Principle of Assay
This Vitamin B12 ELISA Kit is an in vitro competitive enzyme-linked immunosorbent assay for the quantitative measurement of Vitamin B12 in plasma, tissue homogenates and other biological fluids.
A microplate has been precoated with Vitamin B12. Samples, standards and Anti-Vitamin B12 Antibody (Biotin) are added to the wells, where Vitamin B12 in the sample competes with the fixed Vitamin B12 to bind to the antibody. The wells are washed to remove unbound material and HRP-conjugated streptavidin is added. After incubation, the wells are washed again and TMB substrate is added. The TMB substrate is catalyzed by HRP to produce blue colouration. The reaction is terminated by addition of Stop Solution which stops the colour development and produces a colour change from blue to yellow. The intensity of the signal is inversely proportional to the amount of Vitamin B12 in the sample and the intensity is measured at 45 nm.
