBCs). In cultures, RMP production increased as the parasites matured. Hemin and parasite products induced RMP production in URBCs, which was inhibited by N-acetylcysteine, suggesting heme-mediated oxidative stress as a pathway for the generation of RMPs.

Antibodies to actin in autoimmune haemolytic anaemia

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Ritzmann Mathias

2010-03-01

Full Text Available Abstract Background In autoimmune haemolytic anaemia (AIHA, autoreactive antibodies directed against red blood cells are up-regulated, leading to erythrocyte death. Mycoplasma suis infections in pigs induce AIHA of both the warm and cold types. The aim of this study was to identify the target autoantigens of warm autoreactive IgG antibodies. Sera from experimentally M. suis-infected pigs were screened for autoreactivity. Results Actin-reactive antibodies were found in the sera of 95% of all animals tested. The reactivity was species-specific, i.e. reactivity with porcine actin was significantly higher than with rabbit actin. Sera of animals previously immunised with the M. suis adhesion protein MSG1 showed reactivity with actin prior to infection with M. suis indicating that molecular mimicry is involved in the specific autoreactive mechanism. A potentially cross-reactive epitope was detected. Conclusions This is the first report of autoreactive anti-actin antibodies involved in the pathogenesis of autoimmune haemolytic anaemia.

Mouse-specific antibody responses to a monoclonal antibody during repeated immunoscintigraphy investigations: Comparison of antibody titres and imaging studies in a rat model

International Nuclear Information System (INIS)

Pimm, M.V.; Gribben, S.J.; Markham, A.J.; Perkins, A.C.

1990-01-01

As a model for human mouse-specific antibody responses in patients undergoing immunoscintigraphy, we have investigated in rats the production of mouse-specific antibodies (MA) to the mouse monoclonal antibody 791T/36. At intervals of between 5 and 16 weeks the rats were given repeated cycles of intravenous (IV) injections of antibody with or without a simultaneous intradermal (ID) injection. The IV dose was 60 μg/kg, a dose similar to that used in many clinical immunoscintigraphy studies. The ID injection was 2 μg, which mimicks the skin test dose often given in clinical imaging protocols. The study was carried out with both 131 I-labelled antibody and with antibody labelled with 111 In by DTPA chelation. MA was measured with a passive haemagglutination assay using sheep red blood cells coated with the monoclonal antibody. Of rats given ID injections of unlabelled antibody at the same time as the IV imaging doses, 9/20 produced MA during 4 cycles of injections. In contrast, only 2/16 rats given only the IV dose produced MA. Both 131 I- and 111 In-labelled antibody appeared equally immunogenic with 5/18 and 6/18 overall responders, respectively. The production of MA was associated with a significant perturbation in the biodistribution of the IV dose of labelled antibody as seen by gamma-camera imaging of the rats given 111 In-labelled antibody. There was clearance of immune complexes to the liver, this organ accumulating up to 90% of the whole body count rate of radiolabel. MA titres of between 1/100 and 1/78000 caused equal perturbation of biodistribution, although below 1/100 the effect was more variable. (orig.)

A Monoclonal Antibody against Wnt-1 Induces Apoptosis in Human Cancer Cells

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Biao He

2004-01-01

Full Text Available Aberrant activation of the Wingless-type (Wnt/β-catenin signaling pathway is associated with a variety of human cancers. Little is known regarding the role that Wnt ligands play in human carcinogenesis. To test whether a Wnt-1 signal is a survival factor in human cancer cells and thus may serve as a potential cancer therapeutic target, we investigated the effect of inhibition of Wnt-1 signaling in a variety of human cancer cell lines, including non small cell lung cancer, breast cancer, mesothelioma, and sarcoma. Both monoclonal antibody and RNA interference (RNAi were used to inhibit Wnt-1 signaling. We found that incubation of a monoclonal anti-Wnt-1 antibody induced apoptosis and caused downstream protein changes in cancer cells overexpressing Wnt-1. In contrast, apoptosis was not detected in cells lacking or having minimal Wnt-1 expression after the antibody incubation. RNAi targeting of Wnt-1 in cancer cells overexpressing Wnt-1 demonstrated similar downstream protein changes and induction of apoptosis. The antibody also suppressed tumor growth in vivo. Our results indicate that both monoclonal anti-Wnt-1 antibody and Wnt-1 siRNA inhibit Wnt-1 signaling and can induce apoptosis in human cancer cells. These findings hold promise as a novel therapeutic strategy for cancer.

Role of red cells and plasma composition on blood sessile droplet evaporation

Science.gov (United States)

Lanotte, Luca; Laux, Didier; Charlot, Benoît; Abkarian, Manouk

2017-11-01

The morphology of dried blood droplets derives from the deposition of red cells, the main components of their solute phase. Up to now, evaporation-induced convective flows were supposed to be at the base of red cell distribution in blood samples. Here, we present a direct visualization by videomicroscopy of the internal dynamics in desiccating blood droplets, focusing on the role of cell concentration and plasma composition. We show that in diluted suspensions, the convection is promoted by the rich molecular composition of plasma, whereas it is replaced by an outward red blood cell displacement front at higher hematocrits. We also evaluate by ultrasounds the effect of red cell deposition on the temporal evolution of sample rigidity and adhesiveness.

Immunochemical characterization of rhesus proteins with antibodies raised against synthetic peptides.

Science.gov (United States)

Hermand, P; Mouro, I; Huet, M; Bloy, C; Suyama, K; Goldstein, J; Cartron, J P; Bailly, P

1993-07-15

Rabbit polyclonal antibodies were raised against synthetic peptides corresponding to hydrophilic regions of the human Rhesus (Rh) IX cDNA-encoded polypeptide predicted to be extracellularly or intracellularly exposed in the topologic model of the Rh blood group protein. Four antibodies encompassing residues 33-45 (MPC1), 224-233 (MPC4), 390-404 (MPC6), and 408-416 (MPC8) were characterized and compared with a polyclonal anti-Rh protein obtained by immunization with purified Rh proteins. All antibodies had specificity for authentic Rh polypeptides and reacted on Western blot with Rh proteins immunoprecipitated with human monoclonal anti-RhD, -c, and -E. MPC1, but not the other antibodies, agglutinated all human erythrocytes except Rhnull and Rhmod cells, which either lack totally or are severely deficient in Rh proteins, respectively. Immunoblotting analysis with membrane proteins from common and rare variants showed that MPC1 and MPC8 reacted in Western blot with 32-Kd Rh polypeptides from all common red blood cells except those from Rhnull and Rhmod, indicating that peptide regions 33-45 and 408-416 may be common to several if not all Rh proteins, whatever the Rh blood group specificity. MPC4 reacted only with membrane preparations from cells carrying the E antigen, whereas MPC6 recognized preferentially the Rh proteins from E and Ee preparations, suggesting that the protein encoded by the RhIXb cDNA carries the E and/or e antigen(s). Immunoadsorption experiments using inside-out or right-side-out sealed vesicules from DccEE red blood cells as competing antigen showed that the MPC6 and MPC8 antibodies bound only to the cytoplasmic side of the erythrocyte membrane, thus providing evidence for the intracellular orientation of the C-terminal 27 residues of the Rh polypeptides. Attempts to transiently or stably express the Rh polypeptides. Attempts to transiently or stably express the Rh cDNA in eukaryotic cells were largely unsuccessful, suggesting that Rh antigen

The Red Blood Cell Membrane of Preterm Infants in the Early Neonatal Period

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S. A. Perepelitsa

2014-01-01

Full Text Available Objective: to study the nanostructure of red blood cell membranes and erythrocyte index in preterm neonatal infants.Subjects and methods. The trial enrolled 47 neonatal infants, including 33 preterm infants who were included in a study group and 14 fullterm infants who formed a comparative group. The gestational age of the preterm infants was 33.3±1.9 weeks and the birth weight was 2065.4±304.8 g. Red blood cell counts, hemoglobin, and erythrocyte indices were estimat ed and the red blood cells were examined using an atomicforce microscope.Results. At birth, the preterm infants showed macrocytosis, intrauterine poikylocytosis, and the impaired nanostructure of red blood cell membranes. Intrauterine hypoxia affects the red blood cell membrane nanostructures: a phospholipid bilayer and a spectrin matrix, without damaging the membrane protein component. The detected changes are reversible and directed to maintaining the functional ability of red blood cells in a critical situation. At birth, gestational age, a baby's weight, hemoglobin, and blood cholesterol and standard bicarbonate levels influence the parameters of a red blood cell component. The early neonatal period was characterized by an active process on the red blood cell membranes and a change of morphological forms, suggesting the continuing postnatal rearrangement of erythropoiesis and a preterm infant's adaptation to new environmental conditions.

A synthetic glycan microarray enables epitope mapping of plant cell wall glycan-directed antibodies

DEFF Research Database (Denmark)

Ruprecht, Colin; Bartetzko, Max P; Senf, Deborah

2017-01-01

In the last three decades, more than 200 monoclonal antibodies have been raised against most classes of plant cell wall polysaccharides by different laboratories world-wide. These antibodies are widely used to identify differences in plant cell wall components in mutants, organ and tissue types......, and developmental stages. Despite their importance and broad use, the precise binding epitope for only a few of these antibodies has been determined. Here, we use a plant glycan microarray equipped with 88 synthetic oligosaccharides to comprehensively map the epitopes of plant cell wall glycan-directed antibodies....... Our results reveal the binding epitopes for 78 arabinogalactan-, rhamnogalacturonan-, xylan-, and xyloglucan-directed antibodies. We demonstrate that, with knowledge of the exact epitopes recognized by individual antibodies, specific glycosyl hydrolases can be implemented into immunological cell wall...

Red blood cell-deformability measurement: review of techniques.

Science.gov (United States)

Musielak, M

2009-01-01

Cell-deformability characterization involves general measurement of highly complex relationships between cell biology and physical forces to which the cell is subjected. The review takes account of the modern technical solutions simulating the action of the force applied to the red blood cell in macro- and microcirculation. Diffraction ektacytometers and rheoscopes measure the mean deformability value for the total red blood cell population investigated and the deformation distribution index of individual cells, respectively. Deformation assays of a whole single cell are possible by means of optical tweezers. The single cell-measuring setups for micropipette aspiration and atomic force microscopy allow conducting a selective investigation of deformation parameters (e.g., cytoplasm viscosity, viscoelastic membrane properties). The distinction between instrument sensitivity to various RBC-rheological features as well as the influence of temperature on measurement are discussed. The reports quoted confront fascinating possibilities of the techniques with their medical applications since the RBC-deformability has the key position in the etiology of a wide range of conditions.

Fully human monoclonal antibodies from antibody secreting cells after vaccination with Pneumovax®23 are serotype specific and facilitate opsonophagocytosis.

Science.gov (United States)

Smith, Kenneth; Muther, Jennifer J; Duke, Angie L; McKee, Emily; Zheng, Nai-Ying; Wilson, Patrick C; James, Judith A

2013-05-01

B lymphocyte memory generates antibody-secreting cells (ASCs) that represent a source of protective antibodies that may be exploited for therapeutics. Here we vaccinated four donors with Pneumovax®23 and produced human monoclonal antibodies (hmAbs) from ASCs. We have cloned 137 hmAbs and the specificities of these antibodies encompass 19 of the 23 serotypes in the vaccine, as well as cell wall polysaccharide (CWPS). Although the majority of the antibodies are serotype specific, 12% cross-react with two serotypes. The Pneumovax®23 ASC antibody sequences are highly mutated and clonal, indicating an anamnestic response, even though this was a primary vaccination. Hmabs from 64% of the clonal families facilitate opsonophagocytosis. Although 9% of the total antibodies bind to CWPS impurity in the vaccine, none of these clonal families showed opsonophagocytic activity. Overall, these studies have allowed us to address unanswered questions in the field of human immune responses to polysaccharide vaccines, including the cross-reactivity of individual antibodies between serotypes and the percentage of antibodies that are protective after vaccination with Pneumovax®23. Copyright © 2012 Elsevier GmbH. All rights reserved.

Partial Red Blood Cell Exchange in Children and Young Patients with Sickle Cell Disease: Manual Versus Automated Procedure.

Science.gov (United States)

Escobar, Carlos; Moniz, Marta; Nunes, Pedro; Abadesso, Clara; Ferreira, Teresa; Barra, António; Lichtner, Anabela; Loureiro, Helena; Dias, Alexandra; Almeida, Helena

2017-10-31

The benefits of manual versus automated red blood cell exchange have rarely been documented and studies in young sickle cell disease patients are scarce. We aim to describe and compare our experience in these two procedures. Young patients (≤ 21 years old) who underwent manual- or automated-red blood cell exchange for prevention or treatment of sickle cell disease complications were included. Clinical, technical and hematological data were prospectively recorded and analyzed. Ninety-four red blood cell exchange sessions were performed over a period of 68 months, including 57 manual and 37 automated, 63 for chronic complications prevention, 30 for acute complications and one in the pre-operative setting. Mean decrease in sickle hemoglobin levels was higher in automated-red blood cell exchange (p exchange and access alarm on automated-red blood cell exchange. No major complication or alloimunization was recorded. Automated-red blood cell exchange decreased sickle hemoglobin levels more efficiently than manual procedure in the setting of acute and chronic complications of sickle cell disease, with minor technical concerns mainly due to vascular access. The threshold of sickle hemoglobin should be individualized for clinical and hematological goals. In our cohort of young patients, the need for an acceptable venous access was a limiting factor, but iron-overload was avoided. Automated red blood cell exchange is safe and well tolerated. It permits a higher sickle hemoglobin removal efficacy, better volume status control and iron-overload avoidance.

Cell membrane antigen-antibody complex dissociation by the widely used glycine-HCL method: an unreliable procedure for studying antibody internalization.

Science.gov (United States)

Tsaltas, G; Ford, C H

1993-02-01

Methods following the process of binding and internalization of antibodies to cell surface antigens have often employed low pH isoosmolar buffers in order to dissociate surface antigen-antibody complexes. One of the most widely used buffers is a 0.05 M glycine-HCL buffer pH 2.8. Since the efficacy of action of this buffer was critical to a series of internalization experiments employing monoclonal antibodies (Mabs) to carcinoembryonic antigen (CEA) expressing cancer cell lines in this laboratory, we tested its performance in a number of different assays. Our results indicate that this buffer only partially dissociates antigen-antibody bonds and therefore can introduce major inaccuracies in internalization experiments.

Pre-existing neutralizing antibody mitigates B cell dysregulation and enhances the Env-specific antibody response in SHIV-infected rhesus macaques.

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Juan Pablo Jaworski

Full Text Available Our central hypothesis is that protection against HIV infection will be powerfully influenced by the magnitude and quality of the B cell response. Although sterilizing immunity, mediated by pre-formed abundant and potent antibodies is the ultimate goal for B cell-targeted HIV vaccine strategies, scenarios that fall short of this may still confer beneficial defenses against viremia and disease progression. We evaluated the impact of sub-sterilizing pre-existing neutralizing antibody on the B cell response to SHIV infection. Adult male rhesus macaques received passive transfer of a sub-sterilizing amount of polyclonal neutralizing immunoglobulin (Ig purified from previously infected animals (SHIVIG or control Ig prior to intra-rectal challenge with SHIVSF162P4 and extensive longitudinal sampling was performed. SHIVIG treated animals exhibited significantly reduced viral load and increased de novo Env-specific plasma antibody. Dysregulation of the B cell profile was grossly apparent soon after infection in untreated animals; exemplified by a ≈50% decrease in total B cells in the blood evident 2-3 weeks post-infection which was not apparent in SHIVIG treated animals. IgD+CD5+CD21+ B cells phenotypically similar to marginal zone-like B cells were highly sensitive to SHIV infection, becoming significantly decreased as early as 3 days post-infection in control animals, while being maintained in SHIVIG treated animals, and were highly correlated with the induction of Env-specific plasma antibody. These results suggest that B cell dysregulation during the early stages of infection likely contributes to suboptimal Env-specific B cell and antibody responses, and strategies that limit this dysregulation may enhance the host's ability to eliminate HIV.

Apoptotic Effect of Anti myeloma Polyclonal Antibodies on The Growth of Myeloma Cells

International Nuclear Information System (INIS)

Abd El-Ghany, I.Y.; El-Kolaly, M.T.; Moustafa, K.A.; El-Shershaby, H.M.; Sayed, A.A.; Borai, I.H.; El-Lahloby, N.M.

2013-01-01

Multiple myeloma (MM) is a malignancy characterized by proliferation of plasma cells. Cancer immunotherapy is a major branch of biological therapy that utilizes living cells and their products. The aim of this study is to produce and evaluate the antiproliferative effect of anti myeloma polyclonal antibodies (with and without labelling with radioactive isotopes) against the growth of myeloma cells. The production of polyclonal antibodies (PAb) was generated by immunizing five healthy female mature white New-Zealand rabbits with myeloma cells (SP2/OR) through primary injection and five booster doses. The preparation of labelled anti myeloma antibodies was carried out using chloramine-T method and it was purified using PD-10 chromatographic column. The results obtained revealed that anti myeloma polyclonal antibodies inhibited proliferation and induced apoptosis of myeloma cell lines in vitro and induced apoptosis after serial intraperitoneal injection of PAb in ascites bearing mice in vivo. The present study suggested that the effect of labelled anti myeloma antibodies on myeloma cells growth inhibition was more effective than that of anti myeloma antibodies without labelling which is due to the cytotoxic effect of ionizing radiation. Apoptosis triggered by PAb was confirmed by flow cytometry, caspase -8 and -9 and β2-microglobulin.

Fed-batch CHO cell culture for lab-scale antibody production

DEFF Research Database (Denmark)

Fan, Yuzhou; Ley, Daniel; Andersen, Mikael Rørdam

2017-01-01

Fed-batch culture is the most commonly used upstream process in industry today for recombinant monoclonal antibody production using Chinese hamster ovary cells. Developing and optimizing this process in the lab is crucial for establishing process knowledge, which enable rapid and predictable tech......-transfer to manufacturing scale. In this chapter, we will describe stepwise how to carry out fed-batch CHO cell culture for lab-scale antibody production....

Transfer plate radioassay using cell monolayers to detect anti-cell surface antibodies synthesized by lymphocyte hybridomas

International Nuclear Information System (INIS)

Schneider, M.D.; Eisenbarth, G.S.

1979-01-01

A solid phase [ 125 I] Protein A radioassay for anti-cell surface antibodies is described, which employs target cell monolayers cultured on fenestrated polyvinyl chloride 96-well plates ('transfer plates'). The calibrated aperture in the bottom of each well is small enough to retain fluid contents by surface tension during monolayer growth, but also permits fluid to enter the wells when transfer plate are lowered into receptacles containing washing buffer on test sera. To assay for antibodies directed against target cell surface antigens, transfer plates bearing monolayers are inserted into microculture plates with corresponding 96-well geometry, thereby simultaneously sampling 96 wells. This assay allows rapid screening of hundreds of hybrid cell colonies for production of antibodies with desired tissue specificity. (Auth.)

Synergistic cytotoxic effects of antibodies directed against different cell surface determinants

Energy Technology Data Exchange (ETDEWEB)

Elliott, E V; Pindar, A; Stevenson, F K; Stevenson, G T [Southampton General Hospital (UK). Tenovus Research Lab.

1978-03-01

Three antibody populations were raised in rabbits against surface antigens on guinea-pig L/sub 2/C leukaemic lymphocytes: against idiotypic determinants on the lambda chain of the surface immunoglobulin, against C region determinants on the lambda chain, and against the surface antigens recognised by conventional anti-lymphocyte sera. Complement and K-cell cytotoxicities effected by the antibodies on L/sub 2/C cells were studied in vitro. In both cytotoxic systems mixtures of the antibodies revealed synergy, in that the titres of the mixtures exceeded predicted additive titres of their components. The synergy was greater when the mixed antibodies were directed to determinants on the same molecule rather than to determinants on different molecules.

Preparation of anti-Sudan red monoclonal antibody and development of an indirect competitive enzyme-linked immunosorbent assay for detection of Sudan red in chilli jam and chilli oil.

Science.gov (United States)

Xu, Jing; Zhang, Yuanyang; Yi, Jian; Meng, Meng; Wan, Yuping; Feng, Caiwei; Wang, Shanliang; Lu, Xiao; Xi, Rimo

2010-10-01

Sudan dyes are banned to be used in food additives because of the carcinogenicity of their metabolites. A rapid and sensitive indirect competitive enzyme-linked immunosorbent assay (ELISA) was developed to detect the residues of Sudan dyes. Novel immunogen and coating antigen were synthesized via glutaraldehyde linking. The hapten-bovine serum albumin (BSA) was applied as immunogen and the hapten-ovalbumin (OVA) was served as coating antigen. The monoclonal antibody obtained showed high sensitivity to Sudan I with an IC(50) value of 1.7 μg L(-1) in buffer and was suitable to detect the residues of Sudan red in food products. The specificity of the assay was studied by measuring cross-reactivity of the antibody with the structurally related compounds of Sudan II (red (120%). Chilli jam and chilli oil samples spiked with Sudan dyes were analyzed by the method. The detection limit (LOD) of the ELISA method applied in chilli jam and chilli oil was 9.0 μg L(-1) and 19.6 μg L(-1), respectively. The recovery rates of Sudan-I in chilli oil and chilli jam were in the range of 80%-110% with coefficients of variation <25%. The intra-assay variation and inter-assay variation in buffer were both <9%.

Mapping the stem cell state: eight novel human embryonic stem and embryonal carcinoma cell antibodies

DEFF Research Database (Denmark)

Wright, A; Andrews, N; Bardsley, K

2011-01-01

The antigenic profile of human embryonic stem (ES) and embryonal carcinoma (EC) cells has served as a key element of their characterization, with a common panel of surface and intracellular markers now widely used. Such markers have been used to identify cells within the 'undifferentiated state...... of reactivity for all antibodies against both ES and EC cells, suggesting that these markers will afford recognition of unique sub-states within the undifferentiated stem cell compartment....... and EC cells, and herein describe their characterization. The reactivity of these antibodies against a range of cell lines is reported, as well as their developmental regulation, basic biochemistry and reactivity in immunohistochemistry of testicular germ cell tumours. Our data reveal a range...

Autologous Hematopoietic Stem Cell Transplantation to Prevent Antibody Mediated Rejection After Vascularized Composite Allotransplantation

Science.gov (United States)

2017-10-01

Award Number: W81XWH-16-1-0664 TITLE: Autologous Hematopoietic Stem Cell Transplantation to Prevent Antibody-Mediated Rejection after...Annual 3. DATES COVERED 15 Sep 2016 – 14 Sep 2017 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Autologous Hematopoietic Stem Cell Transplantation to...sensitization, autologous hematopoietic stem cell transplantation, antibody mediated rejection, donor specific antibodies 16. SECURITY CLASSIFICATION OF

Antibody-independent control of gamma-herpesvirus latency via B cell induction of anti-viral T cell responses.

Directory of Open Access Journals (Sweden)

Kelly B McClellan

2006-06-01

Full Text Available B cells can use antibody-dependent mechanisms to control latent viral infections. It is unknown whether this represents the sole function of B cells during chronic viral infection. We report here that hen egg lysozyme (HEL-specific B cells can contribute to the control of murine gamma-herpesvirus 68 (gammaHV68 latency without producing anti-viral antibody. HEL-specific B cells normalized defects in T cell numbers and proliferation observed in B cell-/- mice during the early phase of gammaHV68 latency. HEL-specific B cells also reversed defects in CD8 and CD4 T cell cytokine production observed in B cell-/- mice, generating CD8 and CD4 T cells necessary for control of latency. Furthermore, HEL-specific B cells were able to present virally encoded antigen to CD8 T cells. Therefore, B cells have antibody independent functions, including antigen presentation, that are important for control of gamma-herpesvirus latency. Exploitation of this property of B cells may allow enhanced vaccine responses to chronic virus infection.

Identification and prevention of antibody disulfide bond reduction during cell culture manufacturing.

Science.gov (United States)

Trexler-Schmidt, Melody; Sargis, Sandy; Chiu, Jason; Sze-Khoo, Stefanie; Mun, Melissa; Kao, Yung-Hsiang; Laird, Michael W

2010-06-15

In the biopharmaceutical industry, therapeutic monoclonal antibodies are primarily produced in mammalian cell culture systems. During the scale-up of a monoclonal antibody production process, we observed excessive mechanical cell shear as well as significant reduction of the antibody's interchain disulfide bonds during harvest operations. This antibody reduction event was catastrophic as the product failed to meet the drug substance specifications and the bulk product was lost. Subsequent laboratory studies have demonstrated that cells subjected to mechanical shear release cellular enzymes that contribute to this antibody reduction phenomenon (manuscript submitted; Kao et al., 2009). Several methods to prevent this antibody reduction event were developed using a lab-scale model to reproduce the lysis and reduction events. These methods included modifications to the cell culture media with chemicals (e.g., cupric sulfate (CuSO(4))), pre- and post-harvest chemical additions to the cell culture fluid (CCF) (e.g., CuSO(4), EDTA, L-cystine), as well as lowering the pH and air sparging of the harvested CCF (HCCF). These methods were evaluated for their effectiveness in preventing disulfide bond reduction and their impact to product quality. Effective prevention methods, which yielded acceptable product quality were evaluated for their potential to be implemented at manufacturing-scale. The work described here identifies numerous effective reduction prevention measures from lab-scale studies; several of these methods were then successfully translated into manufacturing processes. 2010 Wiley Periodicals, Inc.

Acute and chronic influence of temperature on red blood cell anion exchange.

