Background: Over the past decade laboratories that test for metabolic disorders have introduced tandem mass spectrometry (MS/MS), which is more sensitive, specific, reliable, and comprehensive than traditional assays, into their newborn-screening programs. MS/MS is rapidly replacing these one-analysis, one-metabolite, one-disease classic screening techniques with a oneanalysis, many-metabolites, many-diseases approach that also facilitates the ability to add new disorders to existing newborn-screening panels. Methods: During the past few years experts have authored many valuable articles describing various approaches to newborn metabolic screening by MS/MS. We attempted to document key developments in the introduction and validation of MS/MS screening for metabolic disorders. Our approach used the perspective of the metabolite and which diseases may be present from its detection rather than a more traditional approach of describing a disease and noting which metabolites are increased when it is present. Content: This review cites important historical developments in the introduction and validation of MS/MS screening for metabolic disorders. It also offers a basic technical understanding of MS/MS as it is applied to multianalyte metabolic screening and explains why MS/MS is well suited for analysis of amino acids and acylcarnitines in dried filter-paper blood specimens. It also describes amino acids and acylcarnitines as they are detected and measured by MS/MS and their significance to the identification of specific amino acid, fatty acid, and organic acid disorders. Conclusions: Multianalyte technologies such as MS/MS are suitable for newborn screening and other mass

Medium-chain acyl-CoA dehydrogenase (MCAD) deficiency is the most frequently diagnosed mitochondrial beta-oxidation defect, and it is potentially fatal. Eighty percent of patients are homozygous for a common mutation, 985A-->G, and a further 18% have this mutation in only one disease allele. In addition, a large number of rare disease-causing mutations have been identified and characterized. There is no clear genotype-phenotype correlation. High 985A-->G carrier frequencies in populations of European descent and the usual avoidance of recurrent disease episodes by patients diagnosed with MCAD deficiency who comply with a simple dietary treatment suggest that MCAD deficiency is a candidate in prospective screening of newborns. Therefore, several such screening programs employing analysis of acylcarnitines in blood spots by tandem mass spectrometry (MS/MS) are currently used worldwide. No validation of this method by mutation analysis has yet been reported. We investigated for MCAD mutations in newborns from US populations who had been identified by prospective MS/MS-based screening of 930,078 blood spots. An MCAD-deficiency frequency of 1/15,001 was observed. Our mutation analysis shows that the MS/MS-based method is excellent for detection of MCAD deficiency but that the frequency of the 985A-->G mutant allele in newborns with a positive acylcarnitine profile is much lower than that observed in clinically affected patients. Our identification of a new mutation, 199T-->C, which has never been observed in patients with clinically manifested disease but was present in a large proportion of the acylcarnitine-positive samples, may explain this skewed ratio. Overexpression experiments showed that this is a mild folding mutation that exhibits decreased levels of enzyme activity only under stringent conditions. A carrier frequency of 1/500 in the general population makes the 199T-->C mutation one of the three most prevalent mutations in the enzymes of fatty-acid oxidation.

This review is intended to serve as a practical guide for geneticists to current applications of tandem mass spectrometry to newborn screening. By making dried-blood spot analysis more sensitive, specific, reliable, and inclusive, tandem mass spectrometry has improved the newborn detection of inborn errors of metabolism. Its innate ability to detect and quantify multiple analytes from one prepared blood specimen in a single analysis permits broad recognition of amino acid, fatty acid, and organic acid disorders. An increasing number of newborn screening programs are either utilizing or conducting pilot studies with tandem mass spectrometry. It is therefore imperative that the genetics community be familiar with tandem mass spectrometric newborn screening.

Background: Deaths from inherited metabolic disorders may remain undiagnosed after postmortem examination and may be classified as sudden infant death syndrome. Tandem mass spectrometry (MS/MS) may reveal disorders of fatty acid oxidation in deaths of previously unknown cause. Methods: We obtained filter-paper blood from 7058 infants from United States and Canadian Medical Examiners. Acylcarnitine and amino acid profiles were obtained by MS/MS. Specialized interpretation was used to evaluate profiles for disorders of fatty acid, organic acid, and amino acid metabolism. The analyses of postmortem blood specimens were compared with the analyses of bile specimens, newborn blood specimens, and specimens obtained from older infants at risk for metabolic disorders. Results: Results on 66 specimens suggested diagnoses of metabolic disorders. The most frequently detected disorders were medium-chain and very-long-chain acyl-CoA dehydrogenase deficiencies (23 and 9 cases, respectively), glutaric acidemia type I and II deficiencies (3 and 8 cases, respectively), carnitine palmitoyl transferase type II/translocase deficiencies (6 cases), severe carnitine deficiency (4 cases), isovaleric acidemia/2-methylbutyryl-CoA dehydrogenase deficiencies (4 cases), and long-chain hydroxyacyl-CoA dehydrogenase/trifunctional protein deficiencies (4 cases). Conclusions: Postmortem metabolic screening can explain deaths in infants and children and provide estimates of the number of infant deaths attributable to inborn errors of metabolism. MS/MS is cost-effective for analysis of postmortem specimens and should be considered for routine use by Medical Examiners and pathologists in unexpected/unknown infant and child death.