Introduction Recently, we described a new coupled colorimetric assay for serum activity toward cytidine-S'-monophosphate (5'-nucleotidase activity, EC 126.96.36.199) based on measurement of the equivalent amount of ammonia released from cytidine by cytidine deaminase (EC 188.8.131.52) (1). In developing this assay we used a crude cy tidinedeaminase preparation as the auxiliary enzyme, which resulted in rather high background extinction values and an overcorrection with respect to the reagentcontrol, probably due to an enzyme contaminating the cytidiiie deaminase. To circumvent this problem, we offered an indirect method. In this communication we prove that a simple and rapid purification of the cytidine deaminase is sufficient to give a reliable direct method, with none of the above disadvantages. Materials and Methods Reagents 1. Buffer solution: Dissolve 4.20 g sodium diethylbarbiturate and 6.30 g MgSO4 · 7H2O in about 800 ml of distilled water. Adjust the pH to 7.50 with HC1 (1 mol/1) and dilute to 1,000 ml. 2. Cytidine deaminase solution: Dilute the stock cytidine deaminase (see below) with buffer solution to give an activity of about 225 U/l and dissolve J. Clin. Chem. Clin. Biochem. / Vol. 14,1976 / No. 10 28 mg disodium phenylorthophosphate (British Drug Houses) per 10 ml
Clinical Chemistry and Laboratory Medicine – de Gruyter
Published: Jan 1, 1976
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