Prolactin Milk/Plasma ELISA Assay Kit

$750.00

The Eagle Biosciences Prolactin ELISA Assay Kit is intended  for the quantification of human Prolactin in human plasma and milk samples. The Prolactin ELISA Assay Kit is for research use only and should not be used for diagnostic procedures.

Prolactin Milk/Plasma ELISA Assay Kit

For Research Use Only

Size: 1×96 wells
Sensitivity: 0.138 ng/ml
Dynamic Range: 0.25-100 ng/mL
Incubation Time: 2 hours
Sample Type: Plasma and Milk
Sample Size: 100 µL

Additional Information

Assay Background

Prolactin which is a secreted hormone is a growth regulator for many tissues, including cells of the immune system. It may also play a role in cell survival by suppressing apoptosis, and it is essential for lactation. Alternative splicing results in multiple transcript variants that encode the same protein. Prolactin acts primarily on the mammary gland by promoting lactation.

Assay Principle

This assay employs the sandwich enzyme immunoassay technique for the detection of prolactin in human plasma and milk samples. The microtiter plate is coated with affinity purified prolactin antibody. Human Prolactin in samples or standards will react with antibody coated on the microtiter plate. After washing, a biotin-labeled polyclonal anti-Human Prolactin antibody is added and bound to the captured prolactin on the microtiter plate. Excess antibody is washed away and bound primary antibody is reacted with a Streptavidin conjugated to horseradish peroxidase (HRP). Following an additional washing step, TMB is added in the wells. HRP catalyzes TMB substrate solution, resulting in the development of a blue color. The color development is inhibited by the addition of a stop solution, and the color turns yellow. The yellow color is measured at 450 nm. The concentration of prolactin in test sample is directly proportional to the color intensity, which can be determined by extrapolation to the standard curve.

Assay Procedure

  1. Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal it.
  2. Add 100 µl of standards or unknown samples into appropriate wells. Shake the plate on a microplate shaker at 300 rpm for 30 minutes at RT.
  3. Aspirate each well and wash, repeating the process 2 times for a total 3 washes. Wash by filling each well with 1× assay buffer (300 μl) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each is essential to good performance. After the last wash, remove any remaining buffer by aspirating, decanting or blotting against clean paper towels.
  4. Add 100 µl of primary antibody into each well. Shake the plate on a microplate shaker at 300 rpm for 30 minutes at RT.
  5. Wash as according to step 3.
  6. Add 100 µl of HRP-conjugated Streptavidin (1:25,000 diluted) into each well. Shake the plate on a microplate shaker at 300 rpm for 30 minutes at RT.
  7. Wash as according to step 3.
  8. Add 100 μl of TMB Reagent to each well. Shake plate at 300 rpm for 5-20 minutes at RT in dark. Substrate will change from colorless to different strengths of blue.
  9. Add 50 μl of 1N H2SO4 or HCl to each well. The color of the solution should change from blue to yellow. Mix thoroughly by gently shaking the plate.
  10. Read the OD with a microplate reader at 450 nm immediately.

Manual

Product Manual