City of Hope: Analysis and Purification of CEA samples

Abstract

Carcinoembryonic Antigen (CEA) is an oncofetal antigen that is expressed during human fetal development. While its expression is very limited in adults, CEA can be highly expressed on the surface of adenocarcinomas that form breast, pancreatic, and colorectal cancers. This enables CEA to be used as a tumor marker for cancer diagnosis and immunotherapy. The City of Hope Department of Immunology developed several monoclonal antibodies specific to CEA such as the T84.66 antibody to target cancer cells for radioactive imaging and therapy. Our project this summer was to analyze nine CEA samples that were purified in the 1970s to determine their current quantity, purity and quality. This source of CEA will be used as an antigen for immunoassays as well as an immunogen for the isolation of new human anti-CEA antibodies. After analysis through Western blots, Coomassie Blue staining and glycoprotein staining of the CEA samples run on SDS-PAGE, we discovered that two of the CEA samples were not pure, as a result, we explored re-purification methods. The Enzyme-Linked Immunosorbent Assay (ELISA) was used to quantify the binding activity of the pure CEA samples. Current pilot purification experiments are being performed to purify samples 2 and 3. Further protein analysis will be done by a competitive ELISA to measure the CEA activity per unit of protein. Successful purification, quantification, and analysis of the CEA samples will produce a new supply of CEA that can be used to evaluate biochemically engineered antibodies for purposes of detecting tumors in cancer treatments.