Phillips Donald J, League Stacy C, Weinstein Paula, Hooper W Craig
Centers for Disease Control and Prevention, National Center on Birth Defects and Development Disabilities, Mail Stop D02, 1600 Clifton Rd., NE, Atlanta, GA 30333, USA.
Cytokine. 2006 Nov;36(3-4):180-8. doi: 10.1016/j.cyto.2006.12.002. Epub 2007 Feb 15.
The present study describes positive and negative interference of human cytokine measurement in multiplexed bead-based immunoassays. Significant differences in measured IL-6 and TNF-alpha values in 30 normal human plasma samples were apparent depending on whether measurements were with a 2-plex assay or embedded in a multiplex of 8-or more cytokine antibody pairs, as well as among the kits of 3-different vendors. Sample diluents containing proprietary blocking ingredients were shown to greatly affect the outcome of measured cytokine values. Additionally, recovery of IL-6 and TNF-alpha from spiked samples suggests significant negative interference from either endogenous antibodies, soluble receptors or anti-cytokine antibodies in 10% and 26% of samples, respectively. While it is evident that multiplexed immunoassays hold great promise for cytokine profiling, there are still important issues needing further study. Especially needed are universally optimized sample diluents, uniformly calibrated standards with mass values, and internal assay controls, which should greatly facilitate intralaboratory accuracy and precision and interlaboratory comparisons of cytokine measurements. Possible causes of interference and remedies are discussed.
本研究描述了基于微珠的多重免疫测定中人类细胞因子测量的正干扰和负干扰。30份正常人血浆样本中白细胞介素-6（IL-6）和肿瘤坏死因子-α（TNF-α）的测量值存在显著差异，这取决于测量是采用双靶点检测法，还是嵌入包含8种或更多细胞因子抗体对的多重检测法，以及不同供应商的试剂盒之间的差异。含有专利封闭成分的样本稀释剂被证明会极大地影响细胞因子测量值的结果。此外，加标样本中IL-6和TNF-α的回收率表明，分别有10%和26%的样本受到内源性抗体、可溶性受体或抗细胞因子抗体的显著负干扰。虽然多重免疫测定在细胞因子分析方面具有很大的前景，但仍有一些重要问题需要进一步研究。特别需要的是通用优化的样本稀释剂、具有质量值的统一校准标准品以及内部检测对照，这将极大地促进实验室内部细胞因子测量的准确性和精密度以及实验室间的比较。本文讨论了干扰的可能原因及补救措施。
