Bovine alpha-fetoprotein. Isolation and characterization

Abstract

Bovine AFP was purified by ion exchange chromatograph on C.M. cellulose and DEAE Sephadex A-50, gel filtration and immunosorbent technique. AFP was homogeneous when studied by gel electrophoresis under non denaturing and denaturing conditions, by ultracentrifugation and by immunological methods. The following molecular data were obtained: 1. Sedimentation equilibrium indicated a molecular weight of 66,500 and sedimentation velocity gave s degrees 20, w = 4.71 S. A partial specific volume v = 0.737 ml g-1 was derived from density measurements. 2. From these data, a Stokes radius of 3.26 nm, a diffusion coefficient D20 w = 6.61 10(-7) cm2 sec-1 and a frictional ratio f/fo = 1.21 were calculated. 2. Sodium dodecylsulphate disc electrophoresis suggests a molecular weight of 67,000. 3. Gel filtration pointed to a molecular weight of 75,000. 4. Microheterogeneity of AFP was demonstrated by isoelectric focusing. The isoelectric point of the major component is 4.6. 5. The chemical composition was determined. AFP is a glycoprotein containing 7 per cent carbohydrate including 1.67 per cent hexoses, 2.38 per cent N-acetyl glucosamine and 1.8 per cent N-acetyl neuraminic acid.