Heat treatment of normal human sera reveals antibodies to bactericidal permeability-inducing protein (BPI)
| Autor: | Brownlee Aa, C. M. Lockwood |
|---|---|
| Rok vydání: | 1999 |
| Předmět: |
Hot Temperature
Recombinant Fusion Proteins Immunology Enzyme-Linked Immunosorbent Assay Chromatography Affinity Epitope Immunoglobulin G Antibodies Antineutrophil Cytoplasmic Polyethylene Glycols Epitopes Antigen Antibody Specificity Reference Values Proteinase 3 Humans Immunology and Allergy biology Chemistry Membrane Proteins Blood Proteins Hydrogen-Ion Concentration Fusion protein Blood Preservation Myeloperoxidase biology.protein Original Article Protein G Antibody Antimicrobial Cationic Peptides |
| Zdroj: | Clinical and Experimental Immunology. 117:183-189 |
| ISSN: | 1365-2249 0009-9104 |
| Popis: | SUMMARYHeat treatment of normal sera to 56°C for 30 min, a common procedure for the inactivation of viruses, e.g. HIV, reveals the presence of antibodies to neutrophil cytoplasm antigens (ANCA), as detected by indirect immunofluorescence on ethanol-fixed human neutrophils and by antigen-specific ELISA for BPI. Reactivity was not seen to the other common vasculitis-associated antigens proteinase 3 (PR3) or myeloperoxidase (MPO). The effect of temperature was maximal at 56°C, with substantial antibody demonstrable after only 5 min at this temperature. In experiments using polyethylene glycol (PEG)6000 to remove immune complexes, the effect of heating could be abrogated by preincubation with 8% PEG, which suggested that these anti BPI antibodies might be complexed in sera. After passage of normal plasma over a protein G column, the acid-eluted fraction contained elevated levels of antibodies to BPI but not to other vasculitis-associated antigens such as PR3 or MPO, nor to glomerular basement membrane (GBM), the Goodpasture antigen which is recognized by the pathogenically important human antibodies shown to mediate nephritis in transfer experiments. Moreover the levels of anti-BPI in the IgG fraction could be augmented by preincubation with glycine pH 2.5 for 30 min. This anti-BPI activity could be inhibited by addition of the unbound material from the protein G column and this inhibitory material was not heat-labile at 56°C. The molecular specificity of this autoreactivity was confirmed using recombinant BPI in coincubation experiments and the epitope localized to the C or N terminal moieties by the use of recombinant fusion proteins. |
| Databáze: | OpenAIRE |
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