Vaccine Research and Development and Early Clinical Development Biostatistics, Pfizer Inc., Pearl River, New York, USA
Vaccine Research and Development and Early Clinical Development Biostatistics, Pfizer Inc., Pearl River, New York, USA.
mSphere. 2018 Aug 8;3(4):e00128-18. doi: 10.1128/mSphere.00128-18.
A Luminex-based direct immunoassay (dLIA) platform has been developed to replace the standardized pneumococcal enzyme-linked immunosorbent assay platform. The multiplex dLIA simultaneously measures the concentration of serum immunoglobulin G (IgG) antibodies specific for pneumococcal capsular polysaccharide (PnPS) serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F, and 23F. The assay uses poly-l-lysine (PLL)-conjugated PnPS, chemically coupled to spectrally distinct Luminex microspheres. Assay validation experiments were performed using residual human serum samples obtained from 13-valent pneumococcal conjugate vaccine (13vPnC) clinical studies. Assay results are expressed as IgG antibody concentrations in micrograms per milliliter using the international reference serum, 007sp. The lower limit of quantitation (LLOQ) for all serotypes covered in the 13-plex dLIA fell within the range of 0.002 to 0.038 µg/ml serum IgG. The difference between the lower limit and upper limit of