A procedure using immobilized antibody for the isolation of the beta-subunit of human chorionic gonadotropin from the culture fluid of a beta-subunit-secreting nontrophoblastic cell line.

Antibody produced against the beta-subunit of hCG was linked to Sepharose 4B and used to isolate ectopic hCG beta secreted by DoT cervical cancer cells in culture. beta-Subunit was eluted with 3 M guanidine-hydrochloride, pH 3.0 (three treatments, 15 min each, at 20 C). The eluted hCG beta was desalted with Sephadex G-25 and concentrated by lyophilization. A single batch of antibody-bound Sepharose was used repeatedly to adsorb hCG beta from liter quantities of spent culture fluid containing 10% fetal bovine serum. The material recovered from 15.75 liters of fluid was pooled and treated a second time with immobilized antibody. Using only immobilized antibody, purification of more than 200,000-fold was achieved, with an overall recovery of about 28%.

Duke Scholars

Author Mary T Story Family Medicine and Community Health