An automated p-nitrophenylphosphate serum alkaline phosphatase procedure for the AutoAnalyzer.

Abstract

A new and simplified procedure for the automated determination of serum alkaline phosphatase uses the AutoAnalyzer. The substrate p-nitrophenylphosphate in 2-amino-2-methyl-l-propanol offers the prime advantage of providing directly its own chromogen, p-nitrophenol, following enzyme action. The procedure also permits use of small sample volumes, provides linear reaction rates, and has a simple manifold de-sign. Correlations are presented between the manual procedure and the automated system. T1HIS PAPER reports the development of a simplified automated proce-dure for the determination of serum alkaline phosphatase. The method, developed for use with the AutoAnalyzer,t utilizes a p-nitrophenyl-phosphate substrate which, following enzyme action, produces its own chromogen, p-nitrophenol. One existing automated serum alkaline phosphatase procedure (1) using a phenylphosphate substrate is based on the procedures of Kind and King (2) aimd Powell and Smith (3). Recently, this method has been modified to permit simultaneous determination of both the emmzyme reaction and control (4). The product of enzyme action, phenol, must be further treated with aminoantipyrine and alkaline ferricyanide to produce a pink or reddish color whose absorbance is measured. Both automated methods express the results in King-Armstrommg units (5).