DETECTION OF SECRETED AND INTRACELLULAR ANNEXIN-II BY A RADIOIMMUNOASSAY

A simple radioimmunoaasay for detection of secreted and intracellular annexin II in human cells is presented. Annexin II is a multifunctiona l protein in human cells and may have a role in several types of cance rs. No enzymatic activity has been associated with the protein, thus m aking its detection difficult. Using purified annexin II from human pl acenta, we have developed a sensitive radioimmunoassay protocol. A lin ear response was observed up to a concentration of 0.5 mu g purified p rotein in the assay. Using this radioimmunoassay protocol, annexin II can be detected in undiluted clinical human samples such as bronchoalv eolar lavages and various tissue extracts. We demonstrate the applicab ility of this technique to measure intracellular annexin II in extract s of a human adenocarcinoma cell line (HeLa) and secreted annexin II f rom bronchoalveolar lavage fluid from a human patient. Using HeLa cell extracts and BAL, we observed a linear response with up to 10 mu g to tal protein in the assay. We further demonstrate the applicability of this technique to measure differences in intracellular and secreted an nexin II in the human pancreatic adenocarcinoma cell lines CD-11, CD-1 8 and Capan-2. While CD-11 and CD-18 do not secrete annexin II, the ce ll line Capan-2 secretes high levels of the protein.