An avidin-biotin ELISA assay for the measurement of de novo human IgE synthesis in culture supernatants.
A very sensitive (100 pg/ml) solid-phase enzyme immunoassay (ELISA) for the determination of human IgE has been developed. This assay incorporates the avidin-biotin system to increase sensitivity and can detect as little as 100 pg/ml (10 pg/test) of human IgE. The assay is highly specific and allows quantitative determination of human IgE in supernatants of peripheral blood lymphocytes as well as in serum. The very high sensitivity of the assay was accomplished by optimizing concentrations of the following reagents: (1) affinity-purified rabbit anti-human IgE coating antibodies; (2) biotin-conjugated goat anti-human IgE; (3) avidin-horseradish peroxidase (HRP) conjugate. In summary, the assay described is rapid (6 h), reproducible, isotype specific, and has the sensitivity of radioimmunoassays usually employed for the quantification of IgE. This assay may be utilized in establishing concentrations of in vitro IgE levels synthesized by human peripheral blood lymphocytes (PBL).
D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000276:immunology; D001360:Avidin; D001710:Biotin; D002478:Cells, Cultured; D003470:Culture Media / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006041:Goats; D006801:Humans; D007073:Immunoglobulin E / Q000032:analysis / Q000096:biosynthesis*; D008214:Lymphocytes / Q000276:immunology / Q000378:metabolism; D051379:Mice; D011817:Rabbits
D000888:Antibodies, Anti-Idiotypic; D003470:Culture Media; D001360:Avidin; D007073:Immunoglobulin E; D001710:Biotin