Analysis of Biotinidase Activity in Serum by Digital Imaging Colorimetry Detection

Biotinidase deficiency (BD) is an autosomal recessive inherited disorder of biotin recycling that leads to neurological and cutaneous consequences if left untreated. The clinical features of BD can be ameliorated or prevented by the administration of pharmacological doses of the vitamin biotin. Since it is a treatable disorder, BD is included in the newborn screening program in Türkiye as in many other countries. Therefore, monitoring of biotinidase enzyme activity (BEA) is of vital importance, especially for patients. The aim of this study was to develop a simple and reliable colorimetric method based on digital imaging for the analysis of BEA in serum samples. To determine the optimum distance and LED light source in the analyzer box that we fabricated in the laboratory, images of the solutions in a 96-well microplate were taken with a mobile phone camera, and each color space was examined. The most reliable relationship was between blank subtracted intensities of green channel and analyte concentrations, which was in the range of 35-400 ng/mL p-aminobenzoic acid (r² = 0.999). The limit of detection and limit of quantification were 11 and 35 ng/mL, respectively. The proposed method was successfully applied to serum samples of 60 patients with BD and 60 healthy controls. We claim that the method can be easily performed for determination of BEA anywhere without needing expensive instruments.