Determination of delta-aminolevulinic acid dehydratase (ALAD) by enzyme immunoassay and its application to evaluation of lead exposure

Abstract

Enzyme Immunoassay (EIA) technique using a glass bead was established in order to quantify the total amount of delta-aminolevulinic acid dehydratase (ALAD, EC 4, 2, 1, 24) including an inactivated enzyme and to determine the level of ALAD in erythrocytes of workers with moderate lead exposure. EIA was carried out by competitive antibody binding method in which the antigen (ALAD) was linked to a glass bead (O.D. 7 mm). A standard curve was fitted to the function y = A X B/(x-B) + C, where y is the enzymatic reaction product, x is the concentration of ALAD, and A, B and C are parameters. The coefficients of variation in the intra- and inter-assay were 3.7% and 6.5%, respectively. EIA-based amount of ALAD was 136.7 +/- 23 mg/l erythrocyte (mean +/- S.D.) in 66 workers. This ALAD amount was not found to be correlated with the lead level in the blood but correlated with the duration of lead-exposure. An especially good correlation (r = 0.453) was obtained with the inactivated portion of ALAD (EIA-based amount minus activity-based amount). On the other hand, the ratio of restored activity following heat treatment and addition of Zn and dithiothreitol to non-restored activity was well correlated with the lead level (r = 0.874), but not correlated with the duration of lead-exposure (r = 0.218). The inactivated portion of ALAD based on EIA accordingly may be considered a good indicator of the duration of lead exposure, and the activity-based amount of ALAD may be considered a suitable indicator of the extent of the present lead exposure.