Based on our population reference range for biotinidase enzyme activity screening, we updated the cut-off value to 37 U instead of 44 U. The cut-off was updated in the year 2019, after 7 years and 6 months, which resulted in about a 4% reduction in false positive cases for biotinidase deficiency. The gene discussed is BTD; the disease is hyperinsulinemic hypoglycemia, familial, 4.