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2.
Delayed type hypersensitivity (DTH) to bovine serum albumin (BSA) and to lipid-conjugated BSA were studied comparatively. Unlike the case of BSA with which no DTH can be detected with native antigen, injection of butyric-conjugated BSA (Bu-BSA) in sensitized mice provokes a typical DTH for an early and limited period. Alum-precipitated Bu-BSA (Al-Bu-BSA) provokes from the beginning a stronger DTH which persists a much longer period. 相似文献
3.
Monoclonal antibodies were developed to neutrophil surface antigens in an effort to identify neutrophil subpopulations. Eleven
monoclonals were positive for neutrophils using the ELISA and flow cytometry. Four monoclonals recognised neutrophils and
monocytes, but more importantly each recognised subpopulations of neutrophils. Percentage binding of the same population of
neutrophils by each monoclonal was 19H7, 23%; 36H10, 34%; 46B3, 16%; 9D13, 37%. All four showed additive percentage binding
to neutrophils when reacted with each other. Cow differences in percentage binding of monoclonals were also evident, i.e.,
46B3 — cow no. 8437, 8%; cow no. 2820, 22%. Monocytes showed the most variation, both by cow and by clone within cow, i.e.,
cow no. 8813, 46B3 = 98% and 9D13 = 6%; clone 19H7, cow no. 8813 = 88% and cow no. 8679 = 2%. Percentage binding to eosinophils
was low for all clones except 46B3 which averaged 46%, 5%, 72% and 76% for four cows tested. Percentage binding to lymphocytes
tended to be high for all clones across all cows, averaging 62%. Clone 36H10 tended to be the lowest of the clones, averaging
38%. The ability of these clones to identify subpopulations of neutrophils and the significant among cow variation make them
useful tools for studying the relationship of different cell-surface antigens to cell function within cow and among cows. 相似文献
4.
Rotavirus-specific antibodies were detected in fetal bovine serum, bovine serum albumin, and human serum albumin by radioimmunoprecipitation with the NCDV strain of bovine rotavirus as the detecting antigen. Fetal bovine sera neutralized bovine rotavirus in a plaque reduction neutralization test to titers of 1:20 or greater. Immunoglobulins purified from fetal bovine serum by protein A-agarose affinity chromatography precipitated rotavirus antigens but did not neutralize bovine rotavirus. Rotavirus antibodies in fetal bovine serum and in purified serum albumin preparations may interfere with diagnostic assays for the detection of rotavirus antigens or antibodies. 相似文献
5.
The aim of this study was to investigate the recognition pattern of bovine serum albumin (BSA), a major dietary protein by serum IgG and IgA antibodies. Anti-BSA IgG and IgA antibodies were measured by ELISA technique in 3 different cohorts: 578 unselected persons, 84 new-onset insulin-dependent diabetes mellitus (IDDM) patients and 103 atopic persons. In order to characterize the recognition pattern of the different BSA domains, recombinant BSA and recombinant fragments covering the 3 BSA domains were produced. BSA digestion was monitored in simulated gastric fluid experiments by means of domain specific monoclonal antibodies. IgG and IgA antibody titres to native BSA were highest in IDDM patients. The three major BSA domains were equally well recognized by IgG antibodies of the three cohorts. Interestingly all three study groups showed a dissociation of their IgG and IgA antibody response to the first BSA domain. The ratio of IgG to IgA antibodies recognizing this domain was 93%/42% in controls, 92%/37% in IDDM patients and 80%/47% in atopic persons. In simulated gastric fluid experiments, the first BSA domain was the first to become undetectable to specific monoclonal antibodies during digestion. In conclusion humoral IgG and IgA antibodies recognize the major BSA domains with different frequencies. The N-terminal domain of BSA, the first to be degraded during simulated gastric digestion is less well recognized by IgA antibodies. This suggests that early digestion is negatively correlated to the IgA antibody response and that the IgA response associated to the gut associated lymphoid tissue (GALT) and the systemic IgG antibody responses are independent. 相似文献
6.
Of 89 monoclonal antibodies screened for anti-class I activity in a cytotoxic assay against bovine peripheral-blood lymphocytes, 6 reacted with all lymphocytes from all cattle tested, 72 failed to react at all and 11 reacted with polymorphic determinants. The reactivity of some of the 11 polymorphic monoclonal antibodies was dependent upon the bovine major histocompatibility system (BoLA) class I type. Eight monoclonal antibodies selected for putative anti-class II activity reacted with B-enriched lymphocytes from all cattle tested. 相似文献
7.
