RAPID

EXTRACTION OF HUMAN ERYTHROCYTE CHOLINESTERASE BY ALKALI PSEUDOAGGLUTINATION

In a clinical investigation of cholinergic states we encountered the need for a potent cholinesterase of human origin. The most readily available human source is the erythrocyte, which contains the enzyme in appreciable amounts. Cholinesterase can be liberated from red blood cells by a variety of methods, all of which have in common the destruction of the cell. For our experiments the presence of hemoglobin in the final substrate is a disadvantage. This communication reports a method for rapid extraction of human erythrocyte cholinesterase which is comparatively free of products of hemolysis. The method is based on the observation of one of us1 that pseudoagglutination occurs when erythrocytes are treated with dilute alkali. By extracting chilled human erythrocyte paste for 3 hours with 0.1 volume of 0.85 per cent NaCl solution to which had been added sufficient concentrated NH,OH solution to raise the pH of the suspension to 8.3, maximal esterase extraction with minimal hemolysis is accomplished. The supernatant, when removed from the clumped cells, has an esterase activity approaching that of corpuscle paste and a protein content of about 0.5 per cent. Thus a loo-fold concentration is effected. The cholinesterase extract should immediately be subjected to suction to remove NH3 and so lower the pH toward neutrality. The solution then may be stored indefinitely at ice box temperature, or subjected to further concentration by ammonium sulfate fractionation, or lyophilization.
Halloran General Hospital Staten Island New York Received for publication, January 6,1947 JOHNMENTHA HELMUTH SPRINZ ROBERTBARNARD

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1 J. Lab. and Clin. Med., 23,98 (1937).

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