Quantitative determination of human aldose reductase by enzyme-linked immunosorbent assay: Immunoassay of human aldose reductase

Abstract

An antibody-sandwich enzyme-linked immunosorbent assay (ELISA) for evaluating tissue levels of aldose reductase was developed using a polyclonal antibody prepared against the recombinant enzyme expressed in a baculovirus system. The specificity of this antibody to aldose reductase was verified by immunoprecipitation, immunoblotting and ELISA. The polyclonal antibody did not crossreact with human aldehyde reductase, an enzyme in the same aldo-keto reductase family structurally and functionally related to aldose reductase. The sensitivity and specificity of this assay method enabled direct determination of aldose reductase level in various human tissues including the erythrocyte. The highest level of aldose reductase was detected in the kidney medulla among tissues investigated. More than a 2-fold variability in the erythrocyte aldose reductase was demonstrated among healthy individuals, indicating the heterogeneity of this enzyme expression in a human population. This assay system may be useful for direct measurement of the level of tissue aldose reductase in conjunction with the evaluation of the efficacy of aldose reductase inhibitors prescribed for the treatment of diabetic complications.