Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0393754 (HSA)
 2,996 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Studies concerning the localization of immune complexes in lymphoid follicles and the involvement of these trapped immune complexes in the regulation of the immune response have thus far been performed with poorly defined complexes in terms of size and composition. For that reason, the minimum requirements for trapping in terms of number of antigen- and antibody molecules present in immune complexes could not be determined. We here describe the production and in vivo use of a monomeric HSA-HRP antigen-enzyme conjugate, readily demonstrable in cryostat sections and ELISA. This conjugate was obtained by combining the glutaraldehyde coupling-method with chromatography to fractionate monomeric and multimeric constituents. SDS-PAGE analysis showed that the conjugate consisted of a single molecular species of 109 kDa, whereas the often used periodate oxidation coupling method yielded a heterogeneous population of multimeric, oligomeric and monomeric molecules. We investigated the minimal size requirements for the composition of immune complexes to be trapped in murine spleen follicles using three different conjugates (monomeric HSA-HRP, multimeric HSA-HRP and multimeric HSA-HRP-Penicillin) and a panel of anti-HSA and anti-Penicillin monoclonal antibodies. We demonstrate that the smallest immune complexes, consisting of one antibody and two conjugate molecules, do not localize in splenic follicles. Immune complexes prepared with a single monoclonal antibody localize in follicles only if the epitope recognized occurs repeatedly on the antigen. The relevance of these results for physiological follicular trapping of protein antigens is discussed.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Production of monomeric antigen-enzyme conjugate to study requirements for follicular immune complex trapping. 137 27

SDS-PAGE showed that human salivary alpha-amylase family A (HSA-A) was converted to family B (HSA-B) in human saliva. This conversion did not occur in the supernatant of saliva which had been centrifuged at 105,000 x g for 60 min. An enzyme which catalyzed the conversion existed in the insoluble fraction of human saliva. The enzyme was solubilized with nonionic or zwitterionic detergents, and showed the maximum activity around pH 6. It was stable between pH 4 and 10, and at a temperature lower than 40 degrees C. The enzyme reduced the molecular weight of HSA-A (62,000) to the same molecular weight (58,000) as that of HSA-B without forming any intermediate. It also changed the PAGE pattern of multiple forms of HSA-A to the same pattern as that of HSA-B. It was not inhibited by protease inhibitors, and it did not destroy the reactivity of HSA-A with anti-human salivary alpha-amylase antiserum. The enzyme diminished the reactivity of HSA-A with concanavalin A. These results indicate that HSA-A was converted to HSA-B through the release of sugar chains by the action of the enzyme in the insoluble fraction of human saliva.
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PMID:Evidence for conversion of human salivary alpha-amylase family A to family B by an enzyme action. 142 13

The thermal stability of IL-1 beta in aqueous solution as a function of temperature (5-60 degrees C), pH (2-9), buffer (acetate, citrate, tris, and phosphate), and cyroprotectants (sugars, HSA) was investigated in this study. The analytical methodologies included RP-HPLC, SEC, ELISA, IEF-PAGE, SDS-PAGE, and bioassay. The degradation and inactivation of IL-1 beta at or above 39 degrees C were attributed to autoxidation of the two cysteine residues in the denatured protein, followed by hydrophobic/covalent aggregation and precipitation. At or below 30 degrees C, IEF- and SDS-PAGE results suggest a possible deamidation reaction. The difference in mechanism of degradation precludes the prediction of formulation shelf life from accelerated temperature data. Nonetheless, the good stability observed at 5 degrees C suggests that a solution formulation may be feasible for IL-1 beta.
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PMID:Stability of interleukin 1 beta (IL-1 beta) in aqueous solution: analytical methods, kinetics, products, and solution formulation implications. 187 Oct 44

Coelomic fluids of the two earthworm species E.foetida (E.F.) and L.terrestris (L.T.) have not only the ability to lyse various vertebrate erythrocytes but also to digest vertebrate serum proteins. Both activities are carried by different molecules since hemolysis but not proteolysis was inhibited by simple sugars. In contrary, proteolysis was blocked by PMSF which did not influence hemolysis. Coelomic fluids of E.F. digest effectively vertebrate serum proteins (PIgG, HSA) but not the proteins of L.T. coelomic fluids. The proteolytic activity was detected in approximately 40 000 mol. wt. fraction. After digestion proteolytic fragments were analyzed by immunoelectrophoresis, SDS-PAGE and TCA precipitation. Two of the fragments reacting with PIgG antisera remained intact even after 120 h digestion.
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PMID:Lytic activities in coelomic fluid of Eisenia foetida and Lumbricus terrestris. 352 2

