Evaluation of a Common Internal Standard Material to Reduce Inter-Laboratory Variation and Ensure the Quality, Safety and Efficacy of Expanded Newborn Screening Results When Using Flow Injection Analysis Tandem Mass Spectrometry with Internal Calibration

Abstract

In 2015, the newborn screening (NBS) programmes in England and Wales were expanded to include four additional disorders: Classical Homocystinuria, Isovaleric Acidemia, Glutaric Aciduria Type 1 and Maple Syrup Urine Disease, bringing the total number of analytes quantified to eight: phenylalanine, tyrosine, leucine, methionine, isovalerylcarnitine, glutarylcarnitine, octanoylcarnitine and decanoylcarnitine. Post-implementation, population data monitoring showed that inter-laboratory variation was greater than expected, with 90th centiles varying from 17% to 59%. We evaluated the effect of stable isotope internal standard (IS) used for quantitation on inter-laboratory variation. Four laboratories analysed routine screening samples (n > 101,820) using a common IS. Inter-laboratory variation was determined for the eight analytes and compared with results obtained using an in-house common IS (n > 102,194). A linear mixed-effects model was fitted to the data. Using a common IS mix reduced the inter-laboratory variation significantly (p < 0.05) for five analytes. For three analytes, the lack of significance was explained by use of individual laboratory “calibration factors”. For screening programmes where laboratories adhere to single analyte cut-off values (COVs), it is important that inter-laboratory variation is minimised, primarily to prevent false positive results. Whilst the use of a common IS helps achieve this, it is evident that instrument set-up also contributes to inter-laboratory variation.

Improving Harmonization and Standardization of Expanded Newborn Screening Results by Optimization of the Legacy Flow Injection Analysis Tandem Mass Spectrometry Methods and Application of a Standardized Calibration Approach

“…The postoptimization experiments were performed using a simulated sample (protein-precipitated plasma spiked to achieve analyte concentrations approximating the COV), thus negating differences due to sample extraction. Prior to optimization of the method, it was evident that both intra-and interinstrument variability were greater than expected (4). From a practical perspective, the intrainstrument variability at the COV was large enough to potentially result in a false-negative or falsepositive result (e.g., preoptimization, the %RSD is 46% for C5, 24.3% for Leu, and 22.2% for C5DC (Fig.…”

Current State and Innovations in Newborn Screening: Continuing to Do Good and Avoid Harm

“…Currently, there are no commercially available, DBS-based external calibrators. Recent studies have demonstrated that inter-laboratory variation can be significantly reduced using a common stable isotope [ 28 ] and by retrospectively harmonizing screening results to a common material [ 29 ].…”

“…A major limitation of the utility of DBS specimens for biomarker analysis is the lack of commercially available, matrix-matched certified reference materials (CRMs) for the various analytes in DBS specimens with which to standardise laboratory tests [ 27 , 28 ]. As a result, DBS calibrators tend to be produced in-house by collecting blood from a healthy donor or using residual pooled patient samples and adding an aqueous enrichment prior to application onto the filter paper.…”