Isolation, Characterization, and Synthesis of AlphaFetoprotein from Neonatal Rat Brain

Abstract: Monospecific anti‐rat serum alpha‐fetoprotein (AFP) IgG was coupled to cyanogen bromide‐activated Sepharose‐4B (4.5 mg/ml packed volume of gel) to yield an immunoaffinity matrix. The immunoaffinity column was used to isolate AFP from feto‐neonatal rat brain. The purified AFP was immunologically and electrophoreti‐cally similar to serum AFP. It yielded a single band with a molecular weight of 70,000 on sodium dodecyl sulphate polyacrylamide gel electrophoresis. Polyacrylamide gel electrophoresis of the protein under nondenaturing conditions yielded two charge variants of AFP, reminiscent of AFP from feto‐neonatal rat serum. The AFP was observed to bind estradiol with K_a= 5.8 × 10⁸M⁻¹ and 1.3 × 10⁸M⁻¹ by dextran‐coated charcoal adsorption and Sephadex gel filtration techniques, respectively. Newborn rat brain cells linearly incorporated [¹⁴C]leucine into immunoprecipitable AFP during 6 h in culture. It is, therefore, concluded that feto‐neonatal rat brain contains AFP similar to that present in fetal serum and that it may arise in brain as a result of its in situ synthesis.