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PubMed · 730820

An optimized assay for adenosine deaminase using reverse phase high pressure liquid chromatography.

R Hartwick·A Jeffries·A Krstulovic·P R Brown
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Abstract

A sensitive, optimized assay for adenosine deaminase (E.C. 3.5.4.4), is presented which is based on a reverse phase HPLC analysis of adenosine. In this method a sample of erythrocytes was incubated with adenosine and the decrease in the adenosine concentration with time was analyzed by HPLC. The precision of the method averaged approximately 5% RSD with a sensitivity of about 0.1 U/ml of packed erythrocytes. Comparison with other literature values showed similar activities for adenosine deaminase in erythrocytes (0.229 +/- 0.025) U/ml (alpha = 0.05). The optimization included studies on the ionic strength, pH, enzyme and substrate concentration, and reaction time. The Km for adenosine deaminase was found to be (0.178 +/- 0.018) mM (alpha = 0.05). The method offers several advantages over other assay methods, including an improved capability to discern competing side reactions from other enzymes.

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Adenosine DeaminaseChromatography, High Pressure LiquidErythrocytesHumansNucleoside DeaminasesJournal ArticleResearch Support, U.S. Gov't, P.H.S.

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R Hartwick, A Jeffries, A Krstulovic, P R Brown. 1978. An optimized assay for adenosine deaminase using reverse phase high pressure liquid chromatography.. https://doi.org/10.1093/chromsci%2F16.9.427

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