We compared the efficacy of 4 methods for isolating circulating tumor cells: immunocapture with Ber-EP4–coated magnetic beads, density gradient separation, ammonium chloride, and distilled water–mediated erythrocyte lysis. Human blood from healthy volunteers was mixed with serial dilutions of prostate (LNCaP) and liver (HepG2) derived tumor cells. Isolation of circulating tumor cells was followed by reverse transcriptase–polymerase chain reaction with primers specific for prostate-specific antigen and α-fetoprotein. Ber-EP4 antigen expression was evaluated by immunohistochemistry in 27 hepatocellular carcinomas and 34 prostate adenocarcinomas. Peripheral blood samples from 12 patients with hepatocellular carcinoma and 10 with prostate adenocarcinoma also were tested. Density gradient separation and Ber-EP4 immunocapture were the most sensitive techniques for isolating circulating tumor cells in in vitro tests. Isolation by density gradient separation was significantly more sensitive than Ber- EP4 immunocapture when applied to peripheral blood samples of patients with cancer, a result consistent with the variable expression of Ber-EP4 antigen that we found by immunohistochemistry in prostate adenocarcinomas and hepatocellular carcinomas.
This work was supported by grants from the Association pour la Recherche sur le Cancer, France, Assistance Publique (contract no. CRC95019I), France, and Ligue Nationale contre le Cancer, France.