INS, ELISA

The DiaSource INS-ELISA is a solid phase Enzyme Amplified Sensitivity Immunoassay performed on breakable microtiterplates. The assay uses monoclonal antibodies (MAbs) directed against distinct epitopes of insulin.Calibrators and samples react with the capture monoclonal antibody (MAb 1) coated on microtiter well and with a monoclonal antibody (MAb 2) labelled with horseradish peroxidase (HRP).

After an incubation period allowing the formation of a sandwich: coated MAb 1 – human insulin – MAb 2 – HRP, the microtiterplate is washed to remove unbound enzyme labelled antibody. Bound enzyme-labelled antibody is measured through a chromogenic reaction. Chromogenic solution (TMB ready for use) is added and incubated.

The reaction is stopped with the addition of Stop Solution and the microtiterplate is then read at the appropriate wavelength. The amount of substrate turnover is determined colourimetrically by measuring the absorbance, which is proportional to the insulin concentration.

A calibration curve is plotted and INS concentration in samples is determined by interpolation from the calibration curve.