Science.gov (United States)

Jensen, F B; Wang, T; Brahm, J

2001-01-01

Unidirectional (36)Cl(-) efflux via the red blood cell anion exchanger was measured under Cl(-) self-exchange conditions (i.e. no net flow of anions) in rainbow trout Oncorhynchus mykiss and red-eared freshwater turtle Trachemys scripta to examine the effects of acute temperature changes and acclimation temperature on this process. We also evaluated the possible adaptation of anion exchange to different temperature regimes by including our previously published data on other animals. An acute temperature increase caused a significant increase in the rate constant (k) for unidirectional Cl(-) efflux in rainbow trout and freshwater turtle. After 3 weeks of temperature acclimation, 5 degrees C-acclimated rainbow trout showed only marginally higher Cl(-) transport rates than 15 degrees C-acclimated trout when compared at the same temperature. Apparent activation energies for red blood cell Cl(-) exchange in trout and turtle were lower than values reported in endothermic animals. The Q(10) for red blood cell anion exchange was 2.0 in trout and 2.3 in turtle, values close to those for CO(2) excretion, suggesting that, in ectothermic animals, the temperature sensitivity of band-3-mediated anion exchange matches the temperature sensitivity of CO(2) transport (where red blood cell Cl(-)/HCO(3)(-) exchange is a rate-limiting step). In endotherms, such as man and chicken, Q(10) values for red blood cell anion exchange are considerably higher but are no obstacle to CO(2) transport, because body temperature is normally kept constant at values at which anion exchange rates are high. When compared at constant temperature, red blood cell Cl(-) permeability shows large differences among species (trout, carp, eel, cod, turtle, alligator, chicken and man). Cl(-) permeabilities are, however, remarkable similar when compared at preferred body temperatures, suggesting an appropriate evolutionary adaptation of red blood cell anion exchange function to the different thermal niches occupied

Immunity War: A Novel Therapy for Lymphoma Using T-cell Bispecific Antibodies.

Science.gov (United States)

Prakash, Ajay; Diefenbach, Catherine S

2018-06-08

The activity of T cell mediated immunotherapies in B cell lymphoma has been limited to date. The novel bi-specific antibody CD20-TCB, has a 2:1 antibody design to maximize T cell engagement, and demonstrates activity in preclinical models. This may represent a novel therapeutic approach for patients with relapsed/refractory NHL. Copyright ©2018, American Association for Cancer Research.

DETERMINANTS OF RED-BLOOD-CELL DEFORMABILITY IN RELATION TO CELL AGE

NARCIS (Netherlands)

BOSCH, FH; WERRE, JM; ROERDINKHOLDERSTOELWINDER, B; HULS, T; WILLEKENS, FLA; WICHERS, G; HALIE, MR

Red blood cell (RBC) deformability was determined with an ektacytometer in fractions separated on the basis of differences in cell volume or density. Deformability was measured with ektacytometry (rpm-scan and osmo-scan). We studied three groups of RBC fractions:l. By counterflow centrifugation we

Anti-idiotypic antibodies that protect cells against the action of diphtheria toxin

International Nuclear Information System (INIS)

Rolf, J.M.; Gaudin, H.M.; Tirrell, S.M.; MacDonald, A.B.; Eidels, L.

1989-01-01

An anti-idiotypic serum prepared against the combining site (idiotype) of specific anti-diphtheria toxoid antibodies was characterized with respect to its interaction with highly diphtheria toxin-sensitive Vero cells. Although the anti-idiotypic serum protected Vero cells against the cytotoxic action of diphtheria toxin, it did not prevent the binding of 125 I-labeled diphtheria toxin to the cells but did inhibit the internalization and degradation of 125 I-labeled toxin. This anti-idiotypic serum immunoprecipitated a cell-surface protein from radiolabeled Vero cells with an apparent Mr of approximately 15,000. These results are consistent with the hypothesis that the anti-idiotypic serum contains antibodies that carry an internal image of an internalization site on the toxin and that a cell-surface protein involved in toxin internalization possesses a complementary site recognized by both the toxin and the anti-idiotypic antibodies

Antibody screening & identification in the general patient population at a tertiary care hospital in New Delhi, India.

Science.gov (United States)

Makroo, Raj Nath; Bhatia, Aakanksha; Hegde, Vikas; Chowdhry, Mohit; Thakur, Uday Kumar; Rosamma, N L

2014-09-01

The development of alloantibodies can significantly complicate transfusion therapy and results in difficulties in cross-matching of blood. Most literature on alloimmunization is limited to multitransfused individuals, with very few studies on the general hospital patients. This study was aimed at assessing the frequency and type of unexpected red cell antibodies in the general patient population at a multispecialty tertiary care centre in New Delhi, India. The results of 49,077 antibody screening tests carried out on patients, from January 2009 to December 2012 were analyzed. The clinical and transfusion records were reviewed. The data were compiled and statistically analysed. A total of 49,077 (29,917; 60.96% males and 19,160; 39.04% females) patient samples were screened for the presence of unexpected antibodies. Antibody screening was positive in 403 patients (0.82%). In the serum samples of 164 patients only autoantibodies were identified, 27 revealed autoantibodies with one or more underlying alloantibodies, while 212 patients had only alloantibody/ies in their serum. The overall alloimmunization rate was 0.49 per cent. Antibodies against the Rh system were the most frequent (64.1%), the most common alloantibody identified being anti E (37.2%), followed by anti D (19.2%). Since clinically significant antibodies are frequently detected in our patient population, antibody screening and if required, identification is the need of the hour. Since antibodies against the common Rh and Kell blood group antigens are the most frequent, provision of Rh and Kell matched red cells may be of protective value.

Techniques for measuring red cell, platelet, and WBC survival

International Nuclear Information System (INIS)

Mayer, K.; Freeman, J.E.

1986-01-01

Blood cell survival studies yield valuable information concerning production and destruction of cells circulating in the bloodstream. Methodologies for the measurement of red cell survival include nonisotopic methods such as differential agglutination and hemolysis. The isotopic label may be radioactive or, if not, will require availability of a mass spectrograph. These methods fall into two categories, one where red cells of all ages are labeled ( 51 Cr, DFP32, etc.) and those employing a cohort label of newly formed cells ( 14 C glycine, 75 Se methionine, etc.). Interpretation of results for methodology employed and mechanism of destruction, random or by senescence, are discussed. A similar approach is presented for platelet and leukocyte survival studies. The inherent difficulties and complications of sequestration, storage, and margination of these cells are emphasized and discussed. 38 references

Estimation of transfused red cell survival using an enzyme-linked antiglobulin test

International Nuclear Information System (INIS)

Kickler, T.S.; Smith, B.; Bell, W.; Drew, H.; Baldwin, M.; Ness, P.M.

1985-01-01

An enzyme-linked antiglobulin test (ELAT) method was developed to estimate survival of transfused red cells. This procedure is based on a principle analogous to that of the Ashby technique were antigenically distinct red cells are transfused and their survival studied. The authors compared the ELAT survival to the 51 Chromium method ( 51 Cr) in four patients. Three patients with hypoproliferative anemias showed T 1/2 by ELAT of 17.5, 18, and 17 days versus 18.5, 20, and 19 days by the 51 Cr method. A fourth patient with traumatic cardiac hemolysis had two studies performed. In this case, the ELAT showed a T 1/2 of 10 and 8.1 days while 51 Cr T 1/2 values were 11 and 10.5 days. The ELAT method for measuring red cell survival yielded data which agreed closely with the results of the 51 Cr method. Although 51 Cr is the accepted method for red cell survival, the ELAT method can be used to estimate transfused red cell survival

An improved autoradiographic technique for the detection of antibody-forming cells

International Nuclear Information System (INIS)

Mason, D.W.

1976-01-01

An autoradiographic technique for the detection of antibody-forming cells has been developed for the assay of anti-DNP responses. The lymphoid cell suspension to be assayed was allowed to sediment on to a glass slide coated with DNP-conjugated gelatin to which the secreted antibody bound during subsequent incubation. The bound antibody and its Ig class was revealed by a second incubation using 125 I-anti-immunoglobulin reagents followed by autoradiography. Studies on the sensitivity and specificity of the method are presented and its advantages over other techniques described. The technique should be readily applicable to other haptens

CD133 antibody conjugation to decellularized human heart valves intended for circulating cell capture.

Science.gov (United States)

Vossler, John D; Min Ju, Young; Williams, J Koudy; Goldstein, Steven; Hamlin, James; Lee, Sang Jin; Yoo, James J; Atala, Anthony

2015-09-03

The long term efficacy of tissue based heart valve grafts may be limited by progressive degeneration characterized by immune mediated inflammation and calcification. To avoid this degeneration, decellularized heart valves with functionalized surfaces capable of rapid in vivo endothelialization have been developed. The aim of this study is to examine the capacity of CD133 antibody-conjugated valve tissue to capture circulating endothelial progenitor cells (EPCs). Decellularized human pulmonary valve tissue was conjugated with CD133 antibody at varying concentrations and exposed to CD133 expressing NTERA-2 cl.D1 (NT2) cells in a microflow chamber. The amount of CD133 antibody conjugated on the valve tissue surface and the number of NT2 cells captured in the presence of shear stress was measured. Both the amount of CD133 antibody conjugated to the valve leaflet surface and the number of adherent NT2 cells increased as the concentration of CD133 antibody present in the surface immobilization procedure increased. The data presented in this study support the hypothesis that the rate of CD133(+) cell adhesion in the presence of shear stress to decellularized heart valve tissue functionalized by CD133 antibody conjugation increases as the quantity of CD133 antibody conjugated to the tissue surface increases.

Cardiac arrest due to hyperkalemia following irradiated packed red cells transfusion

Energy Technology Data Exchange (ETDEWEB)

Miyazawa, Kazuharu [Yamamoto-kumiai General Hospital, Noshiro, Akita (Japan); Ohta, Sukejuurou; Kojima, Yukiko; Mizunuma, Takahide; Nishikawa, Toshiaki

1998-11-01

We describe two cases of cardiac arrest due to hyperkalemia following transfusion of irradiated packed red cells. Case 1: Because sudden, rapid and massive hemorrage occurred in a 69-year-old male patient undergoing the left lobectomy of the liver, 8 units of irradiated packed red cells were rapidly transfused, the patient developed cardiac arrest. Serum kalium concentration after transfusion was 7.6 mEq/l. Case 2: A 7-month-old girl scheduled for closure of a ventricular septal defect, developed cardiac arrest due to hyperkalemia at the start of cardiopulmonary bypass. The extracorporeal circuit was primed with 6 units of irradiated packed red blood cells. Serum kalium concentration immediately after the start of cardiopulmonary bypass was 10.6 mEq/l. Analysis of kalium concentration in the pilot tubes of the same packs revealed 56-61 mEq/l. These case reports suggest that fresh irradiated packed red cells should be transfused during massive bleeding and for pediatric patients to prevent severe hyperkalemia. (author)

Fixation effect of SurePath preservative fluids using epidermal growth factor receptor mutation-specific antibodies for immunocytochemistry.

Science.gov (United States)

Kawahara, Akihiko; Taira, Tomoki; Abe, Hideyuki; Watari, Kosuke; Murakami, Yuichi; Fukumitsu, Chihiro; Takase, Yorihiko; Yamaguchi, Tomohiko; Azuma, Koichi; Akiba, Jun; Ono, Mayumi; Kage, Masayoshi

2014-02-01

Cytological diagnosis of respiratory disease has become important, not only for histological typing using immunocytochemistry (ICC) but also for molecular DNA analysis of cytological material. The aim of this study was to investigate the fixation effect of SurePath preservative fluids. Human lung cancer PC9 and 11-18 cell lines, and lung adenocarcinoma cells in pleural effusion, were fixed in CytoRich Blue, CytoRich Red, 15% neutral-buffered formalin, and 95% ethanol, respectively. PC9 and 11-18 cell lines were examined by ICC with epidermal growth factor receptor (EGFR) mutation-specific antibodies, the EGFR mutation DNA assay, and fluorescence in situ hybridization. The effect of antigenic storage time was investigated in lung adenocarcinoma cells in pleural effusion by ICC using the lung cancer detection markers. PC9 and 11-18 cell lines in formalin-based fixatives showed strong staining of EGFR mutation-specific antibodies and lung cancer detection markers by ICC as compared with ethanol-based fixatives. DNA preservation with CytoRich Blue and CytoRich Red was superior to that achieved with 95% ethanol and 15% neutral-buffered formalin fixatives, whereas EGFR mutations by DNA assay and EGFR gene amplification by fluorescence in situ hybridization were successfully identified in all fixative samples. Although cytoplasmic antigens maintained high expression levels, expression levels in nuclear antigens fell as storage time increased. These results indicate that CytoRich Red is not only suitable for ICC with EGFR mutation-specific antibodies, but also for DNA analysis of cytological material, and is useful in molecular testing of lung cancer, for which various types of analyses will be needed in future. © 2013 American Cancer Society.

Wildtype p53-specific Antibody and T-Cell Responses in Cancer Patients

DEFF Research Database (Denmark)

Pedersen, Anders Elm; Stryhn, Anette; Justesen, Sune

2011-01-01

patients. Detection of antibodies against wt p53 protein has been used as a diagnostic and prognostic marker and discovery of new T-cell epitopes has enabled design of cancer vaccination protocols with promising results. Here, we identified wt p53-specific antibodies in various cancer patients......(264-272) in breast cancer patients and against HLA-A*01:01 binding peptide wt p53(226-234) and HLA-B*07:02 binding peptide wt p53(74-82) in renal cell cancer and breast cancer patients, respectively. Finally, we analyzed antibody and T-cell responses against wt p53 15-mer peptides in patients with metastatic renal...

The development of methods for obtaining monoclonal antibody-producing cells

Directory of Open Access Journals (Sweden)

Michał Skowicki

2016-04-01

Full Text Available Monoclonal antibodies (mAbs are biomolecules of great scientific and practical significance. In contrast to polyclonal antibodies from immune sera, they are homogeneous and monospecific, since they are produced by hybridoma cells representing a clone arising from a single cell. The successful technology was described for the first time in 1975; the inventors were later awarded the Nobel Prize. Currently, mAbs are broadly used as a research tool, in diagnostics and medicine in particular for the treatment of cancer or in transplantology. About 47 therapeutics based on monoclonal antibodies are now available in the US and Europe, and the number is still growing. Production of monoclonal antibodies is a multistage, time-consuming and costly process. Growing demand for these molecules creates space for research focused on improvements in hybridoma technology. Lower costs, human labor, and time are important goals of these attempts. In this article, a brief review of current methods and their advances is given.

Safe extension of red blood cell storage life at 4{degree}C

Energy Technology Data Exchange (ETDEWEB)

Bitensky, M.; Yoshida, Tatsuro

1996-04-01

The project sought to develop methods to extend the storage life of red blood cells. Extended storage would allow donor to self or autologous transfusion, expand and stabilize the blood supply, reduce the cost of medical care and eliminate the risk of transfusion related infections, including a spectrum of hepatitides (A, B and C) and HIV. The putative cause of red blood cell spoilage at 4 C has been identified as oxidative membrane damage resulting from deoxyhemoglobin and its denaturation products including hemichrome, hemin and Fe{sup 3+}. Trials with carbon monoxide, which is a stabilizer of hemoglobin, have produced striking improvement of red blood cell diagnostics for cells stored at 4 C. Carbonmonoxy hemoglobin is readily converted to oxyhemoglobin by light in the presence of oxygen. These findings have generated a working model and an approach to identify the best protocols for optimal red cell storage and hemoglobin regeneration.

A comparison of methods of determining the 100 percent survival of preserved red cells

International Nuclear Information System (INIS)

Valeri, C.R.; Pivacek, L.E.; Ouellet, R.; Gray, A.

1984-01-01

Studies were done to compare three methods to determine the 100 percent survival value from which to estimate the 24-hour posttransfusion survival of preserved red cells. The following methods using small aliquots of 51 Cr-labeled autologous preserved red cells were evaluated: First, the 125 I-albumin method, which is an indirect measurement of the recipient's red cell volume derived from the plasma volume measured using 125 I-labeled albumin and the total body hematocrit. Second, the body surface area method (BSA) in which the recipient's red cell volume is derived from a body surface area nomogram. Third, an extrapolation method, which extrapolates to zero time the radioactivity associated with the red cells in the recipient's circulation from 10 to 20 or 15 to 30 minutes after transfusion. The three methods gave similar results in all studies in which less than 20 percent of the transfused red cells were nonviable (24-hour posttransfusion survival values of between 80-100%), but not when more than 20 percent of the red cells were nonviable. When 21 to 35 percent of the transfused red cells were nonviable (24-hour posttransfusion survivals of 65 to 79%), values with the 125 I-albumin method and the body surface area method were about 5 percent lower (p less than 0.001) than values with the extrapolation method. When greater than 35 percent of the red cells were nonviable (24-hour posttransfusion survival values of less than 65%), values with the 125 I-albumin method and the body surface area method were about 10 percent lower (p less than 0.001) than those obtained by the extrapolation method

Anti-idiotypic antibodies that protect cells against the action of diphtheria toxin

Energy Technology Data Exchange (ETDEWEB)

Rolf, J.M.; Gaudin, H.M.; Tirrell, S.M.; MacDonald, A.B.; Eidels, L.

1989-03-01

An anti-idiotypic serum prepared against the combining site (idiotype) of specific anti-diphtheria toxoid antibodies was characterized with respect to its interaction with highly diphtheria toxin-sensitive Vero cells. Although the anti-idiotypic serum protected Vero cells against the cytotoxic action of diphtheria toxin, it did not prevent the binding of /sup 125/I-labeled diphtheria toxin to the cells but did inhibit the internalization and degradation of /sup 125/I-labeled toxin. This anti-idiotypic serum immunoprecipitated a cell-surface protein from radiolabeled Vero cells with an apparent Mr of approximately 15,000. These results are consistent with the hypothesis that the anti-idiotypic serum contains antibodies that carry an internal image of an internalization site on the toxin and that a cell-surface protein involved in toxin internalization possesses a complementary site recognized by both the toxin and the anti-idiotypic antibodies.

Enhanced CDC of B cell chronic lymphocytic leukemia cells mediated by rituximab combined with a novel anti-complement factor H antibody.

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Mark T Winkler

Full Text Available Rituximab therapy for B cell chronic lymphocytic leukemia (B-CLL has met with mixed success. Among several factors to which resistance can be attributed is failure to activate complement dependent cytotoxicity (CDC due to protective complement regulatory proteins, including the soluble regulator complement factor H (CFH. We hypothesized that rituximab killing of non-responsive B-CLL cells could be augmented by a novel human monoclonal antibody against CFH. The B cells from 11 patients with B-CLL were tested ex vivo in CDC assays with combinations of CFH monoclonal antibody, rituximab, and a negative control antibody. CDC of rituximab non-responsive malignant B cells from CLL patients could in some cases be augmented by the CFH monoclonal antibody. Antibody-mediated cytotoxicity of cells was dependent upon functional complement. In one case where B-CLL cells were refractory to CDC by the combination of rituximab plus CFH monoclonal antibody, additionally neutralizing the membrane complement regulatory protein CD59 allowed CDC to occur. Inhibiting CDC regulatory proteins such as CFH holds promise for overcoming resistance to rituximab therapy in B-CLL.

A novel monoclonal anti-CD81 antibody produced by genetic immunization efficiently inhibits Hepatitis C virus cell-cell transmission.

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Isabel Fofana

Full Text Available Hepatitis C virus (HCV infection is a challenge to prevent and treat because of the rapid development of drug resistance and escape. Viral entry is required for initiation, spread, and maintenance of infection, making it an attractive target for antiviral strategies.Using genetic immunization, we produced four monoclonal antibodies (mAbs against the HCV host entry factor CD81. The effects of antibodies on inhibition of HCV infection and dissemination were analyzed in HCV permissive human liver cell lines.The anti-CD81 mAbs efficiently inhibited infection by HCV of different genotypes as well as a HCV escape variant selected during liver transplantation and re-infecting the liver graft. Kinetic studies indicated that anti-CD81 mAbs target a post-binding step during HCV entry. In addition to inhibiting cell-free HCV infection, one antibody was also able to block neutralizing antibody-resistant HCV cell-cell transmission and viral dissemination without displaying any detectable toxicity.A novel anti-CD81 mAb generated by genetic immunization efficiently blocks HCV spread and dissemination. This antibody will be useful to further unravel the role of virus-host interactions during HCV entry and cell-cell transmission. Furthermore, this antibody may be of interest for the development of antivirals for prevention and treatment of HCV infection.

Cre-mediated cell ablation contests mast cell contribution in models of antibody- and T cell-mediated autoimmunity.

Science.gov (United States)

Feyerabend, Thorsten B; Weiser, Anne; Tietz, Annette; Stassen, Michael; Harris, Nicola; Kopf, Manfred; Radermacher, Peter; Möller, Peter; Benoist, Christophe; Mathis, Diane; Fehling, Hans Jörg; Rodewald, Hans-Reimer

2011-11-23

Immunological functions of mast cells remain poorly understood. Studies in Kit mutant mice suggest key roles for mast cells in certain antibody- and T cell-mediated autoimmune diseases. However, Kit mutations affect multiple cell types of both immune and nonimmune origin. Here, we show that targeted insertion of Cre-recombinase into the mast cell carboxypeptidase A3 locus deleted mast cells in connective and mucosal tissues by a genotoxic Trp53-dependent mechanism. Cre-mediated mast cell eradication (Cre-Master) mice had, with the exception of a lack of mast cells and reduced basophils, a normal immune system. Cre-Master mice were refractory to IgE-mediated anaphylaxis, and this defect was rescued by mast cell reconstitution. This mast cell-deficient strain was fully susceptible to antibody-induced autoimmune arthritis and to experimental autoimmune encephalomyelitis. Differences comparing Kit mutant mast cell deficiency models to selectively mast cell-deficient mice call for a systematic re-evaluation of immunological functions of mast cells beyond allergy. Copyright © 2011 Elsevier Inc. All rights reserved.

The Effect of Shape Memory on Red Blood Cell Motions

Science.gov (United States)

Niu, Xiting; Shi, Lingling; Pan, Tsorng-Whay; Glowinski, Roland

2013-11-01

An elastic spring model is applied to study the effect of the shape memory on the motion of red blood cell in flows. In shear flow, shape memory also plays an important role to obtain all three motions: tumbling, swinging, and tank-treading. In Poiseuille flow, cell has an equilibrium shape as a slipper or parachute depending on capillary number. To ensure the tank-treading motion while in slippery shape, a modified model is proposed by introducing a shape memory coefficient which describes the degree of shape memory in cells. The effect of the coefficient on the cell motion of red blood cell will be presented.

Prolonged storage of packed red blood cells for blood transfusion.

Science.gov (United States)

Martí-Carvajal, Arturo J; Simancas-Racines, Daniel; Peña-González, Barbra S

2015-07-14

A blood transfusion is an acute intervention, used to address life- and health-threatening conditions on a short-term basis. Packed red blood cells are most often used for blood transfusion. Sometimes blood is transfused after prolonged storage but there is continuing debate as to whether transfusion of 'older' blood is as beneficial as transfusion of 'fresher' blood. To assess the clinical benefits and harms of prolonged storage of packed red blood cells, in comparison with fresh, on recipients of blood transfusion. We ran the search on 1st May 2014. We searched the Cochrane Injuries Group Specialized Register, Cochrane Central Register of Controlled Trials (CENTRAL, The Cochrane Library), MEDLINE (OvidSP), Embase (OvidSP), CINAHL (EBSCO Host) and two other databases. We also searched clinical trials registers and screened reference lists of the retrieved publications and reviews. We updated this search in June 2015 but these results have not yet been incorporated. Randomised clinical trials including participants assessed as requiring red blood cell transfusion were eligible for inclusion. Prolonged storage was defined as red blood cells stored for ≥ 21 days in a blood bank. We did not apply limits regarding the duration of follow-up, or country where the study took place. We excluded trials where patients received a combination of short- and long-stored blood products, and also trials without a clear definition of prolonged storage. We independently performed study selection, risk of bias assessment and data extraction by at least two review authors. The major outcomes were death from any cause, transfusion-related acute lung injury, and adverse events. We estimated relative risk for dichotomous outcomes. We measured statistical heterogeneity using I(2). We used a random-effects model to synthesise the findings. We identified three randomised clinical trials, involving a total of 120 participants, comparing packed red blood cells with ≥ 21 days storage

New cell line development for antibody-producing Chinese hamster ovary cells using split green fluorescent protein

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Kim Yeon-Gu

2012-05-01

Full Text Available Abstract Background The establishment of high producer is an important issue in Chinese hamster ovary (CHO cell culture considering increased heterogeneity by the random integration of a transfected foreign gene and the altered position of the integrated gene. Fluorescence-activated cell sorting (FACS-based cell line development is an efficient strategy for the selection of CHO cells in high therapeutic protein production. Results An internal ribosome entry site (IRES was introduced for using two green fluorescence protein (GFP fragments as a reporter to both antibody chains, the heavy chain and the light chain. The cells co-transfected with two GFP fragments showed the emission of green fluorescence by the reconstitution of split GFP. The FACS-sorted pool with GFP expression had a higher specific antibody productivity (qAb than that of the unsorted pool. The qAb was highly correlated with the fluorescence intensity with a high correlation coefficient, evidenced from the analysis of median GFP and qAb in individual selected clones. Conclusions This study proved that the fragment complementation for split GFP could be an efficient indication for antibody production on the basis of high correlation of qAb with reconstitution of GFP. Taken together, we developed an efficient FACS-based screening method for high antibody-producing CHO cells with the benefits of the split GFP system.