To define the characteristics and target antigens (Ags) of nephrotoxic antibodies (Abs) and to analyze the factors that govern the evolution of Ab-mediated glomerular injury, we have prepared monoclonal Abs against rat glomerular Ags. BALB/c mice were immunized with Lewis rat cortex or glomeruli, and their spleens were removed and fused with hypoxanthine-aminopterin-thymidine supplement-sensitive myelomas. Hybrids were selected for production of Abs against Lewis rat kidney by indirect immunofluorescence. To date, more than 50 positive hybrids have been selected and their tissue reactivity defined by indirect immunofluorescence and immunoelectron microscopy. Of these, 14 are presented here in detail. One of these monoclonal Abs, K9/4, recognizes a unique Ag present exclusively on the cell surface of rat glomerular visceral epithelial cells. Four Abs (K12/2, K17/4, K12/5, and K12/8) recognize sites within the glomerular basement membrane; K12/2 and K17/4 also bind to vascular basement membranes of the rat, whereas K12/5 and K12/8 bind to glomerular basement membrane, tubular basement membranes, and vascular and epithelial basement membranes in all tissues of the rat. Two hybridomas (K6/1 and K6/3) recognize determinant(s) present on cell surfaces of endothelial and epithelial cells as well as within the glomerular basement membrane. All of these previously mentioned Abs are species restricted (i.e., they bind only to rat tissue) and, with the exception of K9/4, bind upon in vivo administration. Several others, however, recognize ubiquitous Ags that are present on intracellular structures in every species tested. The tissue distribution of these Ags suggests that they are present in contractile or cytoskeletal elements and, as expected from their intracellular location, monoclonal Abs directed against these components do not bind upon in vivo administration. Future studies will be directed at defining the antigenic composition of the glomerular capillary wall and the relevance of such Ags in immune-mediated glomerular injury. 相似文献
8.
Twenty-nine stable hybridoma cell lines secreting monoclonal antibodies to bovine somatotropin (bST) have been produced and characterized. Five of the monoclonal antibodies bind porcine and human somatotropins as well as bST. One of these antibodies was used as a reagent in immunoadsorbent chromatography of recombinant bST or pituitary bST from cell extracts. Following chromatography, the bST preparations retained activity in a rabbit liver radioreceptor assay and in a radioimmunoassay. The immunoadsorbent reagent bound human and porcine somatotropins as well as bovine. 相似文献
9.
Eosinophilic gastroenteritis is a disease characterized histologically by an eosinophilic infiltration of the gut. The cause of this disease remains unclear, although both food allergy and food intolerance have been implicated in its pathogenesis. We report the case of a 22-year-old man in whom gastrointestinal symptoms first appeared in childhood, with involvement of mucosa and muscularis layers of stomach and bowel. He presented high IgE blood levels, and his prick test was positive to bovine, pig, and lamb sera. Immunoblots from calf, pig, and lamb sera, incubated with the patient's serum and revealed by autoradiography, demonstrated the presence of a 65-kDa protein band that was recognized by IgE antibodies but not by IgG. This band corresponded to bovine serum albumin, while IgE did not show reactivity with human albumin. These data suggest a possible role for IgE-mediated hypersensitivity mechanisms in the pathogenesis of eosinophilic gastroenteritis. 相似文献
10.
Two IgG1/kappa class monoclonal antibodies specific for human immune interferon (IFN-gamma), designated B1 and B3, were developed. Specific binding of both monoclonal antibodies to natural or Escherichia coli-derived recombinant human IFN-gamma was demonstrated in a solid-phase radioimmunoassay or by immunoprecipitation. Antibody B3 showed potent neutralizing activity against both natural and recombinant IFN-gamma. Antibody B1, which showed neutralizing activity only when very high concentrations were employed, was used for preparing immunosorbents for affinity chromatography of IFN-gamma. When a highly purified preparation of 125I-labeled natural IFN-gamma was loaded onto the affinity column, all of the biological activity was retained on the column. The bulk of 125I-labeled IFN-gamma bound to the affinity column be eluted in biologically active form, suggesting that antibody B1 could be used for the purification of human IFN-gamma. Analysis of IFN-gamma eluted from the column by NaDodSO4/polyacrylamide gel electrophoresis (SDS-PAGE) indicated that both of the known molecular weight subspecies of IFN-gamma (25,000 and 20,000 MW), as well as the presumed dimer of 45,000 MW, were retained by the B1 antibody affinity column. 相似文献
11.
