Amplified enzyme-linked immunoassay of human proinsulin in serum (detection limit: 0.1 pmol/L).
We describe an amplified enzyme-linked immunoassay of human proinsulin in serum that detects intact proinsulin and both the 32/33 and 65/66 split forms. The method uses the IgG fraction of a polyclonal antibody raised in a guinea pig against intact proinsulin, which we used to coat plastic microtiter plates. A sandwich was formed with proinsulin by using a monoclonal antibody against C-peptide labeled with alkaline phosphatase. We quantified the reaction by using the enzyme amplification procedure, which detected as little intact proinsulin as 0.1 pmol/L. We found no cross-reactivity with C-peptide in the assay, and decreased recovery attributable to the presence of insulin could be demonstrated only with a 30-fold excess of this hormone over proinsulin.
Dhahir
F J
FJ
Department of Clinical Biochemistry, Medical School, University of Newcastle upon Tyne, U.K.
Cook
D B
DB
Self
C H
CH
D000469:Alkaline Phosphatase; D000818:Animals; D000911:Antibodies, Monoclonal; D002096:C-Peptide / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006168:Guinea Pigs; D006801:Humans; D007074:Immunoglobulin G; D015592:Immunoradiometric Assay; D007328:Insulin / Q000097:blood; D008832:Microchemistry; D011384:Proinsulin / Q000097:blood* / Q000276:immunology; D011786:Quality Control; D012016:Reference Values
D000911:Antibodies, Monoclonal; D002096:C-Peptide; D007074:Immunoglobulin G; D007328:Insulin; D011384:Proinsulin; D000469:Alkaline Phosphatase