Atypical and classical memory B cells produce Plasmodium falciparum neutralizing antibodies

DEFF Research Database (Denmark)

Muellenbeck, Matthias F; Ueberheide, Beatrix; Amulic, Borko

2013-01-01

signs of active antibody secretion. AtM and CM were also different in their IgG gene repertoire, suggesting that they develop from different precursors. The findings provide direct evidence that natural Pf infection leads to the development of protective memory B cell antibody responses and suggest......Antibodies can protect from Plasmodium falciparum (Pf) infection and clinical malaria disease. However, in the absence of constant reexposure, serum immunoglobulin (Ig) levels rapidly decline and full protection from clinical symptoms is lost, suggesting that B cell memory is functionally impaired...... that constant immune activation rather than impaired memory function leads to the accumulation of AtM in malaria. Understanding the memory B cell response to natural Pf infection may be key to the development of a malaria vaccine that induces long-lived protection....

The effect of xanthine oxidase and hypoxanthine on the permeability of red cells from patients with sickle cell anemia.

Science.gov (United States)

Al Balushi, Halima W M; Rees, David C; Brewin, John N; Hannemann, Anke; Gibson, John S

2018-03-01

Red cells from patients with sickle cell anemia (SCA) are under greater oxidative challenge than those from normal individuals. We postulated that oxidants generated by xanthine oxidase (XO) and hypoxanthine (HO) contribute to the pathogenesis of SCA through altering solute permeability. Sickling, activities of the main red cell dehydration pathways (P sickle , Gardos channel, and KCl cotransporter [KCC]), and cell volume were measured at 100, 30, and 0 mmHg O 2 , together with deoxygenation-induced nonelectrolyte hemolysis. Unexpectedly, XO/HO mixtures had mainly inhibitory effects on sickling, P sickle , and Gardos channel activities, while KCC activity and nonelectrolyte hemolysis were increased. Gardos channel activity was significantly elevated in red cells pharmacologically loaded with Ca 2+ using the ionophore A23187, consistent with an effect on the transport system per se as well as via Ca 2+ entry likely via the P sickle pathway. KCC activity is controlled by several pairs of conjugate protein kinases and phosphatases. Its activity, however, was also stimulated by XO/HO mixtures in red cells pretreated with N-ethylmaleimide (NEM), which is thought to prevent regulation via changes in protein phosphorylation, suggesting that the oxidants formed could also have direct effects on this transporter. In the presence of XO/HO, red cell volume was better maintained in deoxygenated red cells. Overall, the most notable effect of XO/HO mixtures was an increase in red cell fragility. These findings increase our understanding of the effects of oxidative challenge in SCA patients and are relevant to the behavior of red cells in vivo. © 2018 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society.

An electronic apparatus for early detection of changes in red cell ...

African Journals Online (AJOL)

1989-08-19

Aug 19, 1989 ... An electronic apparatus was developed for anaesthetists to use to detect changes in red cell concentration during sur- gery. The mechanism is based on the relationship between the red cell content and the electrical conductivity of blood. In a pilot study of 170 blood samples, a correlation coefficient.

Epigenetics of peripheral B cell differentiation and the antibody response

Directory of Open Access Journals (Sweden)

Hong eZan

2015-12-01

Full Text Available Epigenetic modifications, such as histone post-translational modifications, DNA methylation, and alteration of gene expression by non-coding RNAs, including microRNAs (miRNAs and long non-coding RNAs (lncRNAs, are heritable changes that are independent from the genomic DNA sequence. These regulate gene activities and, therefore, cellular functions. Epigenetic modifications act in concert with transcription factors and play critical roles in B cell development and differentiation, thereby modulating antibody responses to foreign- and self-antigens. Upon antigen encounter by mature B cells in the periphery, alterations of these lymphocytes epigenetic landscape are induced by the same stimuli that drive the antibody response. Such alterations instruct B cells to undergo immunoglobulin class switch DNA recombination (CSR and somatic hypermutation (SHM, as well as differentiation to memory B cells or long-lived plasma cells for the immune memory. Inducible histone modifications, together with DNA methylation and miRNAs modulate the transcriptome, particularly the expression of activation-induced cytidine deaminase (AID, which is essential for CSR and SHM, and factors central to plasma cell differentiation, such as B lymphocyte-induced maturation protein-1 (Blimp-1. These inducible B cell-intrinsic epigenetic marks guide the maturation of antibody responses. Combinatorial histone modifications also function as histone codes to target CSR and, possibly, SHM machinery to the Ig loci by recruiting specific adaptors that can stabilize CSR/SHM factors. In addition, lncRNAs, such as recently reported lncRNA-CSR and an lncRNA generated through transcription of the S region that form G-quadruplex structures, are also important for CSR targeting. Epigenetic dysregulation in B cells, including the aberrant expression of non-coding RNAs and alterations of histone modifications and DNA methylation, can result in aberrant antibody responses to foreign antigens

Rate of red blood cell destruction varies in different strains of mice infected with Plasmodium berghei-ANKA after chronic exposure

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Kikuchi Mihoko

2009-05-01

Full Text Available Abstract Background Severe malaria anaemia in the semi-immune individuals in the holo-endemic area has been observed to occur at low parasite density with individual variation in the responses. Thus the following has been thought to be involved: auto-immune-mediated mechanisms of uninfected red blood cell destruction, and host genetic factors to explain the differences in individual responses under the same malaria transmission. In this study, the extent of red blood cell (RBC destruction in different strains of semi-immune mice model at relatively low parasitaemia was studied. Methodology To generate semi-immunity, four strains of mice were taken through several cycles of infection and treatment. By means of immunofluorescent assay and ELISA, sera were screened for anti-erythrocyte auto-antibodies, and their relationship with haematological parameters and parasitaemia in the strains of semi-immune mice was investigated. Results Upon challenge with Plasmodium berghei ANKA after generating semi-immune status, different mean percentage haemoglobin (Hb drop was observed in the mice strains (Balb/c = 47.1%; NZW = 30.05%; C57BL/6 = 28.44%; CBA = 25.1%, which occurred on different days for each strain (for Balb/c, mean period = 13.6 days; for C57BL/6, NZW, and CBA mean period = 10.6, 10.8, 10.9 days respectively. Binding of antibody to white ghost RBCs was observed in sera of the four strains of semi-immune mice by immunofluorescence. Mean percentage Hb drop per parasitaemia was highest in Balb/c (73.6, followed by C57BL/6 (8.6, CBA (6.9 and NZW (4.0, p = 0.0005. Consequently, auto-antibodies level to ghost RBC were correlated with degree of anaemia and were highest in Balb/c, when compared with the other strains, p Conclusion The results presented in this study seem to indicate that anti-RBC auto-antibodies may be involved in the destruction of uninfected RBC in semi-immune mice at relatively low parasite burden. Host genetic factors may also

Cost comparison of methods for preparation of neonatal red cell aliquots.

Science.gov (United States)

Lechuga, Diana; Thompson, Christina

2007-01-01

The purpose of this study was to compare the preparation costs of two common methods used for neonatal red blood cell transfusion aliquots. Three months of data from a Level 2 and Level 3 neonatal intensive care unit (NICU) were used to determine the comparative cost for red cell aliquot transfusions using an eight bag aliquot/transfer system or the syringe set system. Using leuko-poor red blood cell blood collected in Adsol and containing approximately 320 ml of red blood cells and supernatant solution, the average cost of neonatal transfusion aliquots was determined using the Charter Medical syringe set and the Charter Medical eight bag aliquot/transfer system. A total of 126 red blood cell transfusion aliquots were used over the three month period. The amount transfused with each aliquot ranged from 5.0 ml - 55.0 ml with an average of 24.0 ml per aliquot. The cost per aliquot using the eight aliquot/transfer set was calculated as $36.25 and the cost per aliquot using the syringe set cost was calculated as $30.71. Additional benefits observed with the syringe set included decreased blood waste. When comparing Charter Medical multiple aliquot bag sets and the Charter Medical syringe aliquot system to provide neonatal transfusions, the use of the syringe system decreased blood waste and proved more cost effective.

Engineering cholesterol-based fibers for antibody immobilization and cell capture

Science.gov (United States)

Cohn, Celine

In 2015, the United States is expected to have nearly 600,000 deaths attributed to cancer. Of these 600,000 deaths, 90% will be a direct result of cancer metastasis, the spread of cancer throughout the body. During cancer metastasis, circulating tumor cells (CTCs) are shed from primary tumors and migrate through bodily fluids, establishing secondary cancer sites. As cancer metastasis is incredibly lethal, there is a growing emphasis on developing "liquid biopsies" that can screen peripheral blood, search for and identify CTCs. One popular method for capturing CTCs is the use of a detection platform with antibodies specifically suited to recognize and capture cancer cells. These antibodies are immobilized onto the platform and can then bind and capture cells of interest. However, current means to immobilize antibodies often leave them with drastically reduced function. The antibodies are left poorly suited for cell capture, resulting in low cell capture efficiencies. This body of work investigates the use of lipid-based fibers to immobilize proteins in a way that retains protein function, ultimately leading to increased cell capture efficiencies. The resulting increased efficiencies are thought to arise from the retained three-dimensional structure of the protein as well as having a complete coating of the material surface with antibodies that are capable of interacting with their antigens. It is possible to electrospin cholesterol-based fibers that are similar in design to the natural cell membrane, providing proteins a more natural setting during immobilization. Such fibers have been produced from cholesterol-based cholesteryl succinyl silane (CSS). These fibers have previously illustrated a keen aptitude for retaining protein function and increasing cell capture. Herein the work focuses on three key concepts. First, a model is developed to understand the immobilization mechanism used by electrospun CSS fibers. The antibody immobilization and cell capturing

Nanostructure of Red Blood Cell Membranes in Premature Neonates with Respiratory Distress Syndrome

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S. A. Perepelitsa

2013-01-01

Full Text Available Objective: to study the nanostructure of red blood cell membranes in premature babies with neonatal respiratory distress syndrome (NRDS, by applying atomic force microscopy. Subjects and methods. The investigation included 27 newborn infants, of them 13 premature babies with NRDS formed a study group. The mean gestational age was 33.1±2.3 weeks; their birth weight was 1800±299.3 g. A comparison group consisted of 14 full-term babies with favorable pregnancy and term labor. The mean gestational age of the babies was 39.4±0.5 weeks; their birth weight was 3131.7±588.8 g; the infants had a one minute Apgar score of 8±0.4. Their red blood cells were examined using an atomic force microscope. The objects to be examined were residual umbilical cord blood (RUCB from the premature infants; central venous blood after 7 hours of birth and neonatal venous blood taken on day 7 of life. Results. RUCB from full-term babies contained planocytes that were a major morphological type of red blood cells. In physiological pregnancy and acute fetal hypoxia, the morphological composition of red blood cells in premature neonates with NRDS was close to that in full-term babies. The planocytes are also a major morphological type of red blood cells in the premature infants; the frequency of their occurrence varies. Stomatocytes are typical of all the neonates in the NRDS group; their frequency levels vary greatly: from 8 to 65% of the total number of erythrocytes. The examination revealed that the premature infants of 31—36 weeks gestation were characterized by abnormal erythrocyte shapes that showed a high variability. At birth, the premature babies were found to have changes in the nanostructure of red blood cell membranes, which were influenced by intrauterine hypoxia. The first-order value reflecting flickering in the red blood cell membrane varies to the most extent. Conclusion. Atomic force microscopy showed that the greatest changes in the structure of red

Targeting to cells of fluorescent liposomes covalently coupled with monoclonal antibody or protein A

Science.gov (United States)

Leserman, Lee D.; Barbet, Jacques; Kourilsky, François; Weinstein, John N.

1980-12-01

Many applications envisioned for liposomes in cell biology and chemotherapy require their direction to specific cellular targets1-3. The ability to use antibody as a means of conferring specificity to liposomes would markedly increase their usefulness. We report here a method for covalently coupling soluble proteins, including monoclonal antibody and Staphylococcus aureus protein A (ref. 4), to small sonicated liposomes, by using the heterobifunctional cross-linking reagent N-hydroxysuccinimidyl 3-(2-pyridyldithio)propionate (SPDP, Pharmacia). Liposomes bearing covalently coupled mouse monoclonal antibody against human β2-microglobulin [antibody B1.1G6 (IgG2a, κ) (B. Malissen et al., in preparation)] bound specifically to human, but not to mouse cells. Liposomes bearing protein A became bound to human cells previously incubated with the B1.1G6 antibody, but not to cells incubated without antibody. The coupling method results in efficient binding of protein to the liposomes without aggregation and without denaturation of the coupled ligand; at least 60% of liposomes bound functional protein. Further, liposomes did not leak encapsulated carboxyfluorescein (CF) as a consequence of the reaction.

Impaired antibody response causes persistence of prototypic T cell-contained virus.

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Andreas Bergthaler

2009-04-01

Full Text Available CD8 T cells are recognized key players in control of persistent virus infections, but increasing evidence suggests that assistance from other immune mediators is also needed. Here, we investigated whether specific antibody responses contribute to control of lymphocytic choriomeningitis virus (LCMV, a prototypic mouse model of systemic persistent infection. Mice expressing transgenic B cell receptors of LCMV-unrelated specificity, and mice unable to produce soluble immunoglobulin M (IgM exhibited protracted viremia or failed to resolve LCMV. Virus control depended on immunoglobulin class switch, but neither on complement cascades nor on Fc receptor gamma chain or Fc gamma receptor IIB. Cessation of viremia concurred with the emergence of viral envelope-specific antibodies, rather than with neutralizing serum activity, and even early nonneutralizing IgM impeded viral persistence. This important role for virus-specific antibodies may be similarly underappreciated in other primarily T cell-controlled infections such as HIV and hepatitis C virus, and we suggest this contribution of antibodies be given consideration in future strategies for vaccination and immunotherapy.

Labelling of red blood cells with 99m pertechnetate

International Nuclear Information System (INIS)

Vyth, A.; Raam, C.F.

1979-07-01

This paper describes a method for labelling red blood cells with 99mTc in vitro, using electrolytically generated stannous ions as the reducing agent for 99mTc-pertechnetate. A labelling of 95% was found. A method for the in vivo labelling of red blood cells is also reported. This involves an injection of a stanno-DTPA-complex followed 20 minutes later by a 99mTc-pertechnetate solution scintillation camera images show more background activity when the in vivo method of labelling is used

Minimal RED Cell Pairs Markedly Improve Electrode Kinetics and Power Production in Microbial Reverse Electrodialysis Cells

KAUST Repository

Cusick, Roland D.

2013-12-17

Power production from microbial reverse electrodialysis cell (MRC) electrodes is substantially improved compared to microbial fuel cells (MFCs) by using ammonium bicarbonate (AmB) solutions in multiple RED cell pair stacks and the cathode chamber. Reducing the number of RED membranes pairs while maintaining enhanced electrode performance could help to reduce capital costs. We show here that using only a single RED cell pair (CP), created by operating the cathode in concentrated AmB, dramatically increased power production normalized to cathode area from both acetate (Acetate: from 0.9 to 3.1 W/m 2-cat) and wastewater (WW: 0.3 to 1.7 W/m2), by reducing solution and charge transfer resistances at the cathode. A second RED cell pair increased RED stack potential and reduced anode charge transfer resistance, further increasing power production (Acetate: 4.2 W/m2; WW: 1.9 W/m2). By maintaining near optimal electrode power production with fewer membranes, power densities normalized to total membrane area for the 1-CP (Acetate: 3.1 W/m2-mem; WW: 1.7 W/m2) and 2-CP (Acetate: 1.3 W/m2-mem; WW: 0.6 W/m2) reactors were much higher than previous MRCs (0.3-0.5 W/m2-mem with acetate). While operating at peak power, the rate of wastewater COD removal, normalized to reactor volume, was 30-50 times higher in 1-CP and 2-CP MRCs than that in a single chamber MFC. These findings show that even a single cell pair AmB RED stack can significantly enhance electrical power production and wastewater treatment. © 2013 American Chemical Society.

Minimal RED Cell Pairs Markedly Improve Electrode Kinetics and Power Production in Microbial Reverse Electrodialysis Cells

KAUST Repository

Cusick, Roland D.; Hatzell, Marta; Zhang, Fang; Logan, Bruce E.

2013-01-01

Power production from microbial reverse electrodialysis cell (MRC) electrodes is substantially improved compared to microbial fuel cells (MFCs) by using ammonium bicarbonate (AmB) solutions in multiple RED cell pair stacks and the cathode chamber. Reducing the number of RED membranes pairs while maintaining enhanced electrode performance could help to reduce capital costs. We show here that using only a single RED cell pair (CP), created by operating the cathode in concentrated AmB, dramatically increased power production normalized to cathode area from both acetate (Acetate: from 0.9 to 3.1 W/m 2-cat) and wastewater (WW: 0.3 to 1.7 W/m2), by reducing solution and charge transfer resistances at the cathode. A second RED cell pair increased RED stack potential and reduced anode charge transfer resistance, further increasing power production (Acetate: 4.2 W/m2; WW: 1.9 W/m2). By maintaining near optimal electrode power production with fewer membranes, power densities normalized to total membrane area for the 1-CP (Acetate: 3.1 W/m2-mem; WW: 1.7 W/m2) and 2-CP (Acetate: 1.3 W/m2-mem; WW: 0.6 W/m2) reactors were much higher than previous MRCs (0.3-0.5 W/m2-mem with acetate). While operating at peak power, the rate of wastewater COD removal, normalized to reactor volume, was 30-50 times higher in 1-CP and 2-CP MRCs than that in a single chamber MFC. These findings show that even a single cell pair AmB RED stack can significantly enhance electrical power production and wastewater treatment. © 2013 American Chemical Society.

An electronic apparatus for early detection of changes in red cell ...

African Journals Online (AJOL)

An electronic apparatus was developed for anaesthetists to use to detect changes in red cell concentration during surgery. The mechanism is based on the relationship between the red cell content and the electrical conductivity of blood. In a pilot study of 170 blood samples, a correlation coefficient of 0,9806 was obtained ...

Optically-driven red blood cell rotor in linearly polarized laser tweezers

Indian Academy of Sciences (India)

We have constructed a dual trap optical tweezers set-up around an inverted microscope where both the traps can be independently controlled and manipulated in all the three dimensions. Here we report our observations on rotation of red blood cells (RBCs) in a linearly polarized optical trap. Red blood cells deform and ...

Impact of Uniform Methods on Interlaboratory Antibody Titration Variability: Antibody Titration and Uniform Methods.

Science.gov (United States)

Bachegowda, Lohith S; Cheng, Yan H; Long, Thomas; Shaz, Beth H

2017-01-01

-Substantial variability between different antibody titration methods prompted development and introduction of uniform methods in 2008. -To determine whether uniform methods consistently decrease interlaboratory variation in proficiency testing. -Proficiency testing data for antibody titration between 2009 and 2013 were obtained from the College of American Pathologists. Each laboratory was supplied plasma and red cells to determine anti-A and anti-D antibody titers by their standard method: gel or tube by uniform or other methods at different testing phases (immediate spin and/or room temperature [anti-A], and/or anti-human globulin [AHG: anti-A and anti-D]) with different additives. Interlaboratory variations were compared by analyzing the distribution of titer results by method and phase. -A median of 574 and 1100 responses were reported for anti-A and anti-D antibody titers, respectively, during a 5-year period. The 3 most frequent (median) methods performed for anti-A antibody were uniform tube room temperature (147.5; range, 119-159), uniform tube AHG (143.5; range, 134-150), and other tube AHG (97; range, 82-116); for anti-D antibody, the methods were other tube (451; range, 431-465), uniform tube (404; range, 382-462), and uniform gel (137; range, 121-153). Of the larger reported methods, uniform gel AHG phase for anti-A and anti-D antibodies had the most participants with the same result (mode). For anti-A antibody, 0 of 8 (uniform versus other tube room temperature) and 1 of 8 (uniform versus other tube AHG), and for anti-D antibody, 0 of 8 (uniform versus other tube) and 0 of 8 (uniform versus other gel) proficiency tests showed significant titer variability reduction. -Uniform methods harmonize laboratory techniques but rarely reduce interlaboratory titer variance in comparison with other methods.

Anti-S100A4 antibody suppresses metastasis formation by blocking stroma cell invasion

DEFF Research Database (Denmark)

Klingelhöfer, Jörg; Grum-Schwensen, Birgitte; Beck, Mette K

2012-01-01

microenvironment, making it an attractive target for anti-cancer therapy. In this study, we produced a function-blocking anti-S100A4 monoclonal antibody with metastasis-suppressing activity. Antibody treatment significantly reduced metastatic burden in the lungs of experimental animals by blocking the recruitment...... of T cells to the site of the primary tumor. In vitro studies demonstrated that this antibody efficiently reduced the invasion of T cells in a fibroblast monolayer. Moreover, it was capable of suppressing the invasive growth of human and mouse fibroblasts. We presume therefore that the antibody exerts...... its activity by suppressing stroma cell recruitment to the site of the growing tumor. Our epitope mapping studies suggested that the antibody recognition site overlaps with the target binding interface of human S100A4. We conclude here that this antibody could serve as a solid basis for development...

Red blood cells, still vital after all these years: Commentary on Canadian Blood Services' International Symposium 2017.

Science.gov (United States)

Qadri, Syed M; Donkor, David A; Yan, Matthew; Ning, Shuoyan; Branch, Donald R; Seghatchian, Jerard; Sheffield, William P

2018-04-01

Canadian Blood Services (CBS), Canada's national blood transfusion service, has for many years sponsored an annual conference, for the education and awareness of interested participants, showcasing the latest evidence-based understanding of both basic science and clinical issues in transfusion medicine and science. The 15th iteration of this symposium took place September 9, 2017 and focused on some of the vital aspects of red blood cells (RBC), in line with the" 3Rs" concept, namely the provision of the Right red blood cell (RBC) product to the Right patient at the Right time. Presentations touched upon: the evolution of blood banking in North America; the monocyte monolayer assay as a predictor of post-transfusion hemolysis; hemoglobin-based oxygen carriers; RBC alloimmunization; serological approaches to complex RBC antibody problems; randomized clinical trials related to the age of stored RBC; RBC genotyping; pathophysiology, prevention and treatment of hemolytic disease of the fetus and newborn (HDFN); and testing and timing in perinatal serology. This commentary provides summaries of all speakers' presentations annotated with relevant references. Special thanks are due to all contributors for their praiseworthy approaches in sharing their experiences and knowledge on this interesting scientific/clinical and management theme. Copyright © 2018 Elsevier Ltd. All rights reserved.

Defective TFH Cell Function and Increased TFR Cells Contribute to Defective Antibody Production in Aging.

Science.gov (United States)

Sage, Peter T; Tan, Catherine L; Freeman, Gordon J; Haigis, Marcia; Sharpe, Arlene H

2015-07-14

Defective antibody production in aging is broadly attributed to immunosenescence. However, the precise immunological mechanisms remain unclear. Here, we demonstrate an increase in the ratio of inhibitory T follicular regulatory (TFR) cells to stimulatory T follicular helper (TFH) cells in aged mice. Aged TFH and TFR cells are phenotypically distinct from those in young mice, exhibiting increased programmed cell death protein-1 expression but decreased ICOS expression. Aged TFH cells exhibit defective antigen-specific responses, and programmed cell death protein-ligand 1 blockade can partially rescue TFH cell function. In contrast, young and aged TFR cells have similar suppressive capacity on a per-cell basis in vitro and in vivo. Together, these studies reveal mechanisms contributing to defective humoral immunity in aging: an increase in suppressive TFR cells combined with impaired function of aged TFH cells results in reduced T-cell-dependent antibody responses in aged mice. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Prevalence of Diego blood group antigen and the antibody in three ethnic population groups in Klang valley of Malaysia

Directory of Open Access Journals (Sweden)

Cheong Tar Wei

2013-01-01

Full Text Available Background: Diego blood group antigen, Di(a, is very rare among Caucasians and Blacks, but relatively common among the South American Indians and Asians of Mongolian origin. The antibody to Di(a is clinically significant to cause hemolytic disease in a new-born or hemolytic transfusion reaction. Objectives: This study was designed to determine the prevalence of Di(a antigen among the blood donors from the three major ethnic groups in Klang Valley of Malaysia as well as to find an incidence of an antibody of the Diego antigen, anti-Di(a, in a tertiary care hospital to ascertain the need to include Di(a+ red cells for an antibody screen cell panel. Materials and Methods: Serological tests were performed by column agglutination technique using commercial reagents and following instruction as per kit insert. Results: Di(a antigen was found with a frequency of 2.1% among the Malaysians donors in three ethnic groups viz, Malay, Chinese and Indian. It was present among 1.25% of 401 Malay, 4.01% of Chinese and 0.88% of 114 Indian origin donors. None of the 1442 patients, including 703 antenatal outpatients, had anti-Di(a in serum. Conclusion: The prevalence of Di(a antigen was found among the donors of all the three ethnic background with varying frequency. Inclusion of Di(a+ red cells in routine antibody screening program would certainly help in detection of this clinically significant antibody and to provide safe blood transfusion in the Klang Valley, though the incidence of antibody appears to be very low in the region.

Prevalence of Diego blood group antigen and the antibody in three ethnic population groups in Klang valley of Malaysia.

