Determination of serum follicle stimulating hormone (FSH) level
Biotin-conjugated anti-FSH and standard or sample is incubated in monoclonal anti-FSH antibody-coated wells. After 15~18 hours incubation and washing, HRP (horseradish peroxidase)-conjugated avidin is added and incubated for 30 minutes. After washing, HRP-complex remaining in wells are reacted with a chromogenic substrate (TMB) for 30 minutes, and reaction is stopped by addition of acidic solution, and absorbance of yellow product is measured spectrophotometrically at 450 nm (sub-wavelength is 620nm). The absorbance is nearly proportional to FSH concentration. The standard curve is prepared by plotting absorbance against standard FSH concentrations (Markkula, Hämäläinen, Loune, & Huhtaniemi, 1995).