Numis Adam L, Fox Christine H, Lowenstein Daniel J, Norris Philip J, Di Germanio Clara
University of California, San Francisco, Department of Neurology & Pediatrics, 675 Nelson Rising Lange, San Francisco, CA 94158 USA.
Department of Neurology, University of California, San Francisco USA.
Heliyon. 2021 Mar 12;7(3):e06445. doi: 10.1016/j.heliyon.2021.e06445. eCollection 2021 Mar.
Multiplex analyses allow for detection of dozens of cytokines/chemokines in small sample volumes. Although several commercially available assay kits are available, there are no comparative data in plasma measurements among pediatric or epilepsy cohorts.
Cohort study of 38 children with epilepsy. We evaluated plasma levels of cytokines/chemokines using three different assays: Luminex® xMAP high-sensitivity (HS) and standard-sensitivity (SS) assays, and Meso-Scale Discovery (MSD). We calculated recovery rates of each analyte, correlation coefficients between assays, and level of agreement between measurements. We repeated analyses in a subset of samples after a single freeze-thaw cycle.
Among ten analytes common to all assays, HS had high recovery (<15% of values extrapolated or out-of- range [OOR]) for all analytes, SS for 50%, and MSD for 40%. While several analytes had a high correlation between assays, Bland-Altman plots demonstrated assays were not interchangeable. For most analytes, a single freeze-thaw cycle decreased cytokines/chemokine measurements. There was good correlation of measurements after a freeze-thaw cycle with acceptable agreement between measurements for six of 13 (46%) analytes using HS, one of 9 (11%) for SS, and none for MSD.
HS assays may optimize yield in plasma for proteins of particular interest in epilepsy research, limit values extrapolated beyond the standard curve, and improve precision compared to other SS and MSD assays.
Our results demonstrate assay choice may be critical to study results and support the need for a standardized approach to biomarker assessment across epilepsy research and other domains.
多重分析能够在小样本量中检测数十种细胞因子/趋化因子。尽管有几种市售检测试剂盒，但在儿科或癫痫队列的血浆测量中尚无比较数据。
对38名癫痫患儿进行队列研究。我们使用三种不同检测方法评估细胞因子/趋化因子的血浆水平：Luminex® xMAP高灵敏度（HS）和标准灵敏度（SS）检测方法，以及Meso-Scale Discovery（MSD）检测方法。我们计算了每种分析物的回收率、检测方法之间的相关系数以及测量值之间的一致性水平。在单次冻融循环后，我们在一部分样本中重复进行分析。
在所有检测方法共有的1种分析物中，HS对所有分析物的回收率都很高（外推值或超出范围[OOR]的值<15%），SS为50%，MSD为40%。虽然几种分析物在检测方法之间具有高度相关性，但Bland-Altman图表明这些检测方法不可互换。对于大多数分析物，单次冻融循环会降低细胞因子/趋化因子的测量值。冻融循环后的测量值之间具有良好的相关性，13种（46%）分析物中有6种使用HS时测量值之间的一致性可接受，9种（11%）中有1种使用SS时可接受，而使用MSD时则没有。
与其他SS和MSD检测方法相比，HS检测方法可能会优化癫痫研究中特定感兴趣蛋白质在血浆中的产量，限制超出标准曲线的外推值，并提高精密度。
我们的结果表明检测方法的选择可能对研究结果至关重要，并支持在癫痫研究和其他领域中采用标准化方法进行生物标志物评估的必要性。