Science.gov (United States)

Wei, Cheong Tar; Al-Hassan, Faisal Muti; Naim, Norris; Knight, Aishah; Joshi, Sanmukh R

2013-01-01

Diego blood group antigen, Di(a), is very rare among Caucasians and Blacks, but relatively common among the South American Indians and Asians of Mongolian origin. The antibody to Di(a) is clinically significant to cause hemolytic disease in a new-born or hemolytic transfusion reaction. This study was designed to determine the prevalence of Di(a) antigen among the blood donors from the three major ethnic groups in Klang Valley of Malaysia as well as to find an incidence of an antibody of the Diego antigen, anti-Di(a), in a tertiary care hospital to ascertain the need to include Di(a+) red cells for an antibody screen cell panel. Serological tests were performed by column agglutination technique using commercial reagents and following instruction as per kit insert. Di(a) antigen was found with a frequency of 2.1% among the Malaysians donors in three ethnic groups viz, Malay, Chinese and Indian. It was present among 1.25% of 401 Malay, 4.01% of Chinese and 0.88% of 114 Indian origin donors. None of the 1442 patients, including 703 antenatal outpatients, had anti-Di(a) in serum. The prevalence of Di(a) antigen was found among the donors of all the three ethnic background with varying frequency. Inclusion of Di(a+) red cells in routine antibody screening program would certainly help in detection of this clinically significant antibody and to provide safe blood transfusion in the Klang Valley, though the incidence of antibody appears to be very low in the region.

Intravenous immunoglobulin in the management of a rare cause of hemolytic disease of the newborn: Anti-SARA antibodies.

Science.gov (United States)

Venkataraman, Rohini; Yusuf, Kamran

2017-01-01

Hemolytic disease of newborn (HDN) is a condition that develops in a fetus, when the IgG molecules produced by the mother pass through the placenta and attack the fetal red blood cells. HDN can occur due to Rh and ABO incompatibilities between the mother and the fetus as well as due to other allo-immune antibodies belonging to Kell (K and k), Duffy (Fya), Kidd (Jka and Jkb), and MNS (M, N, S, and s) systems. Role of intravenous immunoglobulin in management of HDN is not clear.SARA red blood cell antigen, first discovered in 1990 is a low frequency antigen. We report, a multiparous female whose pregnancy was complicated by HDN due to anti-SARA antibodies requiring both exchange transfusion and intravenous immunoglobulin. The response was sustained after intravenous immunoglobulin (IVIG) rather than after exchange transfusion.

Survival of transfused red blood cells: In vivo compatibility testing with chromium-51

International Nuclear Information System (INIS)

Dharkar, D.D.; Pineda, A.A.

1983-01-01

The /sup 51/Cr red cell survival test and specific test for measurement of the disappearance rate of labeled red cells. This procedure can be used for the assessment of red cell compatibility testing in vivo. The authors recommend that more routine transfusions as well as ''difficult'' transfusions be monitored by /sup 51/Cr in vivo compatibility testing before the actual transfusions, so that more consistent and reliable survival values are achieved

EVALUATION OF ANAEMIA USING RED CELL AND RETICULOCYTE PARAMETERS USING AUTOMATED HAEMATOLOGY ANALYSER

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Vidyadhar Rao

2016-06-01

Full Text Available Use of current models of Automated Haematology Analysers help in calculating the haemoglobin contents of the mature Red cells, Reticulocytes and percentages of Microcytic and hypochromic Red cells. This has helped the clinician in reaching early diagnosis and management of Different haemopoietic disorders like Iron Deficiency Anaemia, Thalassaemia and anaemia of chronic diseases. AIM This study is conducted using an Automated Haematology Analyser to evaluate anaemia using the Red Cell and Reticulocyte parameters. Three types of anaemia were evaluated; iron deficiency anaemia, anaemia of long duration and anaemia associated with chronic disease and Iron deficiency. MATERIALS AND METHODS The blood samples were collected from 287 adult patients with anaemia differentiated depending upon their iron status, haemoglobinopathies and inflammatory activity. Iron deficiency anaemia (n=132, anaemia of long duration (ACD, (n=97 and anaemia associated with chronic disease with iron deficiency (ACD Combi, (n=58. Microcytic Red cells, hypochromic red cells percentage and levels of haemoglobin in reticulocytes and matured RBCs were calculated. The accuracy of the parameters was analysed using receiver operating characteristic analyser to differentiate between the types of anaemia. OBSERVATIONS AND RESULTS There was no difference in parameters between the iron deficiency group or anaemia associated with chronic disease and iron deficiency. The hypochromic red cells percentage was the best parameter in differentiating anaemia of chronic disease with or without absolute iron deficiency with a sensitivity of 72.7% and a specificity of 70.4%. CONCLUSIONS The parameters of red cells and reticulocytes were of reasonably good indicators in differentiating the absolute iron deficiency anaemia with chronic disease.

Genetic engineering of cell lines using lentiviral vectors to achieve antibody secretion following encapsulated implantation.

Science.gov (United States)

Lathuilière, Aurélien; Bohrmann, Bernd; Kopetzki, Erhard; Schweitzer, Christoph; Jacobsen, Helmut; Moniatte, Marc; Aebischer, Patrick; Schneider, Bernard L

2014-01-01

The controlled delivery of antibodies by immunoisolated bioimplants containing genetically engineered cells is an attractive and safe approach for chronic treatments. To reach therapeutic antibody levels there is a need to generate renewable cell lines, which can long-term survive in macroencapsulation devices while maintaining high antibody specific productivity. Here we have developed a dual lentiviral vector strategy for the genetic engineering of cell lines compatible with macroencapsulation, using separate vectors encoding IgG light and heavy chains. We show that IgG expression level can be maximized as a function of vector dose and transgene ratio. This approach allows for the generation of stable populations of IgG-expressing C2C12 mouse myoblasts, and for the subsequent isolation of clones stably secreting high IgG levels. Moreover, we demonstrate that cell transduction using this lentiviral system leads to the production of a functional glycosylated antibody by myogenic cells. Subsequent implantation of antibody-secreting cells in a high-capacity macroencapsulation device enables continuous delivery of recombinant antibodies in the mouse subcutaneous tissue, leading to substantial levels of therapeutic IgG detectable in the plasma.

Interplay between Natural Killer Cells and Anti-HER2 Antibodies: Perspectives for Breast Cancer Immunotherapy

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Aura Muntasell

2017-11-01

Full Text Available Overexpression of the human epidermal growth factor receptor 2 (HER2 defines a subgroup of breast tumors with aggressive behavior. The addition of HER2-targeted antibodies (i.e., trastuzumab, pertuzumab to chemotherapy significantly improves relapse-free and overall survival in patients with early-stage and advanced disease. Nonetheless, considerable proportions of patients develop resistance to treatment, highlighting the need for additional and co-adjuvant therapeutic strategies. HER2-specific antibodies can trigger natural killer (NK cell-mediated antibody-dependent cellular cytotoxicity and indirectly enhance the development of tumor-specific T cell immunity; both mechanisms contributing to their antitumor efficacy in preclinical models. Antibody-dependent NK cell activation results in the release of cytotoxic granules as well as the secretion of pro-inflammatory cytokines (i.e., IFNγ and TNFα and chemokines. Hence, NK cell tumor suppressive functions include direct cytolytic killing of tumor cells as well as the regulation of subsequent antitumor adaptive immunity. Albeit tumors with gene expression signatures associated to the presence of cytotoxic lymphocyte infiltrates benefit from trastuzumab-based treatment, NK cell-related biomarkers of response/resistance to HER2-specific therapeutic antibodies in breast cancer patients remain elusive. Several variables, including (i the configuration of the patient NK cell repertoire; (ii tumor molecular features (i.e., estrogen receptor expression; (iii concomitant therapeutic regimens (i.e., chemotherapeutic agents, tyrosine kinase inhibitors; and (iv evasion mechanisms developed by progressive breast tumors, have been shown to quantitatively and qualitatively influence antibody-triggered NK cell responses. In this review, we discuss possible interventions for restoring/enhancing the therapeutic activity of HER2 therapeutic antibodies by harnessing NK cell antitumor potential through

Screening hypochromism (sieve effect) in red blood cells: a quantitative analysis.

Science.gov (United States)

Razi Naqvi, K

2014-04-01

Multiwavelength UV-visible spectroscopy, Kramers-Kronig analysis, and several other experimental and theoretical tools have been applied over the last several decades to fathom absorption and scattering of light by suspensions of micron-sized pigmented particles, including red blood cells, but a satisfactory quantitative analysis of the difference between the absorption spectra of suspension of intact and lysed red blood cells is still lacking. It is stressed that such a comparison is meaningful only if the pertinent spectra are free from, or have been corrected for, scattering losses, and it is shown that Duysens' theory can, whereas that of Vekshin cannot, account satisfactorily for the observed hypochromism of suspensions of red blood cells.

Light scattering by red blood cells in ektacytometry: Fraunhofer versus anomalous diffraction

NARCIS (Netherlands)

Streekstra, G. J.; Hoekstra, A. G.; Nijhof, E. J.; Heethaar, R. M.

1993-01-01

In the present literature on ektacytometry, small angle light scattering by ellipsoidal red blood cells is commonly approximated by Fraunhofer diffraction. Calculations on a sphere with the size and relative refractive index of a red cell, however, show that Fraunhofer diffraction deviates

Ebola virus-like particles produced in insect cells exhibit dendritic cell stimulating activity and induce neutralizing antibodies

International Nuclear Information System (INIS)

Ye Ling; Lin Jianguo; Sun Yuliang; Bennouna, Soumaya; Lo, Michael; Wu Qingyang; Bu Zhigao; Pulendran, Bali; Compans, Richard W.; Yang Chinglai

2006-01-01

Recombinant baculoviruses (rBV) expressing Ebola virus VP40 (rBV-VP40) or GP (rBV-GP) proteins were generated. Infection of Sf9 insect cells by rBV-VP40 led to assembly and budding of filamentous particles from the cell surface as shown by electron microscopy. Ebola virus-like particles (VLPs) were produced by coinfection of Sf9 cells with rBV-VP40 and rBV-GP, and incorporation of Ebola GP into VLPs was demonstrated by SDS-PAGE and Western blot analysis. Recombinant baculovirus infection of insect cells yielded high levels of VLPs, which were shown to stimulate cytokine secretion from human dendritic cells similar to VLPs produced in mammalian cells. The immunogenicity of Ebola VLPs produced in insect cells was evaluated by immunization of mice. Analysis of antibody responses showed that most of the GP-specific antibodies were of the IgG2a subtype, while no significant level of IgG1 subtype antibodies specific for GP was induced, indicating the induction of a Th1-biased immune response. Furthermore, sera from Ebola VLP immunized mice were able to block infection by Ebola GP pseudotyped HIV virus in a single round infection assay, indicating that a neutralizing antibody against the Ebola GP protein was induced. These results show that production of Ebola VLPs in insect cells using recombinant baculoviruses represents a promising approach for vaccine development against Ebola virus infection

Effect of high dose irradiation on the red cell span in rabbits

International Nuclear Information System (INIS)

Kang, T.W.; Koh, J.W.; Woo, K.S.; Lee, O.H.; Youn, C.S.

1982-01-01

As a part of studies on acute effects of high dose irradiation in vivo, the present report was carried out to evaluate the changes of the red cell life span in the white rabbits by a single whole body exposure to gamma rays from 60 Co teletherapy unit. The exposure was done in dose levels of 100, 600 and 900 rads to each experimental group of 10 rabbits. The life span apparent half survival time of red cells, and that the red cell volume in the circulting blood were measured by ICSH Reference method using 51 Cr. (Author)

Monitoring the systemic human memory B cell compartment of melanoma patients for anti-tumor IgG antibodies.

Directory of Open Access Journals (Sweden)

Amy E Gilbert

Full Text Available Melanoma, a potentially lethal skin cancer, is widely thought to be immunogenic in nature. While there has been much focus on T cell-mediated immune responses, limited knowledge exists on the role of mature B cells. We describe an approach, including a cell-based ELISA, to evaluate mature IgG antibody responses to melanoma from human peripheral blood B cells. We observed a significant increase in antibody responses from melanoma patients (n = 10 to primary and metastatic melanoma cells compared to healthy volunteers (n = 10 (P<0.0001. Interestingly, we detected a significant reduction in antibody responses to melanoma with advancing disease stage in our patient cohort (n = 21 (P<0.0001. Overall, 28% of melanoma patient-derived B cell cultures (n = 1,800 compared to 2% of cultures from healthy controls (n = 600 produced antibodies that recognized melanoma cells. Lastly, a patient-derived melanoma-specific monoclonal antibody was selected for further study. This antibody effectively killed melanoma cells in vitro via antibody-mediated cellular cytotoxicity. These data demonstrate the presence of a mature systemic B cell response in melanoma patients, which is reduced with disease progression, adding to previous reports of tumor-reactive antibodies in patient sera, and suggesting the merit of future work to elucidate the clinical relevance of activating humoral immune responses to cancer.

Monitoring the Systemic Human Memory B Cell Compartment of Melanoma Patients for Anti-Tumor IgG Antibodies

Science.gov (United States)

Gilbert, Amy E.; Karagiannis, Panagiotis; Dodev, Tihomir; Koers, Alexander; Lacy, Katie; Josephs, Debra H.; Takhar, Pooja; Geh, Jenny L. C.; Healy, Ciaran; Harries, Mark; Acland, Katharine M.; Rudman, Sarah M.; Beavil, Rebecca L.; Blower, Philip J.; Beavil, Andrew J.; Gould, Hannah J.; Spicer, James; Nestle, Frank O.; Karagiannis, Sophia N.

2011-01-01

Melanoma, a potentially lethal skin cancer, is widely thought to be immunogenic in nature. While there has been much focus on T cell-mediated immune responses, limited knowledge exists on the role of mature B cells. We describe an approach, including a cell-based ELISA, to evaluate mature IgG antibody responses to melanoma from human peripheral blood B cells. We observed a significant increase in antibody responses from melanoma patients (n = 10) to primary and metastatic melanoma cells compared to healthy volunteers (n = 10) (P<0.0001). Interestingly, we detected a significant reduction in antibody responses to melanoma with advancing disease stage in our patient cohort (n = 21) (P<0.0001). Overall, 28% of melanoma patient-derived B cell cultures (n = 1,800) compared to 2% of cultures from healthy controls (n = 600) produced antibodies that recognized melanoma cells. Lastly, a patient-derived melanoma-specific monoclonal antibody was selected for further study. This antibody effectively killed melanoma cells in vitro via antibody-mediated cellular cytotoxicity. These data demonstrate the presence of a mature systemic B cell response in melanoma patients, which is reduced with disease progression, adding to previous reports of tumor-reactive antibodies in patient sera, and suggesting the merit of future work to elucidate the clinical relevance of activating humoral immune responses to cancer. PMID:21559411

Comprehensive Cross-Clade Characterization of Antibody-Mediated Recognition, Complement-Mediated Lysis, and Cell-Mediated Cytotoxicity of HIV-1 Envelope-Specific Antibodies toward Eradication of the HIV-1 Reservoir.

Science.gov (United States)

Mujib, Shariq; Liu, Jun; Rahman, A K M Nur-Ur; Schwartz, Jordan A; Bonner, Phil; Yue, Feng Yun; Ostrowski, Mario A

2017-08-15

Immunotherapy with passive administration of broadly neutralizing HIV-1 envelope-specific antibodies (bnAbs) in the setting of established infection in vivo has yielded mixed results. The contribution of different antibodies toward the direct elimination of infected cells is poorly understood. In this study, we determined the ability of 12 well-characterized anti-HIV-1 neutralizing antibodies to recognize and eliminate primary CD4 T cells infected with HIV-1 belonging to clades A, B, C, and D, via antibody-dependent complement-mediated lysis (ADCML) and antibody-dependent cell-mediated cytotoxicity (ADCC), in vitro We further tested unique combinations of these antibodies to determine the optimal antibody cocktails to be tested in future clinical trials. We report that antibody binding to infected CD4 T cells is highly variable and correlates with ADCML and ADCC processes. Particularly, antibodies targeting the envelope glycan shield (2G12) and V1/V2 site (PG9, PG16, and PGT145) are best at recognizing HIV-1-infected CD4 T cells. However, only PG9 and PG16 and their combinations with other bnAbs sufficiently induced the elimination of HIV-1-infected CD4 T cells by ADCML, ADCC, or both. Notably, CD4 binding site antibodies VRC01, 3BNC117, and NIH45-46 G54W did not exhibit recognition of infected cells and were unable to induce their killing. Future trials geared toward the development of a cure for HIV/AIDS should incorporate V1/V2 antibodies for maximal clearance of infected cells. With the use of only primary immune cells, we conducted a comprehensive cross-clade physiological analysis to aid the direction of antibodies as therapeutics toward the development of a cure for HIV/AIDS. IMPORTANCE Several antibodies capable of neutralizing the majority of circulating HIV-1 strains have been identified to date and have been shown to prevent infection in animal models. However, the use of combinations of such broadly neutralizing antibodies (bnAbs) for the treatment and

Deformation of Two-Dimensional Nonuniform-Membrane Red Blood Cells Simulated by a Lattice Boltzmann Model

International Nuclear Information System (INIS)

Hua-Bing, Li; Li, Jin; Bing, Qiu

2008-01-01

To study two-dimensional red blood cells deforming in a shear Bow with the membrane nonuniform on the rigidity and mass, the membrane is discretized into equilength segments. The fluid inside and outside the red blood cell is simulated by the D2Q9 lattice Boltzmann model and the hydrodynamic forces exerted on the membrane from the inner and outer of the red blood cell are calculated by a stress-integration method. Through the global deviation from the curvature of uniform-membrane, we find that when the membrane is nonuniform on the rigidity, the deviation first decreases with the time increases and implies that the terminal profile of the red blood cell is static. To a red blood cell with the mass nonuniform on the membrane, the deviation becomes more large, and the mass distribution affects the profile of the two sides of the flattened red blood cell in a shear flow. (fundamental areas of phenomenology(including applications))

Mechanical and electrical properties of red blood cells using optical tweezers

International Nuclear Information System (INIS)

Fontes, A; Castro, M L Barjas; Brandão, M M; Fernandes, H P; Huruta, R R; Costa, F F; Saad, S T O; Thomaz, A A; Pozzo, L Y; Barbosa, L C; Cesar, C L

2011-01-01

Optical tweezers are a very sensitive tool, based on photon momentum transfer, for individual, cell by cell, manipulation and measurements, which can be applied to obtain important properties of erythrocytes for clinical and research purposes. Mechanical and electrical properties of erythrocytes are critical parameters for stored cells in transfusion centers, immunohematological tests performed in transfusional routines and in blood diseases. In this work, we showed methods, based on optical tweezers, to study red blood cells and applied them to measure apparent overall elasticity, apparent membrane viscosity, zeta potential, thickness of the double layer of electrical charges and adhesion in red blood cells

Facilitation of syngeneic stem cell engraftment by anti-class I monoclonal antibody pretreatment of unirradiated recipients

International Nuclear Information System (INIS)

Voralia, M.; Semeluk, A.; Wegmann, T.G.

1987-01-01

We have established a murine model of syngeneic bone marrow transplantation based on the use of monoclonal antibody as the sole conditioning regimen in unirradiated recipients. Administration of a single injection of monoclonal antibody directed against major histocompatibility complex-encoded class I determinants facilitated permanent hemopoietic stem cell engraftment without any apparent side-effects. Whereas untreated hosts exhibited a maximal chimerism of 15% at donor cell doses of up to 12 X 10(7) bone marrow cells, pretreatment by 2 mg of anti-class I antibody one week prior to transplantation of 3 X 10(7) syngeneic bone marrow cells resulted in a mean donor representation of about 80%. The antibody can be given up to four weeks prior to transplantation, and the degree of donor engraftment observed is a function of the dose of antibody administered. The fact that specific antibody enhanced engraftment in two strain combinations indicates that antibody is the active agent in facilitating engraftment and that facilitation is not strain-restricted. Anti-class I antibodies of the IgG2a, but not IgG1, isotype are effective in promoting engraftment. Although the isotype requirement suggests a role for antibody-mediated cytotoxicity in promoting stem cell engraftment, the extensive time-frame of facilitation suggests that other effects of the antibody may also be involved. The model of syngeneic bone marrow transplantation we describe here will be useful in studying the mechanisms regulating stem cell engraftment and may have potential clinical application as an approach to autologous marrow transplantation

Artificial Red Cells with Polyhemoglobin Membranes.

Science.gov (United States)

1981-09-01

Y. Reciprocal binding of oxygen and diphosphoglycerate by human hemoglobin. Proc. Natl. Acad. Sci. USA 59:526-532, 1968. 12. Bunn, H. F., Seal, U. S...and Scott, A. F. The role of 2,3- diphosphoglycerate in mediating hemoglobin function of mammalian red cells. Ann. N.Y. Acad. Sciences 241:498-519

Antibody-mediated allotype suppression in adult mice: the role of antigen, effector isotype and regulatory T cells.

Science.gov (United States)

Curling, E M; Dresser, D W

1984-10-01

It has been reported (Contemp. Top. Immunobiol. 1974. 3:41) that allotype-specific T suppressor cells can be induced after monoclonal anti-allotype treatment of neonatal (BALB/c X SJL)F1 (Igha/b) mice. Here we show that (BALB/c X CB20)F1 adult-derived spleen cells (SC) are, by contrast, potently suppressed by monoclonal allotype-specific reagents, (when transferred into irradiated BALB/c recipients) in the absence of primary T suppressor cell induction. Such suppression is only induced in activated B cells [exposed to lipopolysaccharide or sheep red blood cells (SRBC)], and is probably dependent on the isotype of the anti-allotype sera administered. For example, two independently produced IgG1 monoclonal reagents raised against the Igh-1b allotype were poorly suppressive or nonsuppressive, whereas an IgG3 and an IgG2a monoclonal antibody induced a 90% suppression of the target allotype in transferred adult SC. It was found that suppression was not due to a depletion of antigen-specific T cell help since: (a) the addition of SRBC-educated T cells did not break suppression and (b) suppressed SC were as good a source of T cell help as normal SC, in the response of virgin or memory B cell (Thy-1-depleted) responses to SRBC in vivo. Suppression was maintained in suppressed cells which had been rechallenged with SRBC after transfer into a second irradiated recipient, but was not induced in normal SC when these were admixed with an equal number from this suppressed SC population. These findings point to a possible mechanism for the regulation of B cell expression, through the formation of an antibody-Ig receptor complex at the surface of the B lymphocyte. After complexing the target cell is either deleted or inactivated. The response to SRBC was reduced or ablated for at least 70 days after treatment with a single dose of anti-allotype serum.

Red blood cell and platelet genotyping: from current practice to future high-throughput donor typing

NARCIS (Netherlands)

de Haas, M.; van der Schoot, C. E.; Beiboer, S. H. W.; Feskens, M.; Cheroutre, G.; Maaskant-van Wijkb, P. A.

2006-01-01

The molecular basis of almost all red cell and platelet blood group antigens is known. This enables the prediction of red cell or platelet phenotypes based upon the genotypes. In many laboratories, blood group genotyping assays are routinely used in cases where patient red cells cannot be used for

Antiviral Activity of HIV gp120 Targeting Bispecific T Cell Engager (BiTE®) Antibody Constructs.

Science.gov (United States)

Brozy, Johannes; Schlaepfer, Erika; Mueller, Christina K S; Rochat, Mary-Aude; Rampini, Silvana K; Myburgh, Renier; Raum, Tobias; Kufer, Peter; Baeuerle, Patrick A; Muenz, Markus; Speck, Roberto F

2018-05-02

Today's gold standard in HIV therapy is the combined antiretroviral therapy (cART). It requires strict adherence by patients and life-long medication, which can lower the viral load below detection limits and prevent HIV-associated immunodeficiency, but cannot cure patients. The bispecific T cell engaging (BiTE®) antibody technology has demonstrated long-term relapse-free outcomes in patients with relapsed and refractory acute lymphocytic leukemia. We here generated BiTE® antibody constructs that target the HIV-1 envelope protein gp120 (HIV gp120) using either the scFv B12 or VRC01, the first two extracellular domains (1+2) of human CD4 alone or joined to the single chain variable fragment (scFv) of the antibody 17b fused to an anti-human CD3ϵ scFv. These engineered human BiTE® antibody constructs showed engagement of T cells for redirected lysis of HIV gp120-transfected CHO cells. Furthermore, they substantially inhibited HIV-1 replication in PBMCs as well as in macrophages co-cultured with autologous CD8+ T-cells, the most potent being the human CD4(1+2) BiTE® antibody construct and the CD4(1+2)L17b BiTE® antibody construct. The CD4(1+2) h BiTE® antibody construct promoted HIV infection of human CD4-/CD8+ T cells. In contrast, the neutralizing B12 and the VRC01 BiTE® antibody constructs as well as the CD4(1+2)L17b BiTE® antibody construct did not. Thus, BiTE® antibody constructs targeting HIV gp120 are very promising for constraining HIV and warrant further development as novel antiviral therapy with curative potential. Importance HIV is a chronic infection well controlled with the current cART. However, we lack cure of HIV, and the HIV pandemic goes on. Here we showed in vitro and ex vivo t hat a bispecific T-cell engaging (BiTE®) antibody construct targeting HIV gp120 resulted in substantially reduced HIV replication. In addition, these BiTE® antibody constructs display efficient killing of gp120 expressing cells and inhibited replication in ex vivo

Sup(99m) Technetium - labeled red blood cells 'in vitro'

International Nuclear Information System (INIS)

Bernardo Filho, M.; Souza Moura, I.N. de; Boasquevisque, E.M.

