Prolactin (PRL) is a 199 aa, 23kD peptide hormone that is secreted primarily by the pituitary gland in both males and females, though its major roles are in pregnancy and lactation. Prolactin may have a role in breast cancer development, with higher prolactin levels correlating with postmenopausal breast cancer risk. The sensitive quantitative measurement of total rat prolactin antigen in plasma is easily performed with this 96 well strip format ELISA kit. The concentration of prolactin in pooled normal rat plasma determined by in house testing was 6.8 ng/ml. Prolactin levels in pregnant rats are elevated immediately before birth. The assay measures total rat prolactin in the 0.1-100 ng/ml range. Samples with rat prolactin levels above 100 ng/ml should be diluted in blocking buffer before use. Normal plasma may be applied directly to the plate or diluted 1:1 with blocking buffer. Rat prolactin will bind to the capture antibody coated on the microtiter plate. After appropriate washing steps, biotin labeled anti-rat prolactin primary antibody binds to the captured protein. Excess primary antibody is washed away and bound antibody is reacted with peroxidase conjugated streptavidin. Following an additional washing step, TMB substrate is used for color development at 450nm. Color development is proportional to the concentration of prolactin in the samples. A standard calibration curve is prepared using dilutions of purified prolactin and is measured along with the test samples. All reagents and standards are provided in these ELISA kits.