Recently, this laboratory reported an autoreactivity of myoglobin (Mb)3 antisera with the Mb of the species in which the antisera are raised. Also, animals injected with autologous Mb mounted an autoimmune antibody and T-lymphocyte proliferative response against this protein. This posed the possibility that autoimmune recognition might be a general phenomenon not confined only to sequestered proteins such as Mb. Using RSA, we have demonstrated unambiguously that RSA cross-reacted with rabbit antisera to bovine (BSA) or to human (HSA) albumin. In exchange experiments, 125I-labelled RSA was bound by the IgG in the immune complexes isolated from rabbit antisera to BSA or HSA. Also, 125I-antibodies were bound by RSA-adsorbents. Both binding activities were inhibited specifically by RSA. RSA was isolated from three rabbits and each rabbit was immunized with its own RSA. In each rabbit autoantibodies were found by exchange between immune complexes and the rabbit's own [ 125I]-RSA. Also, 125I-antibodies from each rabbit were bound by adsorbents of the rabbit's own RSA. Inhibition studies, data on preimmune sera and on antisera against other proteins confirmed the specificity of the binding. The findings confirm the universality of autoimmune recognition and lend support to our previous suggestion that antigenic sites are ‘structurally-inherent’ in the protein. 相似文献
12.
Monoclonal antibodies to MDP were prepared by hybridization of NSO myeloma cells with spleen cells of BALB/c mice immunized with MDP conjugated to methyl-BSA. Hybridomas secreting anti-MDP antibodies were selected by the binding activity of their supernates to MDP-A-L using a radioimmunoassay. After cloning in soft agar, the specificities of monoclonal anti-MDP antibodies were assayed by an inhibition of ELISA with various derivatives of MDP. Fine structural analysis of specificity for one such clone (2–4) is reported. This antibody recognizes the N-acetyl-muramic acid ( N-AcMur) linked to the dipeptide but not N-Ac-Mur or/and dipeptide alone. The N-Ac group on muramic acid is an important antigenic determinant and the glycopeptide linkage seems to be crucial in presenting the sugar moiety. Conservative substitution of l-Ala (i.e. by l-Ser or l-Val) had no effect on the binding ability to the antibody whereas a radical change i.e. replacement of l-Ala by l-Pro or completely abolished the antigenicity of the molecule. There was no clear correlation between biological activities of various derivatives of MDP and their ability to react with this antibody. Some possible hypotheses explaining this lack of correlation are presented. 相似文献
13.
Mouse monoclonal antibodies directed against nerve growth factor (beta NGF) from bovine seminal plasma have been isolated and characterized. They are produced by hybridomas derived from Sp2/0.Ag14 myeloma cells and spleen cells from BALB/c mice immunized with beta NGF which was purified by the method of Harper et al. [J. biol. Chem. 257, 8541-8548 (1982)]. Five of these hybridomas can be grown in ascites tumor form and secrete antibodies of the IgG1 or IgG2a subclass. When used to probe the components of seminal plasma extracts or purified beta NGF as separated electrophoretically on SDS gels, the antibodies react with the beta NGF band at Mr = 15,000. The antibodies bind to native bovine beta NGF, but bind very poorly to mouse beta NGF. Antibody exclusion and additive-binding experiments indicate that these antibodies bind to the 1 antigenic domain. The cell receptor binding site is probably not close to this domain, as the antibodies fail to block the biological activity of bovine beta NGF on cultures of dissociated neurons from sensory ganglia. These monoclonal antibodies define a region in which bovine beta NGF is structurally different from the closely related molecule mouse beta NGF. 相似文献
15.
An extract of dry‐roasted commercial peanut mix (CPE) was examined for allergenic activity in peanut‐sensitive individuals, using skin tests and radioallergosorbent (RAST) assays. Proteins in the extract were characterized by sodium dodecyl sulfate polyacry‐lamide gel electrophoresis (SDS‐PAGE) and immunoblotting. The proteins were electro‐eluted in three fractions in the ranges 15–25, 26–58 and 65 kDa. The 15–25 kDa molecular weight fraction produced the most reactive skin tests in peanut‐sensitive subjects and was chosen for monoclonal antibody production. Six hybridoma cell lines secreting peanut‐specific antibodies of the IgM isotype (kappa light chain) were produced. Immunopurified CPE proteins were then subjected to SDS‐PAGE, resulting in five major bands with approximate molecular weights of 14, 25, 38, 40 and 44 kDa. Immunoblotting of these separated proteins revealed: (1) three bands with approximate molecular weights of 38, 44 and 65 kDa, which bound IgE from peanut‐sensitive patients; and (2) that the monoclonal antibodies recognized epitopes in bands at approximate molecular weights of 12, 14, 23 and 25 kDa. RAST inhibition assays showed that the affinity‐purified proteins were able to inhibit the binding of serum IgE from peanut‐allergic individuals to solid‐phase CPE. 相似文献
16.