Glucosylated human serum albumin (G-HSA) obtained under incubation with glucose at 37 degrees C for 8 days showed a new fluorescence with a maximum at 430 nm, resulting in quenching of the fluorescence of only one tryptophan residue on HSA. The quantum yield of new fluorescence is 0.024 at 25 degrees C. The analysis of the excitation spectra allowed us to conclude the absence of energy transfer. In G-HSA, non-disulfide cross-linking hexamer was confirmed by SDS-PAGE.
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PMID:New fluorescence of nonenzymatically glucosylated human serum albumin. 648 18

A possible change in the nuclear stability of the human spermatozoa further than ejaculation was investigated. A nuclear chromatin decondensation ability test using 1% SDS + 6 mM EDTA was used on spermatozoa migrated for 1 h in a swim-up migration (in BWW + human serum albumin 0.8%) and capacitated for 5 h in the same medium. The results, analyzed as paired series, showed that (1) capacitated and migrated spermatozoa have a greater nuclear stability than that of the control population (total sperm), (2) there was no significant difference of the nuclear stability between migrated and capacitated spermatozoa, and (3) there was no effect of the media used (BWW + HSA) on the nuclear stability. Thus, it seemed that migrating spermatozoa definitely selects a specific resistant population to decondensing reagents.
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PMID:Effect of migration and capacitation on the nuclear stability of human sperm. 653 42

A technique for isolating and analyzing immune complexes (IC) using rheumatoid factor (RF) as immunoadsorbent is described. It contains several modifications to a previously published original method and has given greatly improved results: a small amount of BSA-anti-BSA (aBSA) model IC was added to human serum and the mixture was filtered through a column of Sephacryl S-300, the large molecular weight fraction was concentrated and added to tubes coated with RF. The bound IC were eluted, radioiodinated and freed to the major contaminants (HSA and immunoglobulins) by adding the corresponding antisera and removing the resulting complexes by coprecipitation or with the aid of protein A-containing Staphylococci. The purified preparations were analyzed by SDS-polyacrylamide gel electrophoresis (PAGE) and autoradiography. BSA was clearly identified in the autoradiograms. An example is given of the application of this technique to the isolation and analysis of IC from an abdominal effusion obtained from a patient with ovarian cancer.
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PMID:An improved technique for the isolation and analysis of immune complexes. 719 13

Covalently cross-linked immune complexes were prepared with multivalent antigens, obtained by coupling varying numbers of 4-azido-4-nitrophenyl groups (NAP) on human serum albumin as the carrier molecule (NAPn . HSA). In this system the haptenic group served to bind the antigen to the antibody (antibodies to NAP) and to form covalent bonds upon photoactivation. The covalently cross-linked immune complexes contained around 30% of antibodies that were dissociable from complexes by SDS polyacrylamide gel electrophoresis. A comparable portion of antibody-combining sites were accessible to the free hapten (NAP . lysine) in molar excess by equilibrium dialysis. The stable, covalently cross-linked complexes with NAP7.0 . HSA and NAP12.9 . HSA were prepared and separated into complexes with varying degrees of lattice by sequential steps of gel filtration. Ag1Ab1 complexes were obtained with reasonable homogeneity. Other preparations contained successively higher lattices but were not homogeneous. When these complexes were injected into mice, the increasing lattice of complexes resulted in increasingly rapid removal of the complexes from the circulation. The antigen, independent of lattice, also contributed to removal of complexes from circulation. NAP12.9 . HSA alone was removed from circulation faster than NAP7.0 . HSA, and Ag1Ab1 complexes with NAP12.9 . HSA were removed faster than Ag1Ab1 complexes with NAP7.0 . HSA. The studied system adds covalently cross-linked immune complexes with multivalent antigens to the armamentarium of covalently cross-linked complexes that previously were obtained only with bivalent affinity labels.
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PMID:Covalently cross-linked immune complexes prepared with multivalent cross-linking antigens. 729 22

