The purpose of this study was to compare performance metrics of postanalytical interpretive tools of the Region 4 Stork collaborative project to the actual outcome based on cutoff values for amino acids and acylcarnitines selected by the California newborn screening program. This study was a retrospective review of the outcome of 176,186 subjects born in California between 1 January and 30 June 2012. Raw data were uploaded to the Region 4 Stork Web portal as .csv files to calculate tool scores for 48 conditions simultaneously using a previously unpublished functionality, the tool runner. Scores for individual target conditions were deemed informative when equal or greater to the value representing the first percentile rank of known true-positive cases (17,099 cases in total). In the study period, the actual false-positive rate and positive predictive value were 0.26 and 10%, respectively. Utilization of the Region 4 Stork tools, simple interpretation rules, and second-tier tests could have achieved a false-positive rate as low as 0.02% and a positive predictive value >50% by replacing the cutoff system with Region 4 Stork tools as the primary method for postanalytical interpretation. Region 4 Stork interpretive tools, second-tier tests, and other evidence-based interpretation rules could have reduced false-positive cases by up to 90% in California. Genet Med 16 12, 889–895.