The tumor suppressor gene p53 is mutated in >50% of hepatocellular carcinomas (HCCs) in many geographic areas of the world, regardless of whether the HCC is associated with hepatitis B or C virus. It is not known whether the host produces antibodies to p53 in HCC; such antibodies have been detected in between 9% to 52% of patients with non-hepatic cancers, but in only 0.4% of healthy blood donors. These antibodies often appear prior to a diagnosis of cancer. Attempts were made to develop an assay to detect antibodies to p53 in HCC patients. An EIA was attempted by using p53 proteins produced by a recombinant vaccinia virus containing the gene for human p53. This was not successful because of a persistently weak signal in the assay. Subsequent approaches used the western blot method. p53 proteins were separated on SDS/PAGE and blotted onto a membrane, which was subsequently cut into narrow strips. Serum from each patient was diluted and incubated with each membrane strip and the reactive serum was identified by exposing the ECL-treated strip to X-ray film. Thirty-six serum samples from HCC patients and controls have been tested multiple times to develop reliable and reproducible conditions for the assay.
