Assay of Aspartate Aminotransferase Activity: Effects of Serum and Serum Proteins on Oxalacetate Decarboxylation and Dialysis

[摘要] Effects of serum and protein on assays measuring aspartate aminotransferase (L-aspartate:2-oxoglutarate aminotransferase, EC 2.6.1.1) were examined. Increasing amounts of normal human serum or human serum albumin in samples with fixed amounts of purified enzyme decrease absorbance values and thus distort the results obtained in an assay in which diazonium-salt chromogenic reagent is used. Serum also directly accelerates disappearance of oxalacetate from an incubation medium similar to that of the enzyme assay studied. Protein accelerates spontaneous decarboxylation of oxalacetate to pyruvate, preventing its quantitation in assays specific for oxalacetate. Primary amine groups of protein are evidently involved, because acylated albumin does not accelerate decarboxylation. This effect is not attributable to metal ions in serum and is not significant in procedures involving use of the less specific dinitrophenylhydrazine, which measure the decarboxylation product, pyruvate. Continuous kinetic assays of aspartate aminotransferase activity, in which oxalacetate is not allowed to accumulate, are unaffected by protein concentration. Fivefold dilution of patients’ sera with physiological saline results in activities that are artifactually high by 56% in affected assays. Varying oxalacetate decarboxylation is also a source of error in the estimation of malate dehydrogenase (L-malate:NAD oxidoreductase; EC 1.1.1.37) activity in body fluids. Protein increases the values obtained by automated procedures requiring dialysis by increasing the amount of oxalacetate diffusing into the chromogenic flow stream. A solution of albumin, about 6 g/dl, is suggested as a dilution fluid for use with the affected assays.

[学科分类] 过敏症与临床免疫学