1983-01-01

A simple technique for the preparation of sup(99m) Tc labeled red blood cells using a comercial kit is described. To each 3ml of plain blood with anti-coagulant was added 1ml of solution of commercial kit with 6.8 μg of stannous chloride. This mixture was incubated in water bath, at 37 0 C, for 60 minutes. Then technetium-99m was added and the mixture was left for another ten minutes, in water bath, at 37 0 C. Under these conditions there was the best labeling of the red blood cells. Similar results were obtained with a solution of stannous chloride prepared freshly. The labeling is strong for 6.8 μg stannous chloride because the labeling was not removed by the several washes of the red blood cells or by the left in water bath. (Author) [pt

Investigation on Cell Proliferation with a New Antibody against Thymidine Kinase 1

Directory of Open Access Journals (Sweden)

Naining Wang

2001-01-01

Full Text Available The cytosolic thymidine kinase 1 (TK1 is one of the enzymes involved in DNA replication. Based on biochemical studies, TK1 is activated at late G1 of cell cycle, and its activity correlates with the cell proliferation. We have developed a polyclonal anti‐TK1 antibody against a synthetic peptide from the C‐terminus of human TK1. Using this antibody, here we demonstrate the exclusive location of TK1 in the cytoplasm of cells. Cell cycle dependent TK1 expression was studied by simultaneous fluorescence staining for TK1 and bromodeoxyuridine, by using elutriated cells, and by quantitation of the amount TK1 in relation to the cellular DNA content. TK1, which was strongly expressed in the cells in S+G2 period, raised at late G1 and decreased during mitosis. The amount of TK1 increased three folds from late G1 to G2. TK1 positive cells were demonstrated in areas of proliferation activity of various normal and malignant tissues. The new anti‐TK1 antibody works in archival specimens and is a specific marker of cell proliferation.

CEACAM1 induces B-cell survival and is essential for protective antiviral antibody production

Science.gov (United States)

Khairnar, Vishal; Duhan, Vikas; Maney, Sathish Kumar; Honke, Nadine; Shaabani, Namir; Pandyra, Aleksandra A.; Seifert, Marc; Pozdeev, Vitaly; Xu, Haifeng C.; Sharma, Piyush; Baldin, Fabian; Marquardsen, Florian; Merches, Katja; Lang, Elisabeth; Kirschning, Carsten; Westendorf, Astrid M.; Häussinger, Dieter; Lang, Florian; Dittmer, Ulf; Küppers, Ralf; Recher, Mike; Hardt, Cornelia; Scheffrahn, Inka; Beauchemin, Nicole; Göthert, Joachim R.; Singer, Bernhard B.; Lang, Philipp A.; Lang, Karl S.

2015-01-01

B cells are essential for antiviral immune defence because they produce neutralizing antibodies, present antigen and maintain the lymphoid architecture. Here we show that intrinsic signalling of CEACAM1 is essential for generating efficient B-cell responses. Although CEACAM1 exerts limited influence on the proliferation of B cells, expression of CEACAM1 induces survival of proliferating B cells via the BTK/Syk/NF-κB-axis. The absence of this signalling cascade in naive Ceacam1−/− mice limits the survival of B cells. During systemic infection with cytopathic vesicular stomatitis virus, Ceacam1−/− mice can barely induce neutralizing antibody responses and die early after infection. We find, therefore, that CEACAM1 is a crucial regulator of B-cell survival, influencing B-cell numbers and protective antiviral antibody responses. PMID:25692415

Thymus cells in myasthenia gravis selectively enhance production of anti-acetylcholine-receptor antibody by autologous blood lymphocytes

International Nuclear Information System (INIS)

Newsom-Davis, J.; Willcox, N.; Calder, L.

1981-01-01

We investigated the role of the thymus in 16 patients with myasthenia gravis without thymoma by studying the production of anti-acetylcholine-receptor antibody by thymic and blood lymphocytes cultured alone or together. In 10 responders (with the highest receptor-antibody titers in their plasma), cultured thymic cells spontaneously produced measurable receptor antibody. Receptor-antibody production by autologous blood lymphocytes was enhanced by the addition of responder's thymic cells, irradiated to abrogate antibody production and suppression (P<0.01). This enhancement was greater and more consistent than that by pokeweed mitogen; it depended on viable thymic cells, appeared to be selective for receptor antibody, and correlated with the ratio of thymic helper (OKT4-positive or OKT4+) to suppressor (OKT8+) T cells (P<0.01). These results suggest that myasthenic thymus contains cell-bound acetylcholine-receptor-like material or specific T cells (or both) that can aid receptor-antibody production. This may be relevant to the benefits of thymectomy in myasthenia and to the breakdown in self-tolerance in this and other autoimmune diseases

Thymus cells in myasthenia gravis selectively enhance production of anti-acetylcholine-receptor antibody by autologous blood lymphocytes

Energy Technology Data Exchange (ETDEWEB)

Newsom-Davis, J.; Willcox, N.; Calder, L.

1981-11-26

We investigated the role of the thymus in 16 patients with myasthenia gravis without thymoma by studying the production of anti-acetylcholine-receptor antibody by thymic and blood lymphocytes cultured alone or together. In 10 responders (with the highest receptor-antibody titers in their plasma), cultured thymic cells spontaneously produced measurable receptor antibody. Receptor-antibody production by autologous blood lymphocytes was enhanced by the addition of responder's thymic cells, irradiated to abrogate antibody production and suppression (P<0.01). This enhancement was greater and more consistent than that by pokeweed mitogen; it depended on viable thymic cells, appeared to be selective for receptor antibody, and correlated with the ratio of thymic helper (OKT4-positive or OKT4+) to suppressor (OKT8+) T cells (P<0.01). These results suggest that myasthenic thymus contains cell-bound acetylcholine-receptor-like material or specific T cells (or both) that can aid receptor-antibody production. This may be relevant to the benefits of thymectomy in myasthenia and to the breakdown in self-tolerance in this and other autoimmune diseases.

Red blood cell alloimmunization in sickle cell disease patients in ...

African Journals Online (AJOL)

Objective: Alloimmunization is a recognized complication of red blood cell (RBC) transfusion and causes delayed hemolytic transfusion reactions and provides problems sourcing compatible blood for future transfusions. The objective of this study was to determine the frequency of RBC alloimmunization in SCD patients in ...

MORPHOMETRIC CHARACTERISTICS OF RED BLOOD CELLS OF Telestes metohiensis (Steindachner, 1901

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Radoslav Dekić

2013-02-01

Full Text Available The paper presents the morphometric characteristics of red blood cells of endemic fish species of Bosnia and Herzegovina, Telestes metohiensis (Steindachner, 1901 inhabiting the Vrijeka river in the Dabar field. A total of 30 fish were sampled during August, 2010. Morphological measurements included the following parameters: axes of the red blood cells and nuclei, the surface of the red blood cells and nuclei and the thickness of the red blood cells. Morphometric characteristics of the erythrocyte maturation stages (acidophilic and polychromatic erythroblasts were also studied as well as their proportion in the peripheral blood. 100 mature forms were measured for each individual. The propotion of the immature forms was expressed per 1000 erythrocytes. Results showed that dimensions of the erythrocytes differed in systematic categories as well as fish types. Dimensions of mature erythrocytes and their maturation stages of the same species differed in shape and size of the nuclei. Proportion of the erythrocyte maturation stages was very low in comparison with the mature erythrocytes, indicating the optimal environmental conditions for the studied species.Key words: morphometric characteristics, erythrocytes, Telestes metohiensis, proportion of immature stages

A single-domain antibody-linked Fab bispecific antibody Her2-S-Fab has potent cytotoxicity against Her2-expressing tumor cells.

Science.gov (United States)

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

2016-12-01

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

Long-term culture and differentiation of porcine red bone marrow hematopoietic cells co-cultured with immortalized mesenchymal cells.

Science.gov (United States)

Garba, Abubakar; Acar, Delphine D; Roukaerts, Inge D M; Desmarets, Lowiese M B; Devriendt, Bert; Nauwynck, Hans J

2017-09-01

Mesenchymal cells are multipotent stromal cells with self-renewal, differentiation and immunomodulatory capabilities. We aimed to develop a co-culture model for differentiating hematopoietic cells on top of immortalized mesenchymal cells for studying interactions between hematopoietic and mesenchymal cells, useful for adequately exploring the therapeutic potential of mesenchymal cells. In this study, we investigated the survival, proliferation and differentiation of porcine red bone marrow hematopoietic cells co-cultured with immortalized porcine bone marrow mesenchymal cells for a period of five weeks. Directly after collection, primary porcine bone marrow mesenchymal cells adhered firmly to the bottom of the culture plates and showed a fibroblast-like appearance, one week after isolation. Upon immortalization, porcine bone marrow mesenchymal cells were continuously proliferating. They were positive for simian virus 40 (SV40) large T antigen and the mesenchymal cell markers CD44 and CD55. Isolated red bone marrow cells were added to these immortalized mesenchymal cells. Five weeks post-seeding, 92±6% of the red bone marrow hematopoietic cells were still alive and their number increased 3-fold during five weekly subpassages on top of the immortalized mesenchymal cells. The red bone marrow hematopoietic cells were originally small and round; later, the cells increased in size. Some of them became elongated, while others remained round. Tiny dendrites appeared attaching hematopoietic cells to the underlying immortalized mesenchymal cells. Furthermore, weekly differential-quick staining of the cells indicated the presence of monoblasts, monocytes, macrophages and lymphocytes in the co-cultures. At three weeks of co-culture, flow cytometry analysis showed an increased surface expression of CD172a, CD14, CD163, CD169, CD4 and CD8 up to 37±0.8%, 40±8%, 41±4%, 23±3% and 19±5% of the hematopoietic cells, respectively. In conclusion, continuous mesenchymal cell

Radioimmunotherapy of Non-Hodgkin's Lymphoma. The interaction of radiation and antibody with lymphoma cells

International Nuclear Information System (INIS)

Illidge, T.M.

1999-06-01

Whilst many patients with indolent Non-Hodgkin's Lymphoma (NHL) can achieve clinical remissions to first-line chemotherapy and/or radiotherapy, most will relapse. Current treatment options for relapsing patients are limited since most patients become resistant to repeated chemotherapy. Death usually occurs within 10 years of diagnosis. Overall, these disappointing results have not changed significantly in a quarter of a century and clearly advocate the urgent priority to research into potential new therapeutic approaches into this diverse and increasingly prevalent group of human tumours. Radioimmunotherapy (RIT) is currently under investigation as a new approach for the treatment of this disease. In this form of treatment, radionuclide-labeled monoclonal antibodies are able to deliver selective systemic irradiation by recognising tumour-associated antigens. The use of RIT with radiolabeled anti-CD20 antibodies in patients with recurrent B-cell lymphoma has resulted in extremely high rates of durable complete remissions. The optimal approach and mechanisms of action of successful RIT remain however largely unknown. The work described in this thesis has focused on clarifying some of the important determinants and mechanisms of effective RIT of syngeneic B-cell lymphoma, both in vivo and in vitro. A successful animal model of RIT in B cell lymphomas was established by initially generating a panel of antibodies against mouse B cell antigens. The in vitro characteristics of these antibodies have been compared with their subsequent performance, in biodistribution studies and RIT in vivo. For the first time in an in vivo model the relative contributions of antibody and irradiation are described. Some antibodies including anti-MHC Class II were shown to be effective delivery vehicles of low doses of Iodine-131. These antibodies, which appear to be inactive delivery vehicles can cure animals with low burdens of tumour. However antibodies such as anti-idiotype and anti-CD40

Preoperative factors associated with red blood cell transfusion in hip fracture patients

DEFF Research Database (Denmark)

Madsen, Christian Medom; Jørgensen, Henrik Løvendahl; Norgaard, Astrid

2014-01-01

Red blood cell (RBC) transfusion is a frequently used treatment in patients admitted with a fractured hip, but the use remains an area of much debate. The aim of this study was to determine preoperative factors associated with the risk of receiving a red blood cell transfusion in hip fracture...

21 CFR 864.8185 - Calibrator for red cell and white cell counting.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Calibrator for red cell and white cell counting. 864.8185 Section 864.8185 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Reagents § 864.8185...

Lentivirus display: stable expression of human antibodies on the surface of human cells and virus particles.

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Ran Taube

Full Text Available BACKGROUND: Isolation of human antibodies using current display technologies can be limited by constraints on protein expression, folding and post-translational modifications. Here we describe a discovery platform that utilizes self-inactivating (SIN lentiviral vectors for the surface display of high-affinity single-chain variable region (scFv antibody fragments on human cells and lentivirus particles. METHODOLOGY/PRINCIPAL FINDINGS: Bivalent scFvFc human antibodies were fused in frame with different transmembrane (TM anchoring moieties to allow efficient high-level expression on human cells and the optimal TM was identified. The addition of an eight amino acid HIV-1 gp41 envelope incorporation motif further increased scFvFc expression on human cells and incorporation into lentiviral particles. Both antibody-displaying human cells and virus particles bound antigen specifically. Sulfation of CDR tyrosine residues, a property recently shown to broaden antibody binding affinity and antigen recognition was also demonstrated. High level scFvFc expression and stable integration was achieved in human cells following transduction with IRES containing bicistronic SIN lentivectors encoding ZsGreen when scFvFc fusion proteins were expressed from the first cassette. Up to 10(6-fold enrichment of antibody expressing cells was achieved with one round of antigen coupled magnetic bead pre-selection followed by FACS sorting. Finally, the scFvFc displaying human cells could be used directly in functional biological screens with remarkable sensitivity. CONCLUSIONS/SIGNIFICANCE: This antibody display platform will complement existing technologies by virtue of providing properties unique to lentiviruses and antibody expression in human cells, which, in turn, may aid the discovery of novel therapeutic human mAbs.

The morphological classification of normal and abnormal red blood cell using Self Organizing Map

Science.gov (United States)

Rahmat, R. F.; Wulandari, F. S.; Faza, S.; Muchtar, M. A.; Siregar, I.

2018-02-01

Blood is an essential component of living creatures in the vascular space. For possible disease identification, it can be tested through a blood test, one of which can be seen from the form of red blood cells. The normal and abnormal morphology of the red blood cells of a patient is very helpful to doctors in detecting a disease. With the advancement of digital image processing technology can be used to identify normal and abnormal blood cells of a patient. This research used self-organizing map method to classify the normal and abnormal form of red blood cells in the digital image. The use of self-organizing map neural network method can be implemented to classify the normal and abnormal form of red blood cells in the input image with 93,78% accuracy testing.

F NMR measurement of intracellular free calcium in human red blood cells

International Nuclear Information System (INIS)

Gupta, R.K.; Schanne, F.A.X.

1986-01-01

Optical techniques for the measurement of intracellular Ca are not readily applicable to the human red cell because of the intense absorption of hemoglobin. The authors have therefore examined the use of 19 F NMR of 5,5'-difluoro-1,2-bis(o-aminophenoxy) ethane-N,N,N',N'-tetra acetic acid (5FBAPTA) introduced non-disruptively by intracellular hydrolysis of the membrane-permeant acetoxymethyl ester derivative. 19 F NMR spectra of 5FBAPTA-containing erythrocytes at 188 MHz displayed two well resolved resonances corresponding to the free and Ca-bound forms of the chelator, the resonance of the free form being ten-fold larger than that of the Ca-bound form. Addition of the ionophore A23187 resulted in the disappearance of the resonance of the free anion and a quantitative increase in the intensity of the resonance of the Ca-complex. From these data, and a K/sub D/ of 708 nM for the Ca-5FBAPTA complex, the authors estimate red cell free Ca to be 70 nM, which is in the range of values obtained for other cells, despite the fact that the human red cell, which lacks intracellular organelles for storing Ca, possesses only 1 μmol total Ca/1. cells in comparison to mmols of total Ca found in other cells. The authors ability to use 19 F NMR to measure free Ca in the red blood cell paves the way for future NMR studies of red cell free Ca concentrations in human essential hypertension as well as in other diseases states in which alterations in cellular Ca homeostasis may be involved

Antibody-dependent cellular cytotoxicity and cytokine/chemokine secretion by KHYG-1 cells stably expressing FcγRIIIA.

Science.gov (United States)

Kobayashi, Eiji; Motoi, Sotaro; Sugiura, Masahito; Kajikawa, Masunori; Kojima, Shuji; Kohroki, Junya; Masuho, Yasuhiko

2014-09-01

Antibody-dependent cellular cytotoxicity (ADCC) mediated by natural killer (NK) cells is a major mechanism of tumor therapy with antibodies. NK cells not only manifest cytotoxicity but also secrete a variety of cytokines/chemokines that regulate immune responses. Using a retroviral vector, in this study we established a KHYG-1 cell line that stably expresses FcγRIIIA (CD16A). The KHYG-1/FcγRIIIA cells exerted potent antibody concentration-dependent ADCC, whereas parental KHYG-1 cells did not. In contrast, without antibody, the natural killer activity of KHYG-1/FcγRIIIA cells was less potent than that of parental KHYG-1 cells. During the course of ADCC, KHYG-1/FcγRIIIA cells secreted IFN-γ and MIP-1α dependent upon antibody concentration, but parental KHYG-1 cells did not. These results suggest that KHYG-1/FcγRIIIA cells would be useful in studies to elucidate the function of NK cells and the mechanism of ADCC. Copyright © 2014 Elsevier B.V. All rights reserved.

Nature of the elements transporting long-chain fatty acids through the red cell membrane

DEFF Research Database (Denmark)

Bojesen, Inge Norby; Bojesen, Eigil

1998-01-01

Docosahexaenoic acid, linoleic acid, red cell membrane, transporting elements, transport kinetics, fatty acid transport......Docosahexaenoic acid, linoleic acid, red cell membrane, transporting elements, transport kinetics, fatty acid transport...

Solid phase radioimmunoassay for detection of malaria antigen. Comparison of monoclonal and polyclonal antibodies

International Nuclear Information System (INIS)

Khusmith, S.; Tharavanij, S.; Patarapotikul, J.; Kasemsuth, R.; Bunnag, D.

1986-01-01

A solid phase competitive binding radioimmunoassay (RIA) was developed for the detection of Plasmodium falciparum in infected blood. A suspension of NP40 treated red blood cells was mixed with labelled antimalarial IgG, incubated and then added to malarial antigen coated microtitre plate. Antimalarial IgGs were purified either from high titre sera from individuals living in a malaria endemic area in Thailand or from a locally produced monoclonal antibody (MAB) which showed a bright generalized immunofluorescent staining pattern against all blood stages of P. falciparum, including gametocytes. This MAB reacted with 27 of 31 P. falciparum isolates from Thailand. Using dilution of red blood cells from in vitro cultures of P. falciparum, the test was found to detect parasites at levels equivalent to 13 and 2.2 parasites/10 6 red blood cells with labelled polyclonal IgG (PIgG) and labelled monoclonal IgG (MIgG), respectively. No false positive results were obtained among samples from non-malarial subjects. Of the samples that gave negative results upon microscopic examination, 50 and 35% were still positive with RIA using MIgG and PIgG, respectively. There was a correlation between RIA and the number of parasites, especially when MIgG was used. The results indicate that the IgG fraction of sera from individuals with natural acquired immunity to malaria showed a lower degree of sensitivity in parasite detection than the IgG from monoclonal antibody. (author)

Hemolytic disease of fetus and newborn due to maternal red blood cell alloantibodies in the Malay population

Science.gov (United States)

Hassan, Mohd Nazri; Mohd Noor, Noor Haslina; Johan Noor, Shah Reza; Sukri, Salamah Ahmad; Mustafa, Rapiaah; Luc Aster, Hans Van Rostenberghe

2014-01-01

Background: Maternal red blood cell (RBC) alloimmunization may lead to production of harmful antibodies that result in hemolytic disease of fetus and newborn (HDFN). There is insufficient data on the prevalence of HDFN due to RBC alloantibodies in the Malay neonatal population. Aim: The aim of this study was to determine the incidence of HDFN in the Malay neonatal population due to clinically significant RBC alloantibodies. Subjects and Methods: A cross sectional study was conducted in Transfusion Medicine Unit, Hospital Universitiy Sains Malaysia over one year period from January to December 2009. A total of 5163 Malay pregnant women who attended labor room for delivery were collected and analyzed prospectively. The blood samples were subjected to the standard immunohematological procedure for RBC antibody screening and identification using reagents of Diamed-ID Gel microtyping system. All the newborns with RBC alloantibody were investigated for the evidence of HDFN. Results: Thirty (0.58%) women were found to have clinically significant RBC alloantibodies. Most of the alloantibodies belonged to Rhesus (Rh) system (56.7%) where anti-E (33.3%) was the most common followed by anti-D (10.0%). Rh antibodies were the main cause of HDFN in fourteen (0.27%) neonates. Anti-D and anti-c were identified to cause moderate to very severe HDFN. Conclusions: With the low prevalence of clinically significant RBC alloantibodies and HDFN, routine antenatal antibody screening practice may not be advised as a routine practice at present, preferably reserved for those women of RhD negative or with history of HDFN, significantly of those attributed to anti-c. PMID:25161351

Naturally occurring anti-glycan antibodies binding to Globo H-expressing cells identify ovarian cancer patients.

Science.gov (United States)

Pochechueva, Tatiana; Alam, Shahidul; Schötzau, Andreas; Chinarev, Alexander; Bovin, Nicolai V; Hacker, Neville F; Jacob, Francis; Heinzelmann-Schwarz, Viola

2017-02-10

Glycosphingolipids are important compounds of the plasma membrane of mammalian cells and a number of them have been associated with malignant transformation and progression, reinforcing tumour aggressiveness and metastasis. Here we investigated the levels of naturally occurring anti-glycan antibodies to Globo H in blood plasma obtained from high-grade serous ovarian cancer patients (SOC) and women without gynaecological malignancies (control) using suspension glycan array technology employing chemically synthesized glycans as antibody targets. We found that anti-human Globo H IgG antibodies were able to significantly discriminate SOC from controls (P anti-Globo H antibodies highly correlated (r = 0.992). The incubation of plasma-derived anti-glycan antibodies with chemically synthesized (presented on fluorescence microspheres) and native Globo H (expressed on Globo H-positive cell lines) revealed strong reactivity of naturally occurring human anti-Globo H antibodies towards its antigen expressed on ovarian cancer cells. Our data demonstrate that human plasma-derived antibodies to Globo H as well as the presence of the antigen might be considered as therapeutic option in ovarian cancer.

Photocleavable DNA Barcoding Antibodies for Multiplexed Protein Analysis in Single Cells.

Science.gov (United States)

Ullal, Adeeti V; Weissleder, Ralph

2015-01-01

We describe a DNA-barcoded antibody sensing technique for single cell protein analysis in which the barcodes are photocleaved and digitally detected without amplification steps (Ullal et al., Sci Transl Med 6:219, 2014). After photocleaving the unique ~70 mer DNA barcodes we use a fluorescent hybridization technology for detection, similar to what is commonly done for nucleic acid readouts. This protocol offers a simple method for multiplexed protein detection using 100+ antibodies and can be performed on clinical samples as well as single cells.

Increased phorbol 12,13-dibutyrate (PDBu) receptor function associated with sickle red cell membrane ghosts

International Nuclear Information System (INIS)

Ramachandran, M.; Nair, C.N.; Abraham, E.C.

1987-01-01

The biological receptor for tumor-promoting phorbol esters has been identified as the Ca 2+ /phospholipid dependent enzyme, protein kinase C. In the red cell, this enzyme is mainly cytosolic but becomes translocated to the membrane if the cellular Ca 2+ is allowed to rise. Since cellular Ca 2+ in sickle red cells is high, it was reasoned that this enzyme may become more membrane-bound. In fact, the authors noticed a four-fold increase in the binding of 3 H-PDBu by membrane ghosts isolated from sickle red cells compared to normal red cells (pmoles PDBu bound/mg protein; normal = 0.3 vs sickle cell = 1.4). Attempts to assay the enzyme directly as phospholipid-activated 32 P incorporation into the acid-precipitable membrane proteins also indicated a two-fold increase in the radiolabelling of sickle cell membrane ghosts. Autophosphorylation of membrane proteins and analysis of the phosphorylation profile by SDS-PAGE and autoradiography revealed phosphorylation predominantly of bands 3, 4.1 and 4.9 which are known protein kinase C substrates for the red cell enzyme. The increased membrane-associated protein kinase C in sickle red cells may have a bearing on the altered membrane properties reported in this condition

Breaking the color barrier - a multi-selective antibody reporter offers innovative strategies of fluorescence detection.

Science.gov (United States)

Gallo, Eugenio; Jarvik, Jonathan W

2017-08-01

A novel bi-partite fluorescence platform exploits the high affinity and selectivity of antibody scaffolds to capture and activate small-molecule fluorogens. In this report, we investigated the property of multi-selectivity activation by a single antibody against diverse cyanine family fluorogens. Our fluorescence screen identified three cell-impermeant fluorogens, each with unique emission spectra (blue, green and red) and nanomolar affinities. Most importantly, as a protein fusion tag to G-protein-coupled receptors, the antibody biosensor retained full activity - displaying bright fluorogen signals with minimal background on live cells. Because fluorogen-activating antibodies interact with their target ligands via non-covalent interactions, we were able to perform advanced multi-color detection strategies on live cells, previously difficult or impossible with conventional reporters. We found that by fine-tuning the concentrations of the different color fluorogen molecules in solution, a user may interchange the fluorescence signal (onset versus offset), execute real-time signal exchange via fluorogen competition, measure multi-channel fluorescence via co-labeling, and assess real-time cell surface receptor traffic via pulse-chase experiments. Thus, here we inform of an innovative reporter technology based on tri-color signal that allows user-defined fluorescence tuning in live-cell applications. © 2017. Published by The Company of Biologists Ltd.

Ocaratuzumab, an Fc-engineered antibody demonstrates enhanced antibody-dependent cell-mediated cytotoxicity in chronic lymphocytic leukemia.

