The assay is an immunometric ('sandwich') EIA for the quantitative
measurement of LH in human serum or plasma. LH in the sample is bound by
2 monoclonal anti-LH antibodies directed at different epitopes. One
antibody is attached to magnetic particles and the other is labelled with
alkaline phosphatase. The assay has four main stages:
Magnetic anti-LH is incubated with sample (100 µL) for 15
minutes at 37°C followed by a magnetic wash step.
The magnetic particles are incubated with alkaline
phophatase labelled anti-LH for 1 hour
at 37°C followed by 2 magnetic wash steps.
A coloured enzyme substrate is incubated with the particles for
30 minutes at 37°C. The presence of alkaline phosphatase causes a
colour change from yellow to pink.
The reaction is terminated by addition of Stop Buffer.
The tubes are then placed in the spectrophotometer or colorimeter.
The optical density (at 550nm and 492nm) of each tube can be measured
and the results calculated using a data processing program.