Purification and characterization of human neuron-specific enolase: radioimmunoassay development.
Human neuron-specific enolase, NSE, (E.C.4.2.1.11), has been purified from the brain. The method includes a salt precipitation and column chromatography on DEAE-Sephacel, Sephacryl S-300, DEAE-Sepharose and Blue-Sepharose. The specific activity of the purified NSE was 51 units/mg protein and the yield was 28%. The molecular weight of human NSE was compared to the bovine form purified by the same procedure. On Sephadex G 150 both human and bovine NSE chromatographed, in the dimeric form, as 77,000 Dalton proteins, while the molecular weight of the monomer was 45,000 Daltons as determined by SDS poly-acrylamide gel electrophoresis. An antiserum, specific for NSE, was raised in rabbits and a radioimmunoassay developed. The contribution of lyzed blood cells to the serum NSE levels in eight healthy donors was determined and correlated with the haemoglobin content. Minimal haemolysis can add 5-10 ng NSE/ ml to the true serum level. For accurate serum NSE determinations, serum samples must be free from haemolysis.
D001921:Brain / Q000201:enzymology*; D002847:Chromatography, Agarose; D004591:Electrophoresis, Polyacrylamide Gel; D006461:Hemolysis; D006801:Humans; D007527:Isoenzymes / Q000302:isolation & purification*; D008970:Molecular Weight; D010751:Phosphopyruvate Hydratase / Q000097:blood / Q000276:immunology / Q000302:isolation & purification*; D011863:Radioimmunoassay
D007527:Isoenzymes; D010751:Phosphopyruvate Hydratase