Objectives: Re-evaluation of the newborn screening for cblC/D defects based on the analysis of false positive and negative rates. Methods: Extended newborn screening was implemented in 2004 and 544,675 newborns were screened by MS/MS. Sequence analysis of genomic DNAwas performed to identify disease-causing mutations in MMACHC and MMADHC. Results: We identified five true-positive cblC defect cases (1/108,935) , one false-negative cblD defect case (1/544,675) and 248 false-positives. All cblC patients showed the most common MMACHC mutation-c.271dupA. The molecular study of cblD patient, a girl identified at 8-month-old, revealed p.R250X mutation in MMADHC gene. At screening time, acylcarnitines presented C3=6.1 μM (cut-off >6.0); Met=15.2 μM (cut-off>12) and C3/Met=0.40; C3/C2=0.35; and C3/C16=2.0. Discussion/Conclusion: Our results revealed that cblC/D defects are important conditions to be screened in our population. At sampling day, normal Met value associated with a borderline C3 value, lead to the misclassification of the cblD-patient as a normal-case. Until now, it was not possible to implement the second-tier tests, but in the future we intent to use this strategy to achieve a better assessment in the screening of these defects. Results: From 1999 to 2007 our recall rate for elevated C3-carnitine or ratio was 217 of 154,000 neonates (0.13%). Retrospective MMA testing of residual samples from these infants detected 19 with elevated MMA, reducing the predicted recall rate to 0.06%. To date 1,200 further MMA tests have been performed, and 48 neonates recalled for a second sample if MMA was above the 99th centile (recall rate below 0.025). 12 had persistent elevated MMA and were referred for clinical assessment. Of these, 11 infants were diagnosed with B12 deficiency and 1 was confirmed to have Cobalamin A deficiency (Dr B Fowler). Conclusion: The incorporation of second tier MMA determination into our routine newborn screening program continues to significantly reduce the false positive rate associated with the measurement of C3-carnitine and ratios. Glutaric Aciduria type 1 (GA1) is a rare disease affecting lysine, hydroxylysine, and tryptophan metabolism. Classically reported presentations include acute childhood encephalopathy precipitated by metabolic stress. However, there is a subset of individuals with GA1 who may remain asymptomatic into adulthood.