Principle of Assay
This direct competitive ELISA is designed to measure the level of S-adenosylhomocysteine (SAH) in the sample. SAH conjugated with macromolecule is immobilised on the micro-titre plate. Standards and samples are pipetted into the wells, and the Anti-SAH antibody (HRP) is added. The free SAH molecule in samples or standards competes with immobilised SAH on the micro-titer plate surface for binding sites of the antibody. After discarding the mixed solution and washing each well, TMB substrate solution is added. The substrate solution turns blue under the effect of HRP, and changes into yellow once stop solution (acid) is added. The colour develops in inverse proportion to the amount of SAH in the sample (or standards). The optical density of the remaining solution (OD450) is measured at 450nm using micro-plate spectrophotometer. The level of SAH in samples can be calculated through standard curve generated with standards.
