Heterophile antibody interference in a multiplexed fluorescent microsphere immunoassay for quantitation of cytokines in human serum
Heterophile antibody interference in a multiplexed fluorescent microsphere immunoassay for quantitation of cytokines in human serum
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DOI：
10.1128/cdli.11.2.325-329.2004
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发表时间：
2004-03-01
期刊：
影响因子：
--
通讯作者：
Hill, HR
中科院分区：
文献类型：
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作者：
Martins, TB;Pasi, BM;Hill, HR
虽然现代免疫测定法提供了用于定量生物流体中细胞因子的灵敏和特异性手段，但嗜异性抗体仍然是这些测定法中细胞因子测量的公认干扰原因。我们已经开发了一种多重荧光微球免疫测定法，仅在75穆尔血清中同时定量10种细胞因子。在该多重测定的开发过程中，还测定了嗜异性抗体引起的测定干扰量，并将检测嗜异性干扰和最小化其影响的方法纳入测定中。与正常血库样本相比，嗜异性抗体导致细胞因子值显著升高。通过使用内部对照鉴别这些假性升高的值，从而鉴别嗜异性抗体结合的试验组分。然后使用该信息设计试验特异性阻断剂和吸收剂，这些阻断剂和吸收剂显示可显著降低假性升高的细胞因子值，同时不影响标准品和对照品值。基于荧光多重微球的免疫测定可用于从单个样品中定量多种细胞因子，并且应该是进一步理解细胞因子在疾病过程中的作用的有用工具。
While modern immunoassays provide sensitive and specific means for the quantitation of cytokines in biological fluids, heterophile antibodies are still a well-recognized cause of interference in the measurement of cytokines in these assays. We have developed a multiplexed fluorescent microsphere immunoassay for the simultaneous quantification of 10 cytokines in only 75 mul of serum. During the development of this multiplexed assay, the amount of assay interference due to heterophile antibodies was also determined, and methods for detecting heterophile interference and minimizing its effect were incorporated into the assay. Heterophile antibodies resulted in significantly elevated cytokine values compared to those of normal blood bank samples. These falsely elevated values, and thus the components of the assay the heterophile antibodies were binding to, were identified through the use of internal controls. This information was then used to design assay-specific blockers and absorbents that were shown to significantly reduce falsely elevated cytokine values while not affecting the standard and control values. The fluorescent multiplexed microsphere-based immunoassay can be used to quantitate multiple cytokines from a single sample and should be a useful tool in furthering our understanding of the role of cytokines in disease processes.