Factors affecting measurement of total alkaline phosphatase activity in human serum, especially wavelength accuracy.

Serum alkaline phosphatase estimations by the Selected Method [Clin. Chem. 21, 1988 (1975)] are very sensitive to small errors in the wavelength setting of the spectrophotometer. With a setting error as small as 3 nm, results can be too low by as much as 23%. The product of the reaction, 4-nitrophenol, can be prepared in very pure form and can be used to check both the wavelength and absorbance accuracy of a spectrophotometer. The molar absorptivity for 4-nitrophenol obtained in the instrument being used should be used in calculating alkaline phosphatase activities.