ELIZEN Proinsulin ELISA Kit (96 Wells)

Principle of the assay: Proinsulin kit is a sensitive two-site sandwich enzyme linked immunosorbent assay. The microtiter plates are coated with a monoclonal antibody (S2) specific for epitope at the C-peptide/insuline A chain junction. S2 is able to bind intact PI, des (31, 32)-PI and split (32, 33)-PI but not insulin, C-peptide and the other “des” and “split” forms. Prior to use a blocking buffer is added to the allocated wells. An aliquot of ample is then added to the wells and incubated. The wells are then washed to remove unbound antibody and other serum compounds. In a second incubation time, an enzyme labelled anti-proinsulin antibody (S53) is incubated in the wells. This antibody is specific for the epitopes at insulin ? chain/C-peptide junction. S53 is able to bind to intact PI, des (64, 65)-PI and split (65, 66)-PI but not insulin, C-peptide and other “des” and “split” forms. The combination of the two Mabs has the ability to detect only the intact human proinsulin. After washing away any unbound enzyme labelled anti-proinsulin antibody, the enzyme activity is measured by adding a chromogen substrate. The intensity of colour development is proportional to the concentration of proinsulin in the sample.

Application: Proinsulin (PI), a polypeptide of 9390 MW (86 amino acids), is synthesized in the b cells of the Islets of Langerhans in the pancreas(1). The sequence of proinsulin is highly conserved in mammalian species and is homologous with IGF-1 and IGF-2 (2). This protein is processed to C-peptide and insulin forms (3). Both are secreted in equimolar amounts into the blood. In normal individuals, proinsulin is present in the circulation in very low concentrations (typical basal values 2-6 pmol/L). The level of proinsulin can be a reflection of b cell status and a consequence of disfunction of PI processing and/or secretion (4). Proinsulin like material is increased in conditions as insulinoma, familial hyperinsulinemia, non-insulin dependent diabetes mellitus (5, 6, 7, 8).