Microassay system for newborn screening for phenylketonuria, maple syrup urine disease, homocystinuria, histidinemia and galactosemia with use of a fluorometric microplate reader

Abstract Microfluorometry for measuring phenylalanine, branched-chain amino acids, homocysteine and histidine in dried blood spots is presented as a new mass screening method for phenylketonuria, maple syrup urine disease, homocystinuria and histidinemia, respectively. Simple and rapid determinations of the metabolites are achieved by adapting each of the optimum fluorogenic reactions to a microplate scale followed by using a highly sensitive microplate reader. Each assay for several hundreds of samples can be completed within 3 h, with fully calculated results by an on-line microcomputer. The proposed system, both with simple procedure and quantitative results, is useful for the multiple routine screening for the four aminoacidopathies, in addition to galactosemia as we reported earlier [20]. (1) The usage of a microplate system for routine microplate screening might be able to open a new style of mass-screening. (2) Measurement of homocysteine seems to be more desirable than measuring methionine in order to detect classic homocystinuria, and to enable detection of the remethylatine defects. However, the recovery procedure of homocysteine needs to be improved; furthermore, the measuring of three branched-chain amino acids will be better than determining only leucine for detecting a mild form of maple syrup urine disease or other conditions which increase the levels of these amino acids. The manner in which we measure phenylalanine in our method is more sensitive than others using the microplate system.