The assay is a direct competitive EIA for the quantitative measurement
of Progesterone in human serum or plasma. Progesterone in the sample is
displaced from serum binding proteins by a blocking reagent and bound by a
monoclonal anti-Progesterone antibody. The complex is isolated by
adsorbtion onto a magnetic anti-mouse IgG separation reagent, and
unoccupied anti-Progesterone sites are bound by alkaline phosphatase
labelled Progesterone tracer. The assay has four main stages:
Anti-Progesterone antibody, blocking reagent and magnetic
anti-mouse IgG are incubated with sample (50 µL) for 2 hours at
37°C followed by a magnetic wash step.
The magnetic particles are incubated with alkaline
phophatase labelled Progesterone tracer for 15 minutes at 37°C
followed by 2 magnetic wash steps.
A coloured enzyme substrate is incubated with the particles for 1 hour
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) of each tube can be measured and the
results calculated using a data processing program.