Phenylketonuria is an inherited metabolic disease, which is characterized by increased level of serum phenylalanine (Phe). The quantitative measurement of Phe in the serum is necessary to confirm the disease, and to distinguish phenylketonuria from other forms of hyperphenylalaninemia. In this study, we report a rapid and inexpensive micro-assay for simultaneous detection and quantitative measurement of serum Phe in dry blood-spots. Analysis of the standard curve showed a broad linear Phe range of 120–1800 μmol L−1. Application of this method in conjunction with the standard Guthrie bacterial inhibition assay and high-pressure liquid chromatography in analyzing 34 samples from phenylketonuria patients and control samples produced comparable results, with the regression equation of Y = 0.994X + 0.996. The advantage of this method over the Guthrie bacterial inhibition assay is its ability to measure the serum Phe quantitatively without false positive results. The method was successfully applied to dried blood-spots as well as serum and whole blood samples. The cost per sample is about 20–50 US cents, which is much less than those of high-pressure liquid chromatography and enzymatic commercial kits. The method can be automated, which is suitable for neonatal and mass phenylketonuria screening, especially in developing countries, where funding is a limiting factor.
This is a preview of subscription content, log in to check access.
Buy single article
Instant access to the full article PDF.
Price includes VAT for USA
Subscribe to journal
Immediate online access to all issues from 2019. Subscription will auto renew annually.
This is the net price. Taxes to be calculated in checkout.
Autti-Ramo I, Makela M, Sintonen H, Koskinen H, Laajalahti L, Halila R, et al. 2005. Expanding screening for rare metabolic disease in the newborn: an analysis of costs, effect and ethical consequences for decision-making in Finland. Acta Paediatr 94: 1126–1136.
Cook BC, Wilhit TR, Smith CH, Landt M, 1995. Reagent-induced variation in specificity of serum phenylalanine assays. Clin Chem 41: 949–950.
Dooley KC, 1992. Enzymatic method for phenyl-ketonuria screening using phenylalanine dehydrogenase. Clin Biochem 25: 271–276.
Faden R, Chwalow AJ, Holtzman NA, Horn SD, 1982. A survey to evaluate parental consent as public policy for neonatal screening. Am J Public Health 72: 1347–1352.
Gerasimova NS, Steklova IV, Tuuminen T, 1989. Fluorimetric method for phenylalanine assay adapted for phenylketonuria screening. Clin Chem 35: 2112–2115.
Guthrie R, Susi A, 1963. A simple phenylalanine method for detecting phenylketonuria in large populations of newborn infants. Pediatrics 32: 338–343.
Hannelius U, Lindgren CM, Melen E, Malmberg A, von Dobeln U, Kere J, 2005. Phenylketonuria screening registry as a resource for population genetic studies. J Med Genet 42: e60.
Hilton MA, 1982. Liquid chromatographic direct determination of phenylalanine and tyrosine in serum or plasma, with application to patients with phenylketonuria. Clin Chem 28: 1215–1218.
Holmgren G, Larsson A, Palmstiena M, Alm J, 1976. The frequency of PKU and hyperphenylalaninemia in Sweden. A study in institutions for mentally retarded as well as in neonates. Clin Gen 10: 313–318.
Kok WTH, Brinkman UA, Frei RW, 1983. Rapid determination of phenylalanine and tyrosine in urine and serum by HPLC with electrochemical detection. J Pharm Biochem Analyt 1: 368–372.
Krause W, Halminski M, McDonald L, Dembure P, Salvo R, Freides D, Elsas L, 1985. Biochemical and neuropsychological effects of elevated plasma phenylalanine in patients with treated phenylketonuria. A model for the study of phenylalanine and brain function in man. J Clin Invest 75: 40–78.
Lee C, 1993. Enzymatic determination of L-phenylalanine in serum. Lab Med 24: 301–304.
Madira WM, Xavier F, Stern J, Wilcox AH, Barron JL, 1992. Determination and assessment of the stability of phenylalanine and tyrosine in blood spots by HPLC. Clin Chem 38: 216–213.
McCaman MW, Robins E, 1962. Fluorimetric method for the determination of phenylalanine in serum. J Lab Clin Med 59: 885–890.
PAHbd. Phenylalanine hydroxylase locus database. http://www.mcgill.ca/pahdb.
Phenylketonuria. Screening and Management. NIH Consensus Statement October 1618, 2000. 17: 1–27.
Qu Y, Miller JB, Slocum RH, Shapira E, 1991. Rapid automated assay for quantitation of isoleucine, leucine, tyrosine and phenylalanine from dried blood filter paper specimens. Clin Chem Acta 203: 191–198.
Randell EW, Lehotay DC, 1996. An automated enzymatic method on the Roche COBAS MIRATM S for monitoring phenylalanine in dried blood spots of patients with phenylketonuria. Clin Chem 29: 133–138.
Rohr FJ, Allred EN, Turner M, Simmons J, Levy HL, 1996. Use of the Guthrie bacterial inhibition assay to monitor blood phenylalanine for dietary treatment of phenylketonuria. Screening 4: 205–211.
Sarkissian CN, Shao Z, Blain F, Peevers R, Su H, Heft R, Chang TM, Scriver CR, 1999. A different approach to treatment of phenylketonuria: phenylalanine degradation with recombinant phenylalanine ammonia lyase. Proc Natl Acad Sci USA 96: 2339–2344.
Scriver CR, Waters PJ, 1996. Monogenic traits are not simple: lessons from phenylketonuria. Trends Genet 15: 267–272.
Scriver CR, 1998. Commentary on: A simple phenylalanine method for detecting phenylketonuria in large populations of newborn infants, by Robert Guthrie and Ada Susi, 1963. Pediatrics 32 338–343; Pediatrics 236–237.
Vallian S, Barahimi E, Moeini H. 2003. Phenylketonuria in Iranian population: a study in institutions for mentally retarded in Isfahan. Mut Res 526: 45–52.
About this article
Cite this article
Vallian, S., Moeini, H. A quantitative bacterial micro-assay for rapid detection of serum phenylalanine in dry blood-spots: Application in phenylketonuria screening. J Appl Genet 47, 79–83 (2006). https://doi.org/10.1007/BF03194603
- bacterial micro-assay