Active Prostate Specific Antigen (PSA)

Prostate Specific Antigen (PSA) is a serine protease enzyme primarily produced by the epithelial cells of the prostate gland. Its key physiological function is to liquefy seminal fluid, facilitating sperm motility. Clinically, measuring serum PSA levels is a cornerstone for screening and monitoring prostate cancer, as elevated concentrations often indicate malignancy, though benign conditions like prostatitis can also cause increases. It is considered an important, though not perfectly specific, tumor marker. PSA can form complexes with other serum proteins, most notably with Alpha-2-Macroglobulin (a2M), a large plasma protease inhibitor. This binding irreversibly encapsulates PSA, effectively neutralizing its enzymatic activity and removing it from serological detection.Thus to detect the activity of recombinant PSA, a functional ELISA assay was performed to evaluate the interaction between recombinant human PSA and recombinant mouse a2M.
Briefly, biotin-linked PSA were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100µl were then transferred to a2M-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST 3 times and incubation with Streptavidin-HRP for 30min, then wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50µl stop solution to the wells and read at 450nm immediately. The binding activity of PSA and a2M was shown in Figure 1, the EC50 for this effect is 4.5 µg/mL.