MONOCLONAL ANTIBODIES 2F5 AND 4G10 AGAINST PROSTATE SPECIFIC ANTIGEN (PSA) COMPLEXED TO alpha 1-ANTICHYMOTRYPSIN

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Abstract
We have shown that an immunoassay (Chugai) for the PSA-ACT complex in serum has 2 to 3 times better specificity than total PSA at sensitivities of 85 to 97% in distinguishing biopsy positive men from biopsy negative men undergoing transrectal ultrasound (TRUS) examination. To increase the specificity of PSA immunoassay for prostate cancer, we produced specific antibodies exclusively against our purified PSA-ACT complex for development of assay kits for PSA-ACT complex. PSA-ACT complex was used as antigen to immunize BALB/c mice. PSA-ACT complex, free PSA, and ACT were used for hybridoma screening. To characterize the new monoclonal antibodies, we used Western blot, immunohistochemistry, and an in house immunoassay. Two monoclonal antibodies, 2F5 and 4G10 were produced exclusively against PSA-ACT complex without any immunoreactivity to ACT or PSA alone. Western blot analysis indicated that 2F5 and 4G10 recognize conformation-dependent epitopes on PSA-ACT complex. Immunohistochemistry studies showed that 2F5 reacted with prostate cancer in about 30% of the cancer cells (sensitivity), but almost never stained normal prostate glands in the peripheral or transition zone tissue (about 100% specificity). Our in-house assay showed that 2F5 can be used as a tracer antibody specifically to detect PSA-ACT complex. Using monoclonal antibody 2F5 as tracer antibody, we have developed a PSA immunoassay exclusively against PSA-ACT complex. This assay should maximize specificity in distinguishing BPH from prostate cancer.