The soluble granule chymase, rat mast cell protease-II (RMCP-II), is abundantly expressed in intestinal mucosal mast cells (MMC) but its function is not known. One hypothesis is that RMCP-II degrades the epithelial basement membrane and promotes the loss of enterocytes typically associated with type I hypersensitivity reactions in the rat. To test this hypothesis more directly, ex vivo perfusion of the cranial mesenteric artery and jejunal lumen was used to monitor the anaphylactic release of RMCP-II and its effects on mucosal permeability and epithelial integrity. Within 2 min of intravascular challenge with soluble adult Nippostrongylus brasiliensis worm antigen there was a 1,000-fold (P < 0.02) increase in the concentration of RMCP-II in the vascular perfusate from the jejunum of Nippostrongylus-sensitized rats but not the controls. Similarly, translocation of RMCP-II into the gut lumen increased 10-fold (P < 0.02) after 2 min only in worm antigen-challenged immune rats. Using an identical protocol, but incorporating Evans blue-labeled human serum albumin (EB-HSA) in the vascular perfusate, the timing of the release of RMCP-II into the two compartments was very similar to the first experiment and furthermore the translocation of EB-HSA increased 18-fold (P < 0.05) after 4 min in sensitized rats challenged with worm antigen. To examine the effects of RMCP-II more directly 1 mg of the highly purified chymase was introduced into the cranial mesenteric artery in ex vivo perfused normal rats. A significant (P < 0.05) 70-fold increase in concentration of RMCP-II in jejunal perfusate occurred after 6 min. In a repeat dose-response experiment, infusion of 0.375, 0.75, or 1.5 mg of RMCP-II, together with EB-HSA, established that the cumulative amounts of RMCP-II and EB-HSA translocated from the vasculature to the gut lumen in each perfusion (during the 10-min period of RMCP-II infusion) were significantly correlated. Analysis of intestinal perfusates by SDS-PAGE and by Western blotting using monoclonal anti-RMCP-II antibody confirmed that there was a concomitant translocation of both the protease and EB-HSA into the gut lumen. Histological evaluation of the mucosa failed to reveal any significant morphological change in any of the experiments. The rapid development of macromolecular leak, its association with the translocation of RMCP-II, and the absence of gross epithelial lesions, suggest for the first time that a mast cell granule chymase increases epithelial permeability via a paracellular route and implies that the substrate may be a protein, or proteins, in the epithelial junctional complex.
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PMID:Release of the mucosal mast cell granule chymase, rat mast cell protease-II, during anaphylaxis is associated with the rapid development of paracellular permeability to macromolecules in rat jejunum. 750 33

A three stage method for the ultrapurification of polyclonal IgE from human serum is reported using anion exchange chromatography followed by monoclonal antibody based positive and negative affinity chromatography. Following dialysis of 25-100 ml of serum (2.3-14 micrograms IgE/ml, n = 4) against 0.05 M Tris pH 8, each specimen was subjected to diethylaminoethyl (DEAE)-cellulose chromatography (serum/matrix = 1/4). IgE was eluted with 0.05 M Tris, 0.05 M NaCl pH 8, yielding an IgE recovery of 61-93%, with removal of approximately 90% of other serum proteins and an IgE purity ([IgE]/[Igs]) of 0.1-1.1%. After adjusting to 0.1 M NaCl and concentrating approximately 30-fold, the eluted IgE was further purified by affinity chromatography using a panel of IUIS/WHO-documented mouse monoclonal anti-human immunoglobulin antibodies (alpha hIg-MAbs). First, the IgE-enriched DEAE-cellulose chromatography fraction was incubated in a batch mode with two alpha hIgE-Fc MAbs (HP6029, HP6061) coupled to CNBr-Sepharose, CL-4B. IgE was eluted with 0.05 M glycine pH 2.8 and immediately neutralized. The IgE recovery was 32-52% and IgE purity was 72-97%. Silver-stained SDS-PAGE and noncompetitive solid-phase two-site immunoenzymetric assays for total human IgA, IgE, IgG and IgM indicated that IgA, IgG and IgM were the only contaminants. Next, the IgE was concentrated 10-30-fold in the presence of 0.1% HSA. One IgE specimen was ultrapurified in a batch mode by negative selection chromatography using three pairs of alpha hIg-MAbs (alpha hIgA: HP6111 + HP6123; alpha hIgG: HP6017 + HP6046; alpha hIgM: HP6081 + HP6083) coupled to CNBr-Sepharose, CL-4B. IgE purity increased from 91% to > 99.9% with approximately 70% recovery of IgE for this step. The ultrapurified IgE antibody was shown to be functionally reactive for allergen and Fc epsilon RI receptors on human basophils. We conclude that alpha hIg-MAbs are powerful tools to facilitate the affinity purification of functionally active human IgE from serum; however, when the analyte is present in low concentration, a carrier protein needs to be added to minimize non-specific loss of the material during this process.
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PMID:Purification of immunoglobulin E (IgE) antibodies from sera with high IgE titers. 787 65


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