BACKGROUND Newborn screening (NBS) laboratories in the United Kingdom adhere to common protocols based on single analyte cutoff values (COVs); therefore, interlaboratory harmonization is of paramount importance. Interlaboratory variation for screening analytes in UK NBS laboratories ranges from 17% to 59%. While using common stable isotope internal standards has been shown to significantly reduce interlaboratory variation, instrument set-up, sample extraction, and calibration approach are also key factors. METHODS Dried blood spot (DBS) extraction processes, instrument set-up, mobile-phase composition, sample introduction technique, and calibration approach of flow injection analysis-tandem mass spectrometry (FIA-MS/MS) methods were optimized. Inter- and intralaboratory variation of methionine, leucine, phenylalanine, tyrosine, isovaleryl-carnitine, glutaryl-carnitine, octanoyl-carnitine, and decanoyl-carnitine were determined pre- and postoptimization, using 3 different calibration approaches. RESULTS Optimal recovery of analytes from DBS was achieved with a 35-min extraction time and 80% methanol (150 μL). Optimized methodology decreased the mean intralaboratory percentage relative SD (%RSD) for the 8 analytes from 20.7% (range 4.1-46.0) to 5.4% (range 3.0-8.5). The alternative calibration approach reduced the mean interlaboratory %RSD for all analytes from 16.8% (range 4.1-25.0) to 7.1% (range 4.1-11.0). Nuclear magnetic resonance analysis of the calibration material highlighted the need for standardization. The purities of isovaleryl-carnitine and glutaryl-carnitine were 85.13% and 69.94% respectively, below the manufacturer's stated values of ≥98%. CONCLUSIONS For NBS programs provided by multiple laboratories using single analyte COVs, harmonization and standardization of results can be achieved by optimizing legacy FIA-MS/MS methods, adopting a common analytical protocol, and using standardized calibration material rather than internal calibration.

中文翻译：

---

通过优化传统流动注射分析串联质谱方法和应用标准化校准方法，提高扩大新生儿筛查结果的协调性和标准化。

背景 英国的新生儿筛查 (NBS) 实验室遵循基于单一分析物截止值 (COV) 的通用协议。因此，实验室间的协调至关重要。英国 NBS 实验室筛选分析物的实验室间变异范围为 17% 至 59%。虽然使用常见的稳定同位素内标已被证明可以显着减少实验室间的差异，但仪器设置、样品提取和校准方法也是关键因素。方法 对干血斑 (DBS) 提取工艺、仪器设置、流动相组成、样品引入技术和流动注射分析-串联质谱 (FIA-MS/MS) 方法的校准方法进行了优化。蛋氨酸、亮氨酸、苯丙氨酸、酪氨酸、异戊酰肉碱、戊二酰肉碱、辛酰肉碱和癸酰肉碱在优化前和优化后使用 3 种不同的校准方法进行测定。结果 在 35 分钟的提取时间和 80% 甲醇 (150 μL) 的条件下，从 DBS 中实现了分析物的最佳回收率。优化的方法将 8 种分析物的平均实验室内相对 SD (%RSD) 从 20.7%（范围 4.1-46.0）降低到 5.4%（范围 3.0-8.5）。替代校准方法将所有分析物的平均实验室间 %RSD 从 16.8%（范围 4.1-25.0）降低到 7.1%（范围 4.1-11.0）。校准材料的核磁共振分析强调了标准化的必要性。异戊酰肉碱和戊二酰肉碱的纯度分别为 85.13% 和 69.94%，低于制造商规定的≥98% 的值。