Ultrasensitive enzyme-linked immunosorbent assay (ELISA) for the detection of picogram quantities of IgE.
Existing methods of measuring IgE in in vitro peripheral blood lymphocyte (PBL) cultures are not sufficiently sensitive to detect IgE when it is present in small amounts. This paper describes a modification of a two-site ELISA which increases the sensitivity of the assay 10-20-fold. By using the Fab' fragment of either rabbit or mouse monoclonal anti-IgE conjugated to alkaline phosphatase (AP) as the detector, the background of the assay was reduced sufficiently to permit signal amplification, using a commercially available amplified AP substrate. With this assay as little as 10 pg/ml of IgE could be detected. The interassay coefficient of variation was 15-18% between 1200 and 100 pg/ml IgE (n = 14) and there was a good correlation with a commercial IgE radioimmunoassay (RIA) (r = 0.98, n = 38).
Kemeny
D M
DM
Department of Allergy and Allied Respiratory Disorders, United Medical School, Guy's Hospital, London, U.K.
Richards
D
D
Durnin
S
S
Johannsson
A
A
D000469:Alkaline Phosphatase; D000911:Antibodies, Monoclonal / Q000276:immunology; D003877:Dermatitis, Contact / Q000276:immunology; D004306:Dose-Response Relationship, Immunologic; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods* / Q000592:standards; D006801:Humans; D006967:Hypersensitivity / Q000276:immunology; D007073:Immunoglobulin E / Q000032:analysis*
D000911:Antibodies, Monoclonal; D007073:Immunoglobulin E; D000469:Alkaline Phosphatase