Thyroxine-binding globulin biosynthesis in isolated monkey hepatocytes.

Thyroxine-binding globulin biosynthesis was demonstrated in hepatocytes isolated from normal adult Rhesus monkeys. Dispersed cells were obtained by in situ liver perfusion with collagenase, hyaluronidase and EDTA. Conditions for optimum cell survival and incorporation of radioactive leucine into newly synthesized proteins were defined. Protein synthesis, and specifically thyroxine-binding globulin synthesis, were shown to continue throughout the incubation period, while cell survival remained high (75% excluded trypan blue after 6h). Incubation medium, cytosol and a particulate fraction (extracted with digitonin) were analyzed for thyroxine-binding globulin. After extensive dialysis and purification by affinity chromatography, newly synthesized thyroxine-binding globulin was identified by specific double-antibody immunoprecipitation and by immunodiffusion and immunoelectrophoresis with autoradiography. Newly synthesized thyroxine-binding globulin was present after 4 h of incubation. After 6 h, the total synthesized had increased to 150% of the 4 h value, while the fraction present in the medium and increased to 300%, indicating probable thyroxine-binding globulin secretion