Science.gov (United States)

Cheney, Carolyn M; Stephens, Deborah M; Mo, Xiaokui; Rafiq, Sarwish; Butchar, Jonathan; Flynn, Joseph M; Jones, Jeffrey A; Maddocks, Kami; O'Reilly, Adrienne; Ramachandran, Abhijit; Tridandapani, Susheela; Muthusamy, Natarajan; Byrd, John C

2014-01-01

Chronic lymphocytic leukemia (CLL) is common in both developed and developing nations where the need for inexpensive and convenient administration of therapy is apparent. Ocaratuzumab is a novel Fc-engineered humanized IgG1 anti-CD20 monoclonal antibody (mAb) designed for effective antibody-dependent cell-mediated cytotoxicity (ADCC) at very low concentrations that may facilitate sub-cutaneous (vs. intravenous) dosing. Here, we report ocaratuzumab's potency against CLL cells. In vitro assessment of ocaratuzumab's direct cytotoxicity (DC), complement-dependent cytotoxicity (CDC), antibody-dependent cellular phagocytosis (ADCP), and ADCC was performed on CLL cells. Ocaratuzumab induced DC, CDC, and ADCP similarly to rituximab or ofatumumab (anti-CD20 mAbs). However, ocaratuzumab showed an advantage in NK cell-mediated ADCC over these antibodies. In allogeneic ADCC, [E:T (effector:target) ratios = 25:1, 12:1, 6:1], ocaratuzumab (10 µg/mL) improved ADCC by ~3-fold compared with rituximab or ofatumumab (P<0.001 all tested E:T ratios). Notably, the superiority of ocaratuzumab-induced ADCC was observed at low concentrations (0.1-10 ug/ml; P<0.03; allogeneic assays). In extended allogeneic ADCC E:T titration, ocaratuzumab (0.1 µg/mL) demonstrated 19.4% more cytotoxicity than rituximab (E:T = 0.38:1; P = 0.0066) and 21.5% more cytotoxicity than ofatumumab (E:T = 1.5:1; P = 0.0015). In autologous ADCC, ocaratuzumab (10 µg/mL) demonstrated ~1.5-fold increase in cytotoxicity compared with rituximab or ofatumumab at all E:T ratios tested (E:Ts = 25:1,12:1,6:1; all P<0.001). Obinutuzumab, a glyco-engineered anti-CD20 mAb, showed no improvement in ADCC activity compared with ocaratuzumab. The enhanced ADCC of ocaratuzumab suggests that it may be effective at low concentrations. If supported by clinical investigation, this feature could potentially allow for subcutaneous dosing at low doses that could expand the potential of administering chemoimmunotherapy in developing

Antibody-dependent cellular cytotoxicity (ADCC) activity of a novel anti-PD-L1 antibody avelumab (MSB0010718C) on human tumor cells

OpenAIRE

Boyerinas, Benjamin; Jochems, Caroline; Fantini, Massimo; Heery, Christopher R.; Gulley, James L.; Tsang, Kwong Yok; Schlom, Jeffrey

2015-01-01

Several anti-PD1/PD-L1 monoclonal antibodies (MAb) are currently providing evidence of clinical benefit in subsets of cancer patients. The mode of action of these MAbs is to inhibit PD1 on immune cells interacting with PD-L1 on tumor cells. These MAbs are either designed or engineered to eliminate antibody-dependent cell-mediated cytotoxicity (ADCC), which, however, has been implicated as an important mechanism in several highly effective MAb-mediated cancer therapies. A fully human anti-PD-L...

Simple and efficient generation of virus-specific T cells for adoptive therapy using anti-4-1BB antibody.

Science.gov (United States)

Imahashi, Nobuhiko; Nishida, Tetsuya; Goto, Tatsunori; Terakura, Seitaro; Watanabe, Keisuke; Hanajiri, Ryo; Sakemura, Reona; Imai, Misa; Kiyoi, Hitoshi; Naoe, Tomoki; Murata, Makoto

2015-01-01

Although recent studies of virus-specific T-cell (VST) therapy for viral infections after allogeneic hematopoietic stem cell transplantation have shown promising results, simple and less time-intensive and labor-intensive methods are required to generate VSTs for the wider application of VST therapy. We investigated the efficacy of anti-CD28 and anti-4-1BB antibodies, which can provide T cells with costimulatory signals similar in strength to those of antigen-presenting cells, in generating VSTs. When peripheral blood mononuclear cells were stimulated with viral peptides together with isotype control, anti-CD28, or anti-4-1BB antibodies, anti-4-1BB antibodies yielded the highest numbers of VSTs, which were on an average 7.9 times higher than those generated with isotype control antibody. The combination of anti-CD28 and anti-4-1BB antibodies did not result in increased numbers of VSTs compared with anti-4-1BB antibody alone. Importantly, the positive effect of anti-4-1BB antibody was observed regardless of the epitopes of the VSTs. In contrast, the capacity of dendritic cells (DCs) to generate VSTs differed considerably depending on the epitopes of the VSTs. Furthermore, the numbers of VSTs generated with DCs were at most similar to those generated with the anti-4-1BB antibody. Generation of VSTs with anti-4-1BB antibody did not result in excessive differentiation or deteriorated function of the generated VSTs compared with those generated with control antibody or DCs. In conclusion, VSTs can be generated rapidly and efficiently by simply stimulating peripheral blood mononuclear cells with viral peptide and anti-4-1BB antibody without using antigen-presenting cells. We propose using anti-4-1BB antibody as a novel strategy to generate VSTs for adoptive therapy.

Maternal red blood cell alloantibodies identified in blood samples obtained from Iranian pregnant women: the first population study in Iran.

Science.gov (United States)

Shahverdi, Ehsan; Moghaddam, Mostafa; Gorzin, Fateme

2017-01-01

The objective was to determine the frequency of occurrence of alloantibodies among pregnant women in Iran. This was a prospective cross-sectional study, which was carried out in the immunohematology reference laboratory of the Iranian Blood Transfusion Organization in Tehran, Iran, in 2008 to 2015. Screening and identification of red blood cell (RBC) alloantibodies was done on the sera of 7340 pregnant females using the standard tube method and gel column agglutination technique. Alloantibodies were identified in the serum of 332 of the 7340 (4.5%) pregnant women. A total of 410 antibodies were detected in 332 positive maternal serum samples with no previous history of blood transfusion. Anti-D was the most common antibody accounting for 70.5% of all the antibodies formed in D- women. The incidence of specific alloimmunization other than Rh group was 14.4%. We concluded that the alloimmunization rate was high in comparison with wide pattern in previous studies. In Iran, like other developing countries, alloimmunization screening tests are performed only to detect anti-D in pregnant D- women. This high rate of alloimmunization, quite possibly, is due to the fact that the majority of blood samples came from pregnant women known to have previous obstetric problems. However, we suggest that RBC antibody screening tests should be extended to all D+ women. © 2016 AABB.

Shape Memory of Human Red Blood Cells

OpenAIRE

Fischer, Thomas M.

2004-01-01

The human red cell can be deformed by external forces but returns to the biconcave resting shape after removal of the forces. If after such shape excursions the rim is always formed by the same part of the membrane, the cell is said to have a memory of its biconcave shape. If the rim can form anywhere on the membrane, the cell would have no shape memory. The shape memory was probed by an experiment called go-and-stop. Locations on the membrane were marked by spontaneously adhering latex spher...

Human Cytomegalovirus Infection Increases Both Antibody- and Non–Antibody-Dependent Cellular Reactivity by Natural Killer Cells

Directory of Open Access Journals (Sweden)

Clive M. Michelo, PhD

2017-12-01

Conclusions. With regard to organ transplantation, these data suggest that CMV infection enhances NK cell alloreactivity, which may pose an additional adverse effect on graft survival, especially in the presence of donor specific antibodies.

Comparative study on the effect of radiation on whole blood and isolate red blood cells

International Nuclear Information System (INIS)

Selim, N.S.

2009-01-01

Assessment of the dielectric properties of red blood cells requires several steps for preparation and isolation from whole blood. These steps may results in changes in the cells properties, and they are time consuming . The present study aims to compare the properties of both whole blood and isolated red blood cells and the effect of gamma radiation on these properties. Adult male rats were exposed to 1, 3.5 and 7 Gy as single dose, from Cs-137 source.The studies dielectric properties, in the frequency range 40 k Hz to 5 MHz, and light scattering studies for suspensions of whole blood and isolated red blood cells from the same groups were measured. The obtained results showed that whole blood and red blood cells suspensions followed the same trend in their response to radiation, which suggests the possibility of using whole blood suspension for the evaluation of the red blood cells properties

Targeting of phage particles towards endothelial cells by antibodies selected through a multi-parameter selection strategy.

Science.gov (United States)

Mandrup, Ole A; Lykkemark, Simon; Kristensen, Peter

2017-02-10

One of the hallmarks of cancer is sustained angiogenesis. Here, normal endothelial cells are activated, and their formation of new blood vessels leads to continued tumour growth. An improved patient condition is often observed when angiogenesis is prevented or normalized through targeting of these genomically stable endothelial cells. However, intracellular targets constitute a challenge in therapy, as the agents modulating these targets have to be delivered and internalized specifically to the endothelial cells. Selection of antibodies binding specifically to certain cell types is well established. It is nonetheless a challenge to ensure that the binding of antibodies to the target cell will mediate internalization. Previously selection of such antibodies has been performed targeting cancer cell lines; most often using either monovalent display or polyvalent display. In this article, we describe selections that isolate internalizing antibodies by sequential combining monovalent and polyvalent display using two types of helper phages, one which increases display valence and one which reduces background. One of the selected antibodies was found to mediate internalization into human endothelial cells, although our results confirms that the single stranded nature of the DNA packaged into phage particles may limit applications aimed at targeting nucleic acids in mammalian cells.

Partitioning of red blood cell aggregates in bifurcating microscale flows

Science.gov (United States)

Kaliviotis, E.; Sherwood, J. M.; Balabani, S.

2017-03-01

Microvascular flows are often considered to be free of red blood cell aggregates, however, recent studies have demonstrated that aggregates are present throughout the microvasculature, affecting cell distribution and blood perfusion. This work reports on the spatial distribution of red blood cell aggregates in a T-shaped bifurcation on the scale of a large microvessel. Non-aggregating and aggregating human red blood cell suspensions were studied for a range of flow splits in the daughter branches of the bifurcation. Aggregate sizes were determined using image processing. The mean aggregate size was marginally increased in the daughter branches for a range of flow rates, mainly due to the lower shear conditions and the close cell and aggregate proximity therein. A counterintuitive decrease in the mean aggregate size was apparent in the lower flow rate branches. This was attributed to the existence of regions depleted by aggregates of certain sizes in the parent branch, and to the change in the exact flow split location in the T-junction with flow ratio. The findings of the present investigation may have significant implications for microvascular flows and may help explain why the effects of physiological RBC aggregation are not deleterious in terms of in vivo vascular resistance.

Benchmarking of commercially available CHO cell culture media for antibody production.

Science.gov (United States)

Reinhart, David; Damjanovic, Lukas; Kaisermayer, Christian; Kunert, Renate

2015-06-01

In this study, eight commercially available, chemically defined Chinese hamster ovary (CHO) cell culture media from different vendors were evaluated in batch culture using an IgG-producing CHO DG44 cell line as a model. Medium adaptation revealed that the occurrence of even small aggregates might be a good indicator of cell growth performance in subsequent high cell density cultures. Batch experiments confirmed that the culture medium has a significant impact on bioprocess performance, but high amino acid concentrations alone were not sufficient to ensure superior cell growth and high antibody production. However, some key amino acids that were limiting in most media could be identified. Unbalanced glucose and amino acids led to high cell-specific lactate and ammonium production rates. In some media, persistently high glucose concentrations probably induced the suppression of respiration and oxidative phosphorylation, known as Crabtree effect, which resulted in high cell-specific glycolysis rates along with a continuous and high lactate production. In additional experiments, two of the eight basal media were supplemented with feeds from two different manufacturers in six combinations, in order to understand the combined impact of media and feeds on cell metabolism in a CHO fed-batch process. Cell growth, nutrient consumption and metabolite production rates, antibody production, and IgG quality were evaluated in detail. Concentrated feed supplements boosted cell concentrations almost threefold and antibody titers up to sevenfold. Depending on the fed-batch strategy, fourfold higher peak cell concentrations and eightfold increased IgG titers (up to 5.8 g/L) were achieved. The glycolytic flux was remarkably similar among the fed-batches; however, substantially different specific lactate production rates were observed in the different media and feed combinations. Further analysis revealed that in addition to the feed additives, the basal medium can make a considerable

Simultaneous development of antibody-dependent cellular cytotoxicity (ADCC) and natural killer (NK) activity in irradiated mice reconstituted with bone marrow cells

International Nuclear Information System (INIS)

Sihvola, M.; Hurme, M.

1987-01-01

Spleen cells from irradiated, bone marrow-reconstituted mice were tested for their ability to mediate antibody-dependent cellular cytotoxicity against P815 target (ADCC-P815), ADCC against sheep red blood cells (ADCC-SRBC), and natural killer (NK) activity judged as YAC-1 lysis at different times after bone marrow reconstitution. Donor-derived ADCC-P815 effectors were found to appear in the spleens 10-12 days after bone marrow reconstitution simultaneously with the appearance of donor-derived NK cells. NK cells recently derived from bone marrow are known to express the Thy-1 antigen; the phenotype of the ''early'' ADCC-P815 effectors was found to be the same as that of NK cells, i.e., Thy-1+, asialo-GM1+. These data suggest that ADCC-P815 effector cells belong to the NK cell population. ADCC-SRBC, in contrast to ADCC-P815 and NK activity, was already high on Day 7 after bone marrow reconstitution. However, it was mediated partly by recipient-derived effectors. ADCC-SRBC effectors were characterized to be different from ADCC-P815 effectors

Ex-vivo expansion of red blood cells: how real for transfusion in humans?

Science.gov (United States)

Migliaccio, Anna Rita; Masselli, Elena; Varricchio, Lilian; Whitsett, Carolyn

2012-03-01

Blood transfusion is indispensable for modern medicine. In developed countries, the blood supply is adequate and safe but blood for alloimmunized patients is often unavailable. Concerns are increasing that donations may become inadequate in the future as the population ages prompting a search for alternative transfusion products. Improvements in culture conditions and proof-of-principle studies in animal models have suggested that ex-vivo expanded red cells may represent such a product. Compared to other cell therapies transfusion poses the unique challenge of requiring great cell doses (2.5×10(12) cells vs 10(7) cells). Although production of such cell numbers is theoretically possible, current technologies generate red cells in numbers sufficient only for safety studies. It is conceived that by the time these studies will be completed, technical barriers to mass cell production will have been eliminated making transfusion with ex-vivo generated red cells a reality. Copyright © 2011 Elsevier Ltd. All rights reserved.

CARbodies: Human Antibodies Against Cell Surface Tumor Antigens Selected From Repertoires Displayed on T Cell Chimeric Antigen Receptors

Directory of Open Access Journals (Sweden)

Vanesa Alonso-Camino

2013-01-01

Full Text Available A human single-chain variable fragment (scFv antibody library was expressed on the surface of human T cells after transduction with lentiviral vectors (LVs. The repertoire was fused to a first-generation T cell receptor ζ (TCRζ-based chimeric antigen receptor (CAR. We used this library to isolate antibodies termed CARbodies that recognize antigens expressed on the tumor cell surface in a proof-of-principle system. After three rounds of activation-selection there was a clear repertoire restriction, with the emergence dominant clones. The CARbodies were purified from bacterial cultures as soluble and active proteins. Furthermore, to validate its potential application for adoptive cell therapy, human T cells were transduced with a LV encoding a second-generation costimulatory CAR (CARv2 bearing the selected CARbodies. Transduced human primary T cells expressed significant levels of the CARbodies-based CARv2 fusion protein on the cell surface, and importantly could be specifically activated, after stimulation with tumor cells. This approach is a promising tool for the generation of antibodies fully adapted to the display format (CAR and the selection context (cell synapse, which could extend the scope of current adoptive cell therapy strategies with CAR-redirected T cells.

Hemolytic disease of the fetus and newborn caused by an antibody to a low-prevalence antigen, anti-SARA.

Science.gov (United States)

Towns, Dale; Hannon, Judith; Hendry, Julia; Barnes, Janet; Goldman, Mindy

2011-09-01

The first case describing the SARAH (SARA) antigen occurred in 1990, in an Australian blood donor. Hemolytic disease of the fetus and newborn (HDFN) due to anti-SARA has not been previously described. We report a case of HDFN in a multiparous female. The pregnancy was unremarkable except that she was involved in a seemingly minor motor vehicle accident at 25 weeks' gestation. Routine prenatal antibody screening was negative throughout the pregnancy. She presented at 37 weeks' gestation because of decreased fetal movements. Labor was induced and a 2702-g infant male was delivered. The infant's hemoglobin was 49 g/L and the bilirubin was 153 µmol/L. Blood samples from the parents and infant were referred to Canadian Blood Services National Immunohematology Reference Laboratory and subsequently to the Australian Red Cross Red Cell Reference Service. The father's and infant's red blood cells were confirmed to be SARA positive, and the mother's plasma contained anti-SARA. The infant was successfully treated with a double-volume exchange transfusion. This is the first example of HDFN associated with this antibody. © 2011 American Association of Blood Banks.

Splenic TFH expansion participates in B-cell differentiation and antiplatelet-antibody production during immune thrombocytopenia.

Science.gov (United States)

Audia, Sylvain; Rossato, Marzia; Santegoets, Kim; Spijkers, Sanne; Wichers, Catharina; Bekker, Cornelis; Bloem, Andries; Boon, Louis; Flinsenberg, Thijs; Compeer, Ewoud; van den Broek, Theo; Facy, Olivier; Ortega-Deballon, Pablo; Berthier, Sabine; Leguy-Seguin, Vanessa; Martin, Laurent; Ciudad, Marion; Samson, Maxime; Trad, Malika; Lorcerie, Bernard; Janikashvili, Nona; Saas, Philippe; Bonnotte, Bernard; Radstake, Timothy R D J

2014-10-30

Antiplatelet-antibody-producing B cells play a key role in immune thrombocytopenia (ITP) pathogenesis; however, little is known about T-cell dysregulations that support B-cell differentiation. During the past decade, T follicular helper cells (TFHs) have been characterized as the main T-cell subset within secondary lymphoid organs that promotes B-cell differentiation leading to antibody class-switch recombination and secretion. Herein, we characterized TFHs within the spleen of 8 controls and 13 ITP patients. We show that human splenic TFHs are the main producers of interleukin (IL)-21, express CD40 ligand (CD154), and are located within the germinal center of secondary follicles. Compared with controls, splenic TFH frequency is higher in ITP patients and correlates with germinal center and plasma cell percentages that are also increased. In vitro, IL-21 stimulation combined with an anti-CD40 agonist antibody led to the differentiation of splenic B cells into plasma cells and to the secretion of antiplatelet antibodies in ITP patients. Overall, these results point out the involvement of TFH in ITP pathophysiology and the potential interest of IL-21 and CD40 as therapeutic targets in ITP. © 2014 by The American Society of Hematology.

Determination of stimulation effective dose of gamma ray on guinea pig to be used for antibody production

International Nuclear Information System (INIS)

Arifin, Muchson; Soewarsono, M.

1983-01-01

The experiment has been performed on guinea pigs which were irradiated by gamma rays with doses of 0, 150, 300 and 600 rad. Observations were carried out on peripheral blood in regard to red blood cell, white blood cell and haemoglobine, every week after irradiation. +he result obtained showed that 150 rad was the optimal stimulation effective dose for antibody production in guinea pigs. (author)

Determination of stimulation effective dose of gamma ray on guinea pig to be used for antibody production

Energy Technology Data Exchange (ETDEWEB)

Arifin, M; Soewarsono, M [National Atomic Energy Agency, Jakarta (Indonesia). Pasar Djumat Research Centre

1983-04-01

The experiment has been performed on guinea pigs which were irradiated by gamma rays with doses of 0, 150, 300 and 600 rad. Observations were carried out on peripheral blood in regard to red blood cell, white blood cell and haemoglobine, every week after irradiation. The result obtained showed that 150 rad was the optimal stimulation effective dose for antibody production in guinea pigs. 10 refs.

Image-based model of the spectrin cytoskeleton for red blood cell simulation.

Science.gov (United States)

Fai, Thomas G; Leo-Macias, Alejandra; Stokes, David L; Peskin, Charles S

2017-10-01

We simulate deformable red blood cells in the microcirculation using the immersed boundary method with a cytoskeletal model that incorporates structural details revealed by tomographic images. The elasticity of red blood cells is known to be supplied by both their lipid bilayer membranes, which resist bending and local changes in area, and their cytoskeletons, which resist in-plane shear. The cytoskeleton consists of spectrin tetramers that are tethered to the lipid bilayer by ankyrin and by actin-based junctional complexes. We model the cytoskeleton as a random geometric graph, with nodes corresponding to junctional complexes and with edges corresponding to spectrin tetramers such that the edge lengths are given by the end-to-end distances between nodes. The statistical properties of this graph are based on distributions gathered from three-dimensional tomographic images of the cytoskeleton by a segmentation algorithm. We show that the elastic response of our model cytoskeleton, in which the spectrin polymers are treated as entropic springs, is in good agreement with the experimentally measured shear modulus. By simulating red blood cells in flow with the immersed boundary method, we compare this discrete cytoskeletal model to an existing continuum model and predict the extent to which dynamic spectrin network connectivity can protect against failure in the case of a red cell subjected to an applied strain. The methods presented here could form the basis of disease- and patient-specific computational studies of hereditary diseases affecting the red cell cytoskeleton.

Mathematical analysis of 51Cr-labelled red cell survival curves in congenital haemolytic anaemias

International Nuclear Information System (INIS)

Kasfiki, A.G.; Antipas, S.E.; Dimitriou, P.A.; Gritzali, F.A.; Melissinos, K.G.

1982-01-01

The parameters of 51 Cr labelled red cell survival curves were calculated in 26 patients with homozygous β-thalassaemia, 8 with sickle-cell anaemia and 3 with s-β-thalassaemia, using a non-linear weighted least squares analysis computer program. In thalassaemic children the calculated parameters denote that the shorting of the mean cell life is due to early senescence alone, while there is some evidence that in thalassaemic adults additional extracellular destruction mechanisms participate as well. Red cell survival curves from patients with sickle-cell anaemia and s-β-thalassaemia resemble each other, while their parameters indicate an initial rapid loss of radioactivity, early senescence and the presence of extracellular red cell destruction factors. (orig.)

Cation Homeostasis in Red Cells From Patients With Sickle Cell Disease Heterologous for HbS and HbC (HbSC Genotype

Directory of Open Access Journals (Sweden)

A. Hannemann

2015-11-01

Full Text Available Sickle cell disease (SCD in patients of HbSC genotype is considered similar, albeit milder, to that in homozygous HbSS individuals — but with little justification. In SCD, elevated red cell cation permeability is critical as increased solute loss causes dehydration and encourages sickling. Recently, we showed that the KCl cotransporter (KCC activity in red cells from HbSC patients correlated significantly with disease severity, but that in HbSS patients did not. Two transporters involved in red cell dehydration, the conductive channels Psickle and the Gardos channel, behaved similarly in red cells from the two genotypes, but were significantly less active in HbSC patients. By contrast, KCC activity was quantitatively greater in HbSC red cells. Results suggest that KCC is likely to have greater involvement in red cell dehydration in HbSC patients, which could explain its association with disease severity in this genotype. This work supports the hypothesis that SCD in HbSC patients is a distinct disease entity to that in HbSS patients. Results suggest the possibility of designing specific treatments of particular benefit to HbSC patients and a rationale for the development of prognostic markers, to inform early treatment of children likely to develop more severe complications of the disease.

Liquid Storage at 4 deg C of Previously Frozen Red Cells

Science.gov (United States)

1987-12-01

adenosine tnphosphate (ATP). 2.3- acceptable red cell function. A post-thaw storage ca- diphosphoglycerate (2.3-DPG), glucose, supernatant hemo...and Received for publication September 22. 1986; revision received supernatant hemoglobin levels within the acceptable range, November 29, 1986, and...percent. All units were sterile at the end of the 21-day post- thaw storage period. 6.9 The mean red cell ATP and 2,3-DPG levels are shown in Figure 1

[Adult T-cell leukemia/lymphoma associated with unusual positivity of anti-ATLA (adult T-cell leukemia-cell-associated antigen) antibodies].

Science.gov (United States)

Eto, T; Okamura, H; Okamura, T; Gondo, H; Kudo, J; Shibuya, T; Harada, M; Niho, Y

1990-03-01

A 56-year-old female was admitted because of generalized lymphadenopathy. Based upon histological findings of biopsied lymph node, malignant lymphoma, diffuse large cell type was diagnosed. The surface marker analysis showed that malignant cells were positive for CD4 and CD2 but negative for CD8. Although anti-ATLA (adult T-cell leukemia associated antigen) antibody was negative with the use of a gelatin particle agglutination method (P.A.), other methods such as an indirect immunofluorescence assay (I.F.), an enzyme-linked immunosorbent assay (E.I.A.) and a Western blotting assay revealed the positivity for anti-ATLA antibody. Adult T-cell leukemia/lymphoma (ATL/L) was confirmed by the presence of monoclonal integration of HTLV-I proviral DNA in biopsied specimen. This case, showing a pattern of P.A. (-) and I.F. (+), is extremely unusual, because I.F. and P.A. show highly close correlation. Thus, it is important to employ different methods for screening of anti-ATLA antibodies in the diagnosis of ATL/L.