Sera from patients undergoing hemodialysis with formaldehyde (F)-sterilized dialyzers were studied to determine if antibodies against F conjugated to human serum albumin (HSA) could be detected. F-human serum albumin (F-HSA) conjugates were prepared using ratios of F to HSA that did not precipitate the HSA. The F-HSA conjugates migrate differently electrophoretically than HSA with an increased negative charge of F-HSA as compared with HSA. The F-HSA was used in an ELISA. The results demonstrated that in certain sera, IgG, IgM, IgA and IgE antibodies against F-HSA could be measured. In the highest titered sera, it was shown that the IgG antibody was not directed against F alone or F-lysine but against an antigenic grouping of F-HSA. No correlation of either IgG or IgE antibodies with immune complex or allergic reactions was found in this series of dialysis patients. Some sera from dialysis patients had antibody activity against HSA. Sera from 2 physicians with rhinitis after F exposure had no antibody activity against F-HSA or HSA. Two nurses with a history of F-induced asthma had no IgG antibodies but did have IgE antibodies against F-HSA and HSA. This spectrum of immunologic responses is analogous to responses in dogs immunized with F or F dog albumin. We have not been able to identify anti HSA antibodies in patients reactive to other hapten-HSA compounds and it is suggested that anti HSA antibodies in F-exposed humans may relate to the F exposure. 相似文献
17.
Thirty-one stabile murine monoclonal antibody (MAb) producing cell lines to progesterone were generated by using a short and a long immunization protocol. Long-term immunization with high doses of 11alpha-hydroxyprogesterone-hemisuccinate-bovine serum albumin (11alpha-OH-P-HS-BSA) antigen led to very good antibody response in Balb/c mice. The donor mouse produced antiserum with a high titre of 1/250,000. Eleven MAbs were selected for further characterization since they showed high sensitivities (<35 pg/well to inhibit 50% of the tracer) in bridge homologous enzyme immunoassay (EIA). The results were compared to the donor mouse polyclonal antiserum. The MAbs and the donor mouse antiserum were generally found to be highly specific, when tested with 30 different steroids. Employing MAb 9C11, with affinity constant, K(alpha), to 11alpha-OH-P-HS of 1.1 x 10(10) M(-1), a bridge heterologous microtitre plate EIA for milk progesterone was developed, using the second-antibody coating technique and horseradish peroxidase (HRP) as an enzyme label. The assay is simple and convenient to use, as it permits direct addition of undiluted milk samples, at the same time maintaining high sensitivity, high precision, and a wide range of optical density (OD) values. The major advantage of the assay developed, compared to previously published direct addition milk progesterone immunoassays, is that progesterone concentrations, measured by the EIA, were not influenced by changing milk fat concentrations, even when milk samples containing up to 10% of milk fat were used for analysis. 相似文献
18.
Balb/c mice were immunized with albumin purified from sailfish (Istiophorus albicans) serum. Hybridomas were produced and screened by ELISA for reactivity with the purified albumins of sailfish, blue marlin (Makaira nigricans) and white marlin (Tetrapturus albidus). Monoclonal antibodies (MAbs) from 16 different clones exhibited activity against sailfish albumin. Thirteen of the MAbs showed cross-reactivity with the marlin species. Three MAbs exhibited distinct specificity for sailfish albumin. One of these species specific MAbs (M2D1) was conjugated to horseradish peroxidase (HRP) in order to construct an ELISA for identification of sailfish from serum. The ELISA for sailfish correctly identified eight sailfish from 26 billfish serum samples. The MAb-peroxidase conjugate was highly specific toward sailfish in that no reaction against heterologous species was detected. 相似文献
19.
The importance of tryptophan in the combining sites of anti-dinitrophenyl antibodies is evaluated from a series of model studies, and from experiments with a Dnp-binding monoclonal antibody protein A3. The thermodynamic parameters characterising the formation of a Dnp/tryptophan complex are determined. It is concluded that a Dnp/tryptophan interaction could provide about a third of the binding energy of Dnp ligands in such proteins as the mouse IgA myeloma 315. 相似文献
20.
Cross-reactions between bovine serum albumin and equine serum albumin were studied using heterologous soluble complexes and specifically purified cross-reacting antibody. Experiments with soluble complexes showed that homologous antigen can displace heterologous antigen specifically bound to antibody but heterologous antigen cannot displace homologous antigen. On gel precipitation tests a specific precipitation resulted when heterologous soluble complex reacted with homologous antigen. By using equine serum albumin conjugated to polyaminopolystyrene the cross-reacting antibodies from anti-bovine serum albumin imune sera could be isolated. These are divalent 7S, γ-globulin antibodies. A figure of cross-reaction was obtained when these purified antibodies were tested by double diffusion in agar with bovine and equine serum albumins. The results obtained both with soluble complexes and with purified antibody support the view that cross-reacting antibody is more avid for the homologous than for the heterologous antigen. 相似文献
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