The autonomous production of individual serum proteins by tissue in culture

Abstract

A method is described for the autonomous production of individual serum proteins by tissue culture. The tissue is incubated with nutrient medium containing a C14-labelled amino acid (tyrosine or leucine), the soluble proteins are separated and purified by ultrafiltration, f ractiona-tion with ethanol and starch-gel electrophoresis, and the radioactivity of the individual proteins is measured. The proteins of the nutrient medium contained no radiocarbon after incubation with the radioactive amino acid in the absence of tissue. When tissue was incubated with medium and carboxyl-labelled leucine, and the isolated protein was treated with ninhydrin appreciable quantities of radioactive CO2 were evolved only if the protein had been hydrolysed. By these methods, the incorporation of radioactive tyrosine or leucine into serum albumin and a-, [beta]- and gamma-globulin was demonstrated with chick-embryo-mesenchyma tissue, human tumor HeLa tissue, and human transformed-liver HLM tissue. A crude estimate of the rates of production that 15 mg of albumin was produced/g of dry HeLa tissue in 48 hr., and 24 mg/g in chick-mesenchyma tissue. The identity of the labelled serum albumin was confirmed immunologically by treatment with antisera to chick- and human-serum albumin. In the experiments with chick mesenchyma, the bulk of the radiocarbon was found in the precipitate only after reaction with anti-chick serum albumin, whereas with HeLa and HLM tissue the opposite was true.