Principle of Assay
Human SEMA4A ELISA Kit (A310444) employs the sandwich enzyme immunoassay technique for the quantitative measurement of human SEMA4A in serum, plasma or other biological fluids. An antibody specific for SEMA4A has been pre-coated onto a 96-well microtiter plate. The standards and test samples are added into the wells and the SEMA4A present in each sample is bound to the wells by the immobilized antibody. Biotinylated Anti-SEMA4A Antibody, which also binds the SEMA4A present in each sample, and Streptavidin-HRP, which binds the Biotinylated Anti-SEMA4A Antibody, are added and the microtiter plate is incubated. Following incubation, unbound Biotinylated Anti-SEMA4A Antibody and unbound Streptavidin-HRP are removed by washing, and two substrate solutions are added to the wells. Color develops in proportion to the amount of SEMA4A captured in each well. The color development is stopped by addition of stop solution which changes the color from blue to yellow and the intensity of the color is then measured. The concentration of SEMA4A in the samples can then be calculated by reading the O.D. absorbance at 450nm in a microplate reader and referring to the standard curve.