Development of novel monoclonal antibodies that define differentiation stages of human stromal (mesenchymal) stem cells

DEFF Research Database (Denmark)

Andersen, Ditte Caroline; Kortesidis, Angela; Zannettino, Andrew C W

2011-01-01

Human mesenchymal stem cells (hMSC) are currently being introduced for cell therapy, yet, antibodies specific for native and differentiated MSCs are required for their identification prior to clinical use. Herein, high quality antibodies against MSC surface proteins were developed by immunizing...... fewer differentiated alkaline phosphatase(+) cells compared to STRO-1(+/-)/Collagen VI(+) hMSC, suggesting that Collagen VI on the cell membrane exclusively defines differentiated MSCs. In conclusion, we have generated a panel of high quality antibodies to be used for characterization of MSCs...... mice with hMSC, and by using a panel of subsequent screening methods. Flow cytometry analysis revealed that 83.5, 1.1, and 8.5% of primary cultures of hMSC were double positive for STRO-1 and either of DJ 3, 9, and 18, respectively. However, none of the three DJ antibodies allowed enrichment...

The Rac Activator DOCK2 Mediates Plasma Cell Differentiation and IgG Antibody Production.

Science.gov (United States)

Ushijima, Miho; Uruno, Takehito; Nishikimi, Akihiko; Sanematsu, Fumiyuki; Kamikaseda, Yasuhisa; Kunimura, Kazufumi; Sakata, Daiji; Okada, Takaharu; Fukui, Yoshinori

2018-01-01

A hallmark of humoral immune responses is the production of antibodies. This process involves a complex cascade of molecular and cellular interactions, including recognition of specific antigen by the B cell receptor (BCR), which triggers activation of B cells and differentiation into plasma cells (PCs). Although activation of the small GTPase Rac has been implicated in BCR-mediated antigen recognition, its precise role in humoral immunity and the upstream regulator remain elusive. DOCK2 is a Rac-specific guanine nucleotide exchange factor predominantly expressed in hematopoietic cells. We found that BCR-mediated Rac activation was almost completely lost in DOCK2-deficient B cells, resulting in defects in B cell spreading over the target cell-membrane and sustained growth of BCR microclusters at the interface. When wild-type B cells were stimulated in vitro with anti-IgM F(ab') 2 antibody in the presence of IL-4 and IL-5, they differentiated efficiently into PCs. However, BCR-mediated PC differentiation was severely impaired in the case of DOCK2-deficient B cells. Similar results were obtained in vivo when DOCK2-deficient B cells expressing a defined BCR specificity were adoptively transferred into mice and challenged with the cognate antigen. In addition, by generating the conditional knockout mice, we found that DOCK2 expression in B-cell lineage is required to mount antigen-specific IgG antibody. These results highlight important role of the DOCK2-Rac axis in PC differentiation and IgG antibody responses.

Method for extending the useful shelf-life of refrigerated red blood cells by flushing with inert gas

Science.gov (United States)

Bitensky, M.W.; Yoshida, Tatsuro

1997-04-29

A method is disclosed using oxygen removal for extending the useful shelf-life of refrigerated red blood cells. A cost-effective, 4 C storage procedure that preserves red cell quality and prolongs post-transfusion in vivo survival is described. Preservation of adenosine triphosphate levels and reduction in hemolysis and in membrane vesicle production of red blood cells stored at 4 C for prolonged periods of time is achieved by removing oxygen from the red blood cells at the time of storage; in particular, by flushing with an inert gas. Adenosine triphosphate levels of the stored red blood cells are boosted in some samples by addition of ammonium phosphate. 4 figs.

A Recombinant Antibody with the Antigen-Specific, Major Histocompatibility Complex-Restricted Specificity of T Cells

Science.gov (United States)

Andersen, Peter S.; Stryhn, Anette; Hansen, Bjarke E.; Fugger, Lars; Engberg, Jan; Buus, Soren

1996-03-01

Specific recognition of peptide/major histocompatibility complex (MHC) molecule complexes by the T-cell receptor is a key reaction in the specific immune response. Antibodies against peptide/MHC complexes would therefore be valuable tools in studying MHC function and T-cell recognition and might lead to novel approaches in immunotherapy. However, it has proven difficult to generate antibodies with the specificity of T cells by conventional hybridoma techniques. Here we report that the phage display technology is a feasible alternative to generate antibodies recognizing specific, predetermined peptide/MHC complexes.

Quantifying changes in the cellular thiol-disulfide status during differentiation of B cells into antibody-secreting plasma cells

DEFF Research Database (Denmark)

Hansen, Rosa Rebecca Erritzøe; Otsu, Mieko; Braakman, Ineke

2013-01-01

by the differentiation, steady-state levels of glutathionylated protein thiols are less than 0.3% of the total protein cysteines, even in fully differentiated cells, and the overall protein redox state is not affected until late in differentiation, when large-scale IgM production is ongoing. A general expansion......Plasma cells produce and secrete massive amounts of disulfide-containing antibodies. To accommodate this load on the secretory machinery, the differentiation of resting B cells into antibody-secreting plasma cells is accompanied by a preferential expansion of the secretory compartments of the cells...... of the ER does not affect global protein redox status until an extensive production of cargo proteins has started....

Antibodies to the human T-cell lymphoma/leukemia virus type I in Dutch haemophiliacs

NARCIS (Netherlands)

Goudsmit, J.; Miedema, F.; Breederveld, C.; Terpstra, F.; Roos, M.; Schellekens, P.; Melief, C.

1986-01-01

95 Dutch haemophiliacs were tested for antibodies to membrane antigens on cells infected with human T-cell leukemia virus type I (HTLV-I-MA) by indirect immunofluorescence and to purified HTLV-I by enzyme-linked immunosorbent assay. Antibodies to HTLV-I-MA were present in 8 of 95 (8%) haemophiliacs,

Method and cell lines for the production of monoclonal antibodies to human glycophorin A

Science.gov (United States)

Bigbee, W.L.; Fong, S.S.N.; Jensen, R.H.; Vanderlaan, M.

Cloned mouse hybridoma cell lines have been established which continuously produce antibodies that differentiate between the M and N forms of human glycophorin A. These antibodies have potential application as human blood group reagents, as markers for terminally differentiated erythroid cells and as immunofluorescent labels of somatically variant human erythrocytes.

Fab antibody fragment-functionalized liposomes for specific targeting of antigen-positive cells

Czech Academy of Sciences Publication Activity Database

Ohradanova-Repic, A.; Nogueira, E.; Hartl, I.; Gomes, A.C.; Preto, A.; Steinhuber, E.; Muehlgrabner, V.; Repic, M.; Kuttke, M.; Zwirzitz, A.; Prouza, M.; Suchánek, M.; Wozniak-Knopp, G.; Hořejší, Václav; Schabbauer, G.; Cavaco-Paulo, A.; Stockinger, H.

2018-01-01

Roč. 14, č. 1 (2018), s. 123-130 ISSN 1549-9634 Institutional support: RVO:68378050 Keywords : Active targeting * Liposome functionalization * Immunoliposome * Antibody engineering * Recombinant Fab antibody fragment Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Cell biology Impact factor: 5.720, year: 2016

Proteomic analysis identifies differentially expressed proteins after red propolis treatment in Hep-2 cells.

Science.gov (United States)

Frozza, Caroline Olivieri da Silva; Ribeiro, Tanara da Silva; Gambato, Gabriela; Menti, Caroline; Moura, Sidnei; Pinto, Paulo Marcos; Staats, Charley Christian; Padilha, Francine Ferreira; Begnini, Karine Rech; de Leon, Priscila Marques Moura; Borsuk, Sibele; Savegnago, Lucielli; Dellagostin, Odir; Collares, Tiago; Seixas, Fabiana Kömmling; Henriques, João Antonio Pêgas; Roesch-Ely, Mariana

2014-01-01

Here we investigated alterations in the protein profile of Hep-2 treated with red propolis using two-dimensional electrophoresis associated to mass spectrometry and apoptotic rates of cells treated with and without red propolis extracts through TUNEL and Annexin-V assays. A total of 325 spots were manually excised from the two-dimensional gel electrophoresis and 177 proteins were identified using LC-MS-MS. Among all proteins identified that presented differential expression, most were down-regulated in presence of red propolis extract at a concentration of 120 μg/mL (IC50): GRP78, PRDX2, LDHB, VIM and TUBA1A. Only two up-regulated proteins were identified in this study in the non-cytotoxic (6 μg/mL) red propolis treated group: RPLP0 and RAD23B. TUNEL staining assay showed a markedly increase in the mid- to late-stage apoptosis of Hep-2 cells induced by red propolis at concentrations of 60 and 120 μg/mL when compared with non-treated cells. The increase of late apoptosis was confirmed by in situ Annexin-V analysis in which red propolis extract induced late apoptosis in a dose-dependent manner. The differences in tumor cell protein profiles warrant further investigations including isolation of major bioactive compounds of red propolis in different cell lines using proteomics and molecular tests to validate the protein expression here observed. Copyright © 2013 Elsevier Ltd. All rights reserved.

Certain Red Blood Cell Indices of Maternal and Umbilical Cord ...

African Journals Online (AJOL)

Uche

Background: Umbilical cord blood analysis may give a clue to the state of health of both pregnant mothers and their neonates. However ... Keywords: Umbilical cord blood; maternal blood; haemoglobin concentration; packed cell volume; red cell indices. Received on .... The packed cell volume was measured using the.

Impact of azacitidine on red blood cell alloimmunisation in myelodysplastic syndrome.

Science.gov (United States)

Ortiz, Sebastián; Orero, Maria T; Javier, Karla; Villegas, Carolina; Luna, Irene; Pérez, Pedro; Roig, Mónica; López, María; Costa, Sofía; Carbonell, Félix; Collado, Rosa; Ivars, David; Linares, Mariano

2017-09-01

The incidence of alloimmunisation in myelodysplastic syndromes (MDS) during the era of supportive treatment ranges from 9 to 56%. However, it is unknown if the widespread use of hypomethylating agents has changed the risk of immunisation. The aim of this study is to evaluate the impact of azacitidine (AZA) therapy on red blood cell (RBC) alloimmunisation in transfused patients with MDS, myelodysplastic syndromes/myeloproliferative syndromes (MDS/MPS) and secondary acute myeloid leukaemia (AML). We have analysed retrospectively all patients with MDS, MDS/MPS and secondary AML from MDS, who received their first transfusion in our hospital between January 1995 and December 2014. We have assessed the impact of age, sex, RBC and platelets units transfused, and AZA treatment on developing alloantibodies. In our study, the number of RBC units transfused increased the risk of developing alloantibodies. However aging and the treatment with AZA were associated with a lower rate of alloimmunisation. Patients with MDS, MDS/MPS and secondary AML who received treatment with AZA developed RBC antibodies at a lower rate than control group. We suggest that aging and immunosuppression due to AZA therapy could develop an immunological tolerance with a weak response to allotransfusions.

Clinical Utility of Red Cell Distribution Width in Alcoholic and Non-alcoholic Liver Cirrhosis

OpenAIRE

Milić, Sandra; Mikolašević, Ivana; Radić, Mladen; Hauser, Goran; Štimac, Davor

2011-01-01

Red blood cell distribution width (RDW) is a measure of the variation of red blood cell width that is reported as a part of standard complete blood count. Red blood cell distribution width results are often used together with mean corpuscular volume (MCV) results to figure out mixed anemia. The aim of our study was to compare the values of RDW in alcoholic and non-alcoholic liver cirrhosis and to determine if RDW follows the severity of disease according to Child-Pugh score. We re...

An Fc engineering approach that modulates antibody-dependent cytokine release without altering cell-killing functions.

Science.gov (United States)

Kinder, Michelle; Greenplate, Allison R; Strohl, William R; Jordan, Robert E; Brezski, Randall J

2015-01-01

Cytotoxic therapeutic monoclonal antibodies (mAbs) often mediate target cell-killing by eliciting immune effector functions via Fc region interactions with cellular and humoral components of the immune system. Key functions include antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), and complement-dependent cytotoxicity (CDC). However, there has been increased appreciation that along with cell-killing functions, the induction of antibody-dependent cytokine release (ADCR) can also influence disease microenvironments and therapeutic outcomes. Historically, most Fc engineering approaches have been aimed toward modulating ADCC, ADCP, or CDC. In the present study, we describe an Fc engineering approach that, while not resulting in impaired ADCC or ADCP, profoundly affects ADCR. As such, when peripheral blood mononuclear cells are used as effector cells against mAb-opsonized tumor cells, the described mAb variants elicit a similar profile and quantity of cytokines as IgG1. In contrast, although the variants elicit similar levels of tumor cell-killing as IgG1 with macrophage effector cells, the variants do not elicit macrophage-mediated ADCR against mAb-opsonized tumor cells. This study demonstrates that Fc engineering approaches can be employed to uncouple macrophage-mediated phagocytic and subsequent cell-killing functions from cytokine release.

Consequences of dysregulated complement regulators on red blood cells

NARCIS (Netherlands)

Thielen, Astrid J. F.; Zeerleder, Sacha; Wouters, Diana

2018-01-01

The complement system represents the first line of defense that is involved in the clearance of pathogens, dying cells and immune complexes via opsonization, induction of an inflammatory response and the formation of a lytic pore. Red blood cells (RBCs) are very important for the delivery of oxygen

Training the next generation analyst using red cell analytics

Science.gov (United States)

Graham, Meghan N.; Graham, Jacob L.

2016-05-01

We have seen significant change in the study and practice of human reasoning in recent years from both a theoretical and methodological perspective. Ubiquitous communication coupled with advances in computing and a plethora of analytic support tools have created a push for instantaneous reporting and analysis. This notion is particularly prevalent in law enforcement, emergency services and the intelligence community (IC), where commanders (and their civilian leadership) expect not only a birds' eye view of operations as they occur, but a play-by-play analysis of operational effectiveness. This paper explores the use of Red Cell Analytics (RCA) as pedagogy to train the next-gen analyst. A group of Penn State students in the College of Information Sciences and Technology at the University Park campus of The Pennsylvania State University have been practicing Red Team Analysis since 2008. RCA draws heavily from the military application of the same concept, except student RCA problems are typically on non-military in nature. RCA students utilize a suite of analytic tools and methods to explore and develop red-cell tactics, techniques and procedures (TTPs), and apply their tradecraft across a broad threat spectrum, from student-life issues to threats to national security. The strength of RCA is not always realized by the solution but by the exploration of the analytic pathway. This paper describes the concept and use of red cell analytics to teach and promote the use of structured analytic techniques, analytic writing and critical thinking in the area of security and risk and intelligence training.

Antibody-dependent NK cell activation is associated with late kidney allograft dysfunction and the complement-independent alloreactive potential of donor-specific antibodies

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Tristan Legris

2016-08-01

Full Text Available Although kidney transplantation remains the best treatment for end-stage renal failure, it is limited by chronic humoral aggression of the graft vasculature by donor-specific antibodies (DSAs. The complement-independent mechanisms that lead to the antibody-mediated rejection (ABMR of kidney allografts remain poorly understood. Increasing lines of evidence have revealed the relevance of natural killer (NK cells as innate immune effectors of antibody-dependent cellular cytotoxicity, but few studies have investigated their alloreactive potential in the context of solid organ transplantation. Our study aimed to investigate the potential contribution of the antibody-dependent alloreactive function of NK cells to kidney graft dysfunction. We first conducted an observational study to investigate whether the cytotoxic function of NK cells is associated with chronic allograft dysfunction. The NK-Cellular Humoral Activation Test (NK-CHAT was designed to evaluate the recipient and antibody-dependent reactivity of NK cells against allogeneic target cells. The release of CD107a/Lamp1+ cytotoxic granules, resulting from the recognition of rituximab-coated B cells by NK cells, was analyzed in 148 kidney transplant recipients (KTRs, mean graft duration: 6.2 years. Enhanced ADCC responsiveness was associated with reduced graft function and identified as an independent risk factor predicting a decline in the estimated glomerular filtration rate (eGFR over a 1-year period (hazard ratio: 2.83. In a second approach, we used the NK-CHAT to reveal the cytotoxic potential of circulating alloantibodies in vitro. The level of CD16 engagement resulting from the in vitro recognition of serum-coated allogeneic B cells or splenic cells was further identified as a specific marker of DSA-induced ADCC. The NK-CHAT scoring of sera obtained from 40 patients at the time of transplant biopsy was associated with ABMR diagnosis. Our findings indicate that despite the administration

Development of hyper osmotic resistant CHO host cells for enhanced antibody production.

Science.gov (United States)

Kamachi, Yasuharu; Omasa, Takeshi

2018-04-01

Cell culture platform processes are generally employed to shorten the duration of new product development. A fed-batch process with continuous feeding is a conventional platform process for monoclonal antibody production using Chinese hamster ovary (CHO) cells. To establish a simplified platform process, the feeding method can be changed from continuous feed to bolus feed. However, this change induces a rapid increase of osmolality by the bolus addition of nutrients. The increased osmolality suppresses cell culture growth, and the final product concentration is decreased. In this study, osmotic resistant CHO host cells were developed to attain a high product concentration. To establish hyper osmotic resistant CHO host cells, CHO-S host cells were passaged long-term in a hyper osmotic basal medium. There were marked differences in cell growth of the original and established host cells under iso- (328 mOsm/kg) or hyper-osmolality (over 450 mOsm/kg) conditions. Cell growth of the original CHO host cells was markedly decreased by the induction of osmotic stress, whereas cell growth of the hyper osmotic resistant CHO host cells was not affected. The maximum viable cell concentration of hyper osmotic resistant CHO host cells was 132% of CHO-S host cells after the induction of osmotic stress. Moreover, the hyper osmotic resistant characteristic of established CHO host cells was maintained even after seven passages in iso-osmolality basal medium. The use of hyper osmotic resistance CHO host cells to create a monoclonal antibody production cell line might be a new approach to increase final antibody concentrations with a fed-batch process. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Antibody Fc engineering improves frequency and promotes kinetic boosting of serial killing mediated by NK cells

Science.gov (United States)

Romain, Gabrielle; Senyukov, Vladimir; Rey-Villamizar, Nicolas; Merouane, Amine; Kelton, William; Liadi, Ivan; Mahendra, Ankit; Charab, Wissam; Georgiou, George; Roysam, Badrinath; Lee, Dean A.

2014-01-01

The efficacy of most therapeutic monoclonal antibodies (mAbs) targeting tumor antigens results primarily from their ability to elicit potent cytotoxicity through effector-mediated functions. We have engineered the fragment crystallizable (Fc) region of the immunoglobulin G (IgG) mAb, HuM195, targeting the leukemic antigen CD33, by introducing the triple mutation Ser293Asp/Ala330Leu/Ile332Glu (DLE), and developed Time-lapse Imaging Microscopy in Nanowell Grids to analyze antibody-dependent cell-mediated cytotoxicity kinetics of thousands of individual natural killer (NK) cells and mAb-coated target cells. We demonstrate that the DLE-HuM195 antibody increases both the quality and the quantity of NK cell-mediated antibody-dependent cytotoxicity by endowing more NK cells to participate in cytotoxicity via accrued CD16-mediated signaling and by increasing serial killing of target cells. NK cells encountering targets coated with DLE-HuM195 induce rapid target cell apoptosis by promoting simultaneous conjugates to multiple target cells and induce apoptosis in twice the number of target cells within the same period as the wild-type mAb. Enhanced target killing was also associated with increased frequency of NK cells undergoing apoptosis, but this effect was donor-dependent. Antibody-based therapies targeting tumor antigens will benefit from a better understanding of cell-mediated tumor elimination, and our work opens further opportunities for the therapeutic targeting of CD33 in the treatment of acute myeloid leukemia. PMID:25232058

Deep coverage mouse red blood cell proteome: a first comparison with the human red blood cell

DEFF Research Database (Denmark)

Pasini, Erica M; Kirkegaard, Morten; Salerno, Doris

2008-01-01

Mice have close genetic/physiological relationships to humans, breed rapidly, and can be genetically modified, making them the most used mammal in biomedical research. Because the red blood cell (RBC) is the sole gas transporter in vertebrates, diseases of the RBC are frequently severe; much...... proteome have been confirmed here. This comparison sheds light on several open issues in RBC biology and provides a departure point for more comprehensive understanding of RBC function....

Structural Changes in the Surface of Red Blood Cell Membranes during Long-Term Donor Blood Storage

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V. V. Moroz

2012-01-01

Full Text Available Objective: to study changes in the surface of red blood cell membranes of donor blood at the macro- and ultrastructural level during its storage for 30 days and to evaluate the functional state of the red blood cell membrane during the whole storage period. Material and methods. The investigation was conducted on human whole blood and packed red blood cells placed in the specialized packs containing the preservative CPDA-1, by using calibrated electroporation and atomic force microscopy and measuring plasma pH. Conclusion. The long-term, up to 30-day, storage of whole blood and packed red blood cells at 4°C was attended by lower plasma pH and increased hemolysis rate constant during calibrated electroporation and by the development of oxidative processes. The hemolysis rate constant was also higher in the packed red blood cells than that in the whole blood. On days 5—6, the membrane structure showed defects that developed, as the blood was stored, and caused irreversible cell membrane damage by day 30. Key words: donor blood, red blood cell membranes, atomic force microscopy.

Effects of 4000 rad irradiation on the in vitro storage properties of packed red cells

International Nuclear Information System (INIS)

Moore, G.L.; Ledford, M.E.

1985-01-01

Immunosuppressed patients who require red cell transfusions receive irradiated (1500-3000 rad) packed red cells. These cells are irradiated immediately before infusion. If a large group of patients become immunosuppressed due to exposure to radiation or chemicals, the ability to supply large volumes of irradiated blood at the time of use might not be possible. An alternate solution to providing quantities of irradiated blood is to irradiate the units prior to storage. This study presents in vitro data comparing storage of paired packed red cell units either irradiated or not irradiated. Five units of fresh blood drawn into citrate-phosphate-dextrose-adenine (CPDA-1) were packed to a hematocrit of 75 +/- 1 percent, and then each unit was divided in two equal parts. One of each pair was irradiated (4000 rads), and both parts of each unit were stored for 35 days at 4 degrees C. Samples were analyzed every 7 days. Irradiation caused a slight drop in red cell adenosine triphosphate and 2,3 diphosphoglycerate and a slight increase in plasma hemoglobin compared to controls. Methemoglobin, pH, and glucose consumption were identical to the controls. The evidence indicates that irradiation did not cause biochemical or metabolic changes in the red cells that would lead us to suspect a difference between irradiated and nonirradiated stored red cells in function or viability. These negative findings require in vivo confirmation

Measurement of the viability of stored red cells by the single-isotope technique using 51Cr. Analysis of validity

International Nuclear Information System (INIS)

Beutler, E.; West, C.

1984-01-01

A single-isotope 51 Cr method often is used to evaluate the viability of stored red cells. In this technique, the red cell mass is measured by back-extrapolation to time zero (t0) of the radioactivity of the blood between 5 and 20 minutes after infusion of the sample. If there is early destruction of stored cells, this method provides an overestimate of the red cell mass and, hence, of the viability of the stored cells. Freshly drawn red cells from normal donors were labeled with /sup 99m/Tc, and cells from the same donor which had been stored in citrate-phosphate-dextrose-adenine-one (CPDA-1) for periods ranging from 7 to 49 days were labeled with 51 Cr. A comparison of the ''true red cell mass'' as determined with /sup 99m/Tc with the back-extrapolated red cell mass from stored 51 Cr-labeled cells has made it possible to define the magnitude of error introduced by early loss of red cells. The overestimation of red cell mass and viability was diminished if only the 51 Cr radioactivity between 5 and 15 minutes after infusion was used in back-extrapolating to t0. The degree of overestimation of red cell mass was greatest when the red cell viability had declined to very low levels. However, in the entire range of 10 to 80 percent viability, the overestimate of viability was usually less than 4 percent. The overestimate of viability proved to be quite similar for all samples and may be taken into account when using the single-isotope technique for measurement of red cell viability

Bone scan and red blood cell scan in a patient with epidermal naevus syndrome

International Nuclear Information System (INIS)

Becker, W.; Wolf, F.; Stosiek, N.; Peters, K.P.

1990-01-01

A bone scan and red blood cell scan in the rare epidermal naevus syndrome, associated with multiple haemangiomes of the bone and hypophosphataemic osteomalacia in a 20-year-old man are reported. The typical pattern of osteomalacia on the bone scan was associated with lesions of increased bone metabolism in the peripheral bones. The haemangiomas did not pool labelled red blood cells. Thus, the bone scan seems to be suited for diagnosing the complete extent of haemangiomas in bone, but they could not be specifically proven by red blood cell pooling. (orig.)

Human Cell Line-Derived Monoclonal IgA Antibodies for Cancer Immunotherapy

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Felix Hart

2017-05-01

Full Text Available IgA antibodies have great potential to improve the functional diversity of current IgG antibody-based cancer immunotherapy options. However, IgA production and purification is not well established, which can at least in part be attributed to the more complex glycosylation as compared to IgG antibodies. IgA antibodies possess up to five N-glycosylation sites within their constant region of the heavy chain as compared to one site for IgG antibodies. The human GlycoExpress expression system was developed to produce biotherapeutics with optimized glycosylation and used here to generate a panel of IgA isotype antibodies directed against targets for solid (TA-mucin 1, Her2, EGFR, Thomsen–Friedenreich and hematological (CD20 cancer indications. The feasibility of good manufacturing practice was shown by the production of 11 g IgA within 35 days in a one liter perfusion bioreactor, and IgA antibodies in high purity were obtained after purification. The monoclonal IgA antibodies possessed a high sialylation degree, and no non-human glycan structures were detected. Kinetic analysis revealed increased avidity antigen binding for IgA dimers as compared to monomeric antibodies. The IgA antibodies exhibited potent Fab- and Fc-mediated functionalities against cancer cell lines, whereby especially granulocytes are recruited. Therefore, for patients who do not sufficiently benefit from therapeutic IgG antibodies, IgA antibodies may complement current regiment options and represent a promising strategy for cancer immunotherapy. In conclusion, a panel of novel biofunctional IgA antibodies with human glycosylation was successfully generated.

