Insulin (rat/mouse)
This Rat / Mouse Insulin ELISA kit is used for the non-radioactive quantification of insulin in mouse and rat sera. Plasma samples may also be used but application to samples of other biological fluids may need validation by the user. One kit is sufficient to measure 39 unknown samples in duplicate.
This assay is a Sandwich ELISA based, sequentially, on: 1) capture of insulin molecules from samples to the wells of a microtiter plate coated by pre-titered amount of a monoclonal mouse anti-rat insulin antibodies and the binding of biotinylated polyclonal antibodies to the captured insulin, 2) wash away of unbound materials from samples, 3) binding of horseradish peroxidase to the immobilized biotinylated antibodies, 4) wash away of free enzyme conjugates, and 5) quantification of immobilized antibody-enzyme conjugates by monitoring horseradish peroxidase activities in the presence of the substrate 3,3’,5,5’-tetramethylbenzidine. The enzyme activity is measured spectrophotometrically by the increased absorbency at 450 nm, corrected from the absorbency at 590nm, after acidification of formed products. Since the increase in absorbency is directly proportional to the amount of captured insulin in the unknown sample, the latter can be derived by interpolation from a reference curve generated in the same assay with reference standards of known concentrations of rat insulin.