Interlaboratory proficiency, intermethod comparison, and calibrator suitability in assay of serum aspartate...

Sources of variation in assays of aspartate aminotransferase (EC 2.6.1.1) activity were examined in an interlaboratory survey and through an examination of materials used as calibration materials in these assays. Four highly stable lyophilized specimens containing human cytoplasmic enzyme, with activities of 0, 22, 46, and 96 U/liter at 30 degrees C and optimal substrate concentrations, were assayed by 319 laboratories. Mean values obtained on these specimens by laboratories using 2,4-dinitrophenylhydrazine kits varied among manufacturers and deviated from values expected from this procedure. The average coefficient of variation (CV) with these kits was greater than 20%. Automated continuous-flow procedures with use of diazonium salt showed the best precision (av CV, less than 10%). However, the automated continuous-flow malate dehydrogenase/NADH coupled method produced an average CV greater than 20%. Results from each of the automated methods were related to a reference malate dehydrogenase/NADH coupled continuous kinetic assay method by temperature relationships alone. Mean values from manual diazonium salt procedures were 1.7-fold greater than similar reference values (av CV was 18%). The higher results were attributed to the use of poorly-defined units and to an artifact caused by chromophore stabilizers in this procedure when aqueous samples are used. The average CV in continuous kinetic methods varied among kit manufacturers, ranging from 6 to 28% for the specimen of highest activity. Variations in results were much larger at 366 nm than at 340 nm than at 340ity. Variations in results were much larger at 366 nm than at 340 nm. Interassay relationships of these methods are presented. Concentrations of pyruvate in commercially available calibration materials differed between manufacturers, varied in stability, and deviated from the expected concentration. For some colorimetric assays the precision attained on reported absorbance values for the enzyme specimens was of the same order of magnitude as that for pyruvate standards. Other sources of error are revealed by the interlaboratory survey. The value of commercially available sources of enzyme activity as calibration or control materials was assessed by evaluating the following properties: activity at suboptimal concentrations of L-aspartate or 2-oxoglutarate, temperature effects, preincubation lability owing to aspartate and phosphate, pyridoxal phosphate saturation, contamination with glutamate dehydrogenase, and manufacturer's rated activity. These properties are compared to those of human cytoplasmic enzyme in a human serum matrix.