[Monoclonal antibodies ICO-02 to blast cell antigens in patients with chronic myeloleukemia in blast crisis].

Science.gov (United States)

Baryshnikov, A Iu

1984-01-01

Mice were immunized with blood cells of a patient with chronic granulocytic leukemia, and their cells were subsequently used for the preparation of hybridoma ICO-02. This hybridoma is continuously producing monoclonal antibodies which reacted with cells in 4 out of 13 patients with blastic crisis of chronic granulocytic leukemia and in 6 out of 38 patients with acute lymphoblastic leukemia. Antibodies reacted with blast cells in 2 out of 3 patients with undifferentiated blastic crisis of chronic myelocytic leukemia and in 2 out of 5 patients with lymphoid variant of blastic crisis of chronic granulocytic leukemia. Cells of 6 patients with acute lymphoblastic leukemia which reacted with the monoclonal antibodies had immunological markers of T lymphocytes bone-marrow precursors. Monoclonal antibodies did not react with cells of blood and bone marrow from healthy people and from patients with chronic lymphocytic leukemia, acute myeloblastic leukemia, acute myelomonocytic leukemia, acute monoblastic leukemia and lymphosarcoma.

Hemolytic disease of fetus and newborn due to maternal red blood cell alloantibodies in the Malay population

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Mohd Nazri Hassan

2014-01-01

Full Text Available Background: Maternal red blood cell (RBC alloimmunization may lead to production of harmful antibodies that result in hemolytic disease of fetus and newborn (HDFN. There is insufficient data on the prevalence of HDFN due to RBC alloantibodies in the Malay neonatal population. Aim: The aim of this study was to determine the incidence of HDFN in the Malay neonatal population due to clinically significant RBC alloantibodies. Subjects and Methods: A cross sectional study was conducted in Transfusion Medicine Unit, Hospital Universitiy Sains Malaysia over one year period from January to December 2009. A total of 5163 Malay pregnant women who attended labor room for delivery were collected and analyzed prospectively. The blood samples were subjected to the standard immunohematological procedure for RBC antibody screening and identification using reagents of Diamed-ID Gel microtyping system. All the newborns with RBC alloantibody were investigated for the evidence of HDFN. Results: Thirty (0.58% women were found to have clinically significant RBC alloantibodies. Most of the alloantibodies belonged to Rhesus (Rh system (56.7% where anti-E (33.3% was the most common followed by anti-D (10.0%. Rh antibodies were the main cause of HDFN in fourteen (0.27% neonates. Anti-D and anti-c were identified to cause moderate to very severe HDFN . Conclusions: With the low prevalence of clinically significant RBC alloantibodies and HDFN, routine antenatal antibody screening practice may not be advised as a routine practice at present, preferably reserved for those women of RhD negative or with history of HDFN, significantly of those attributed to anti-c.

Red Cell Indexes Made Easy Using an Interactive Animation: Do Students and Their Scores Concur?

Science.gov (United States)

Kachroo, Upasana; Vinod, Elizabeth; Balasubramanian, Sivakumar; W., Jesi; Prince, Neetu

2018-01-01

A good understanding of red cell indexes can aid medical students in a considerable manner, serving as a basis to unravel both concepts in red cell physiology and abnormalities associated with the same. In this study, we tried to assess whether an interactive animation was helpful in improving student comprehension and understanding of red cell…

Monoclonal antibody studies in B(non-T)-cell malignancies.

Science.gov (United States)

Shimoyama, M; Minato, K; Tobinai, K; Nagai, M; Hirose, M

1983-09-01

Tumor cells suspensions prepared from 129 B- or non-T cell malignancies were investigated with a panel of 10 monoclonal antibodies and conventional surface marker techniques. Surface immunoglobulin (sIg) and B1 antigen proved to be the most useful markers for B-cell lineage. Six major subtypes of acute lymphoblastic leukemia (ALL) of non-T cell nature are now recognized by these immunological techniques, including null-ALL, Ia-ALL, lymphoid stem cell ALL, pre-pre-B ALL, pre-B ALL and B-ALL. In cases of chronic leukemias and lymphomas of non-T cell nature, 80% of the tumor was defined by sIg and 88% by B1 antigen as definitely of B-cell lineage. The clonal character was also defined in 68% of the tumor on the basis of the detection of predominant single light chain in sIg. Ia-like antigen was detected in almost all cases (96%). Leukemic cells from all cases of chronic lymphocytic leukemia (CLL), chronic lymphosarcoma cell leukemia (CLsCL) and hairy cell leukemia (HCL) reacted with OKIa1 and anti-B1, and leukemic cells from most of them with anti-pan T monoclonal antibody (10.2). In more than half of CLL and CLsCL, leukemic cells were reactive with J5, OKM1, 9.6 and OKT8, but not with OKT3, OKT4 and OKT6. HCL cells had almost the same reactivity with these monoclonal antibodies as CLL and CLsCL cells except that J5 remained unreactive. These results indicated that Japanese CLL, CLsCL and HCL were different from Western ones at least with respect to surface marker characteristics. In cases of lymphomas, heavy chains of sIg were expressed in polyclonal fashion, especially in follicular lymphoma and diffuse lymphomas of medium sized cell type and large cell type, indicating that lymphomas of these types may originate from follicular center cells of the heavy chain switching stage. Anti-T monoclonals were also reactive with lymphoma cells. In about half of follicular lymphomas and diffuse lymphomas of the medium sized cell type, lymphoma cells reacted with 10.2, and less

Antibody formation in mouse bone marrow. IV. The influence of splenectomy on the bone marrow plaque-forming cell response to sheep red blood cells

International Nuclear Information System (INIS)

Benner, R.; Oudenaren, A. van

1975-01-01

Mouse bone marrow is barely capable of plaque-forming cell (PFC) activity during the primary response to sheep red blood cells (SRBC). However, during the secondary response, it becomes the major center of activity containing IgM-, IgG- and IgA-PFC. In the present paper the influence of splenectomy was studied on primary and secondary PFC activity in the bone marrow. Differences in primary and secondary bone marrow PFC responses are probably related to the presence of B and T memory cells in situ. Therefore the effect of splenectomy on the appearance of B and T memory cells in the bone marrow was also investigated. iv.plenectomy before intravenous (iv) immunization with 4 x 10 8 SRBC prevented any primary PFC activity in the bone marrow. The influence of splenectomy before priming on secondary PFC activity in the bone marrow depended on the priming dose of SRBC. Splenectomy before priming with 10 7 SRBC iv completely prevented IgM-, IgG-, and IgA-PFC activity in the bone marrow upon subsequent boosting with 4 x 10 8 SRBC iv. By means of cell transfer experiments it was shown that after splenectomy no B or T memory cells appeared in the bone marrow after priming with 10 7 SRBC iv. Cell transfer experiments showed that splenectomy before priming with 10 7 SRBC iv not only interfered with the appearance of B and T memory cells in the bone marrow, but also with the appearance of B memory cells in peripheral lymph nodes, mesenteric lymph node, Peyer's patches, thymus, and blood. Immunization of spenectomized mice with 4 x 10 8 SRBC iv induced the appearance of B memory cells in peripheral lymph nodes, mesenteric lymph node, Peyer's patches, thymus, and blood

Enhanced killing of chordoma cells by antibody-dependent cell-mediated cytotoxicity employing the novel anti-PD-L1 antibody avelumab

OpenAIRE

Fujii, Rika; Friedman, Eitan R.; Richards, Jacob; Tsang, Kwong Y.; Heery, Christopher R.; Schlom, Jeffrey; Hodge, James W.

2016-01-01

Chordoma, a rare bone tumor derived from the notochord, has been shown to be resistant to conventional therapies. Checkpoint inhibition has shown great promise in immune-mediated therapy of diverse cancers. The anti-PD-L1 mAb avelumab is unique among checkpoint inhibitors in that it is a fully human IgG1 capable of mediating antibody-dependent cell-mediated cytotoxicity (ADCC) of PD-L1-expressing tumor cells. Here, we investigated avelumab as a potential therapy for chordoma. We examined 4 ch...

The Rac Activator DOCK2 Mediates Plasma Cell Differentiation and IgG Antibody Production

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Miho Ushijima

2018-02-01

Full Text Available A hallmark of humoral immune responses is the production of antibodies. This process involves a complex cascade of molecular and cellular interactions, including recognition of specific antigen by the B cell receptor (BCR, which triggers activation of B cells and differentiation into plasma cells (PCs. Although activation of the small GTPase Rac has been implicated in BCR-mediated antigen recognition, its precise role in humoral immunity and the upstream regulator remain elusive. DOCK2 is a Rac-specific guanine nucleotide exchange factor predominantly expressed in hematopoietic cells. We found that BCR-mediated Rac activation was almost completely lost in DOCK2-deficient B cells, resulting in defects in B cell spreading over the target cell-membrane and sustained growth of BCR microclusters at the interface. When wild-type B cells were stimulated in vitro with anti-IgM F(ab′2 antibody in the presence of IL-4 and IL-5, they differentiated efficiently into PCs. However, BCR-mediated PC differentiation was severely impaired in the case of DOCK2-deficient B cells. Similar results were obtained in vivo when DOCK2-deficient B cells expressing a defined BCR specificity were adoptively transferred into mice and challenged with the cognate antigen. In addition, by generating the conditional knockout mice, we found that DOCK2 expression in B-cell lineage is required to mount antigen-specific IgG antibody. These results highlight important role of the DOCK2–Rac axis in PC differentiation and IgG antibody responses.

B cell-targeted therapy with anti-CD20 monoclonal antibody in a mouse model of Graves' hyperthyroidism.

Science.gov (United States)

Ueki, I; Abiru, N; Kobayashi, M; Nakahara, M; Ichikawa, T; Eguchi, K; Nagayama, Y

2011-03-01

Graves' disease is a B cell-mediated and T cell-dependent autoimmune disease of the thyroid which is characterized by overproduction of thyroid hormones and thyroid enlargement by agonistic anti-thyrotrophin receptor (TSHR) autoantibody. In addition to antibody secretion, B cells have recently been recognized to function as antigen-presenting/immune-modulatory cells. The present study was designed to evaluate the efficacy of B cell depletion by anti-mouse (m) CD20 monoclonal antibody (mAb) on Graves' hyperthyroidism in a mouse model involving repeated injection of adenovirus expressing TSHR A-subunit (Ad-TSHR289). We observe that a single injection of 250 µg/mouse anti-mCD20 mAb eliminated B cells efficiently from the periphery and spleen and to a lesser extent from the peritoneum for more than 3 weeks. B cell depletion before immunization suppressed an increase in serum immunoglobulin (Ig)G levels, TSHR-specific splenocyte secretion of interferon (IFN)-γ, anti-TSHR antibody production and development of hyperthyroidism. B cell depletion 2 weeks after the first immunization, a time-point at which T cells were primed but antibody production was not observed, was still effective at inhibiting antibody production and disease development without inhibiting splenocyte secretion of IFN-γ. By contrast, B cell depletion in hyperthyroid mice was therapeutically ineffective. Together, these data demonstrate that B cells are critical not only as antibody-producing cells but also as antigen-presenting/immune-modulatory cells in the early phase of the induction of experimental Graves' hyperthyroidism and, although therapeutically less effective, B cell depletion is highly efficient for preventing disease development. © 2011 The Authors. Clinical and Experimental Immunology © 2011 British Society for Immunology.

B cell-targeted therapy with anti-CD20 monoclonal antibody in a mouse model of Graves' hyperthyroidism

Science.gov (United States)

Ueki, I; Abiru, N; Kobayashi, M; Nakahara, M; Ichikawa, T; Eguchi, K; Nagayama, Y

2011-01-01

Graves' disease is a B cell-mediated and T cell-dependent autoimmune disease of the thyroid which is characterized by overproduction of thyroid hormones and thyroid enlargement by agonistic anti-thyrotrophin receptor (TSHR) autoantibody. In addition to antibody secretion, B cells have recently been recognized to function as antigen-presenting/immune-modulatory cells. The present study was designed to evaluate the efficacy of B cell depletion by anti-mouse (m) CD20 monoclonal antibody (mAb) on Graves' hyperthyroidism in a mouse model involving repeated injection of adenovirus expressing TSHR A-subunit (Ad-TSHR289). We observe that a single injection of 250 µg/mouse anti-mCD20 mAb eliminated B cells efficiently from the periphery and spleen and to a lesser extent from the peritoneum for more than 3 weeks. B cell depletion before immunization suppressed an increase in serum immunoglobulin (Ig)G levels, TSHR-specific splenocyte secretion of interferon (IFN)-γ, anti-TSHR antibody production and development of hyperthyroidism. B cell depletion 2 weeks after the first immunization, a time-point at which T cells were primed but antibody production was not observed, was still effective at inhibiting antibody production and disease development without inhibiting splenocyte secretion of IFN-γ. By contrast, B cell depletion in hyperthyroid mice was therapeutically ineffective. Together, these data demonstrate that B cells are critical not only as antibody-producing cells but also as antigen-presenting/immune-modulatory cells in the early phase of the induction of experimental Graves' hyperthyroidism and, although therapeutically less effective, B cell depletion is highly efficient for preventing disease development. PMID:21235532

T cell regulation of the thymus-independent antibody response to trinitrophenylated-Brucella abortus (TNP-BA)

Energy Technology Data Exchange (ETDEWEB)

Tanay, A.; Strober, S.

1985-06-01

The authors have previously observed a reduction of the T cell-dependent primary antibody response to dinitrophenylated keyhole limpet hemocyanin, and an enhancement of the T cell-independent response to trinitrophenylated Brucella abortus (TNP-BA) in BALB/c mice after treatment with total lymphoid irradiation (TLI). To elucidate the relative contribution of T and B cells to the enhanced T cell-independent antibody responses after TLI, a syngeneic primary adoptive transfer system was utilized whereby irradiated hosts were reconstituted with unfractionated spleen cells or a combination of purified T and B cells from TLI-treated and untreated control mice. Antibody responses of purified splenic B cells from TLI-treated BALB/c mice (TLI/B) to TNP-BA were enhanced 10-fold as compared with those of unfractionated (UF) spleen cells or B cells from normal (NL) BALB/c mice (NL/UF and NL/B, respectively). Splenic T cells from normal animals (NL/T) suppressed the anti-TNP-BA response of TLI/B by more than 100-fold. NL/T neither suppressed nor enhanced the response of NL/B. On the other hand, T cells from TLI-treated mice (TLI/T) enhanced by 100-fold the anti-TNP-BA response of NL/B, but neither suppressed nor enhanced the response of TLI/B. Thus, T cells can regulate the T cell-independent antibody response to TNP-BA. However, experimental manipulation of the T and B cell populations is needed to demonstrate the regulatory functions.

T cell regulation of the thymus-independent antibody response to trinitrophenylated-Brucella abortus (TNP-BA)

International Nuclear Information System (INIS)

Tanay, A.; Strober, S.

1985-01-01

The authors have previously observed a reduction of the T cell-dependent primary antibody response to dinitrophenylated keyhole limpet hemocyanin, and an enhancement of the T cell-independent response to trinitrophenylated Brucella abortus (TNP-BA) in BALB/c mice after treatment with total lymphoid irradiation (TLI). To elucidate the relative contribution of T and B cells to the enhanced T cell-independent antibody responses after TLI, a syngeneic primary adoptive transfer system was utilized whereby irradiated hosts were reconstituted with unfractionated spleen cells or a combination of purified T and B cells from TLI-treated and untreated control mice. Antibody responses of purified splenic B cells from TLI-treated BALB/c mice (TLI/B) to TNP-BA were enhanced 10-fold as compared with those of unfractionated (UF) spleen cells or B cells from normal (NL) BALB/c mice (NL/UF and NL/B, respectively). Splenic T cells from normal animals (NL/T) suppressed the anti-TNP-BA response of TLI/B by more than 100-fold. NL/T neither suppressed nor enhanced the response of NL/B. On the other hand, T cells from TLI-treated mice (TLI/T) enhanced by 100-fold the anti-TNP-BA response of NL/B, but neither suppressed nor enhanced the response of TLI/B. Thus, T cells can regulate the T cell-independent antibody response to TNP-BA. However, experimental manipulation of the T and B cell populations is needed to demonstrate the regulatory functions

Highly multiplexed and quantitative cell-surface protein profiling using genetically barcoded antibodies.

Science.gov (United States)

Pollock, Samuel B; Hu, Amy; Mou, Yun; Martinko, Alexander J; Julien, Olivier; Hornsby, Michael; Ploder, Lynda; Adams, Jarrett J; Geng, Huimin; Müschen, Markus; Sidhu, Sachdev S; Moffat, Jason; Wells, James A

2018-03-13

Human cells express thousands of different surface proteins that can be used for cell classification, or to distinguish healthy and disease conditions. A method capable of profiling a substantial fraction of the surface proteome simultaneously and inexpensively would enable more accurate and complete classification of cell states. We present a highly multiplexed and quantitative surface proteomic method using genetically barcoded antibodies called phage-antibody next-generation sequencing (PhaNGS). Using 144 preselected antibodies displayed on filamentous phage (Fab-phage) against 44 receptor targets, we assess changes in B cell surface proteins after the development of drug resistance in a patient with acute lymphoblastic leukemia (ALL) and in adaptation to oncogene expression in a Myc-inducible Burkitt lymphoma model. We further show PhaNGS can be applied at the single-cell level. Our results reveal that a common set of proteins including FLT3, NCR3LG1, and ROR1 dominate the response to similar oncogenic perturbations in B cells. Linking high-affinity, selective, genetically encoded binders to NGS enables direct and highly multiplexed protein detection, comparable to RNA-sequencing for mRNA. PhaNGS has the potential to profile a substantial fraction of the surface proteome simultaneously and inexpensively to enable more accurate and complete classification of cell states. Copyright © 2018 the Author(s). Published by PNAS.

Identification and red blood cell automated counting from blood smear images using computer-aided system.

Science.gov (United States)

Acharya, Vasundhara; Kumar, Preetham

2018-03-01

Red blood cell count plays a vital role in identifying the overall health of the patient. Hospitals use the hemocytometer to count the blood cells. Conventional method of placing the smear under microscope and counting the cells manually lead to erroneous results, and medical laboratory technicians are put under stress. A computer-aided system will help to attain precise results in less amount of time. This research work proposes an image-processing technique for counting the number of red blood cells. It aims to examine and process the blood smear image, in order to support the counting of red blood cells and identify the number of normal and abnormal cells in the image automatically. K-medoids algorithm which is robust to external noise is used to extract the WBCs from the image. Granulometric analysis is used to separate the red blood cells from the white blood cells. The red blood cells obtained are counted using the labeling algorithm and circular Hough transform. The radius range for the circle-drawing algorithm is estimated by computing the distance of the pixels from the boundary which automates the entire algorithm. A comparison is done between the counts obtained using the labeling algorithm and circular Hough transform. Results of the work showed that circular Hough transform was more accurate in counting the red blood cells than the labeling algorithm as it was successful in identifying even the overlapping cells. The work also intends to compare the results of cell count done using the proposed methodology and manual approach. The work is designed to address all the drawbacks of the previous research work. The research work can be extended to extract various texture and shape features of abnormal cells identified so that diseases like anemia of inflammation and chronic disease can be detected at the earliest.

Geometrical Aspects During Formation of Compact Aggregates of Red Blood Cells

Directory of Open Access Journals (Sweden)

Cardoso A.V.

2002-01-01

Full Text Available In the past forty years considerable progress has been achieved on the knowledge of human blood as a non-Newtonian shear-thinning suspension, whose initial state, that is at rest (stasis or at very low shear rates, has a gel-like internal structure which is destroyed as shear stress increases. The main goal of this communication is to describe the role of geometrical aspects during RBC (red blood cell aggregate formation, growth and compaction on naturally aggregate (porcine blood and non-aggregate (bovine blood samples. We consider how these aspects coupled with tension equilibrium are decisive to transform red cell linear roleaux to three-dimensional aggregates or clusters. Geometrical aspects are also crucial on the compaction of red blood cell aggregates. These densely packed aggregates could precipitate out of blood- either as dangerous deposits on arterial walls, or as clots which travel in suspension until they block some crucial capillary.

Membrane transport of anandamide through resealed human red blood cell membranes

DEFF Research Database (Denmark)

Bojesen, I.N.; Hansen, Harald S.

2005-01-01

The use of resealed red blood cell membranes (ghosts) allows the study of the transport of a compound in a nonmetabolizing system with a biological membrane. Transmembrane movements of anandamide (N-arachidonoylethanolamine, arachidonoylethanolamide) have been studied by exchange efflux experiments...... at 0°C and pH 7.3 with albumin-free and albumin-filled human red blood cell ghosts. The efflux kinetics is biexponential and is analyzed in terms of compartment models. The distribution of anandamide on the membrane inner to outer leaflet pools is determined to be 0.275 ± 0.023, and the rate constant...... of unidirectional flux from inside to outside is 0.361 ± 0.023 s. The rate constant of unidirectional flux from the membrane to BSA in the medium ([BSA]) increases with the square root of [BSA] in accordance with the theory of an unstirred layer around ghosts. Anandamide passed through the red blood cell membrane...

Fc receptors for mouse IgG1 on human monocytes: polymorphism and role in antibody-induced T cell proliferation.

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Tax, W J; Hermes, F F; Willems, R W; Capel, P J; Koene, R A

1984-09-01

In previous studies, it was shown that there is polymorphism in the mitogenic effect of mouse IgG1 monoclonal antibodies against the T3 antigen of human T cells. This polymorphism implies that IgG1 anti-T3 antibodies are not mitogenic for T cells from 30% of healthy individuals. The present results demonstrate that this polymorphism is caused by polymorphism of an Fc receptor for mouse IgG1, present on human monocytes. The Fc receptor for murine IgG1 could be detected by a newly developed rosetting assay on monocytes from all individuals responsive to the mitogenic effect of IgG1 anti-T3 antibodies. This Fc receptor was not detectable on monocytes from those individuals exhibiting no mitogenic responses to IgG1 anti-T3 monoclonal antibodies. Cross-linking of T3 antigens appears to be essential for antibody-induced mitosis of T cells, because mononuclear cells that did not proliferate in response to WT 31 (an IgG1 antibody against T3 antigen) showed a proliferative response to Sepharose beads coated with WT 31. The Fc receptor--if functionally present--may be involved in the cross-linking of T3 antigens through anti-T3 antibodies. Further evidence for the involvement of this Fc receptor in antibody-induced T cell proliferation was provided by inhibition studies. Immune complexes containing IgG1 antibodies were able to inhibit the proliferative response to IgG1 anti-T3 antibodies. This inhibition by immune complexes appears to be mediated through the monocyte Fc receptor for mouse IgG1. These findings are important for the interpretation of previously described inhibitory effects of anti-T cell monoclonal antibodies on T cell proliferation, and show that such inhibitory effects may be monocyte-mediated (via immune complexes) rather than caused by a direct involvement of the respective T cell antigens in T cell mitosis. The Fc receptor for mouse IgG1 plays a role in antibody-induced T cell proliferation. Its polymorphism may have important implications for the

Nonreassuring fetal heart rate patterns and nucleated red blood cells in term neonates.

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Kovalak, E Ebru; Dede, F Suat; Gelisen, Orhan; Dede, Hulya; Haberal, Ali

2011-05-01

The aim of this study was to evaluate the association between nonreassuring fetal heart rate patterns during labor and umbilical cord nucleated red blood cell counts. Nucleated red blood cell data was collected prospectively from 41 singleton term neonates presented with nonreassuring fetal heart rate patterns and/or meconium stained amniotic fluid during labor (study group) and from 45 term neonates without any evidence of nonreassuring fetal status (controls). Umbilical artery pH, blood gases and base excess were also determined to investigate the correlation between independent variables. The median nucleated red blood cells per 100 white blood cells were 13 (range 0-37) in the study group and 8 (range 0-21) in the control group. Stepwise regression analysis have identified meconium stained amniotic fluid (R(2) = 0.15, p patterns. Nucleated red blood cells in the cord blood of newborns were found to be elevated in patients with nonreassuring FHR patterns during labor. However, the wide range and the poor correlation of NRBC count with umbilical artery pH and blood gas values limit its clinical utility as a marker for fetal hypoxia.

Effect of producer cell line on functional activity of anti-D monoclonal antibodies destined for prevention of rhesus sensitization.

Science.gov (United States)

Olovnikova, N I; Ershler, M A; Belkina, E V; Nikolaeva, T L; Miterev, G Yu

2009-04-01

The ability of anti-D antibodies to cause antigen-specific immunosuppression depends on their interaction with low-affinity Fcgamma-receptors. Human monoclonal antibodies to D antigen of the rhesus system were investigated by antibody-dependent cytotoxicity assay in order to estimate their ability to induce hemolysis mediated by low-affinity Fcgamma receptors. We demonstrate that affinity of monoclonal antibodies to receptors of this type does not depend on primary structure of Fc-fragment, but depends on the producer cell line which expresses the antibodies. Monoclonal IgG1 antibodies interacting with FcgammaRIIa and FcgammaRIII lost this property, if they were secreted by human-mouse heterohybridoma, but not by human B-cell line. On the opposite, monoclonal antibodies that could not activate low-affinity Fcgamma receptors were highly active after human cells fusion with rat myeloma YB2/0. Hemolytic activity of IgG3 remained unchanged after fusion of human cells with rodent cells.

A novel monoclonal antibody of human stem cell factor inhibits umbilical cord blood stem cell ex vivo expansion