The quantitation of biotinidase activity in dried blood spots using microtiter transfer plates: identification of biotinidase-deficient and heterozygous individuals.

A simple and rapid method for the quantitation of biotinidase activity in blood-soaked filter paper spots was developed. The assay measures the release of p-aminobenzoate from N-biotinyl-p-aminobenzoate. A microtiter transfer plate is used to rapidly separate the reaction solution from the filter paper spots. Color is developed and the absorbance is determined using a microplate reader. The biotinidase activity in frozen filter spots correlates well with the activity in serum (r = 0.94). The enzyme activities of obligate heterozygotes for biotinidase deficiency were significantly different from those with normal activity (P = 0.03). This rapid screening procedure can be used to quantitate biotinidase activity in newborn screening samples, identify heterozygotes, and estimate the gene frequency and incidence of biotinidase deficiency in large populations. In addition, the use of microtiter transfer plates can be applied to other assays in which the separation of the incubation solution from a filter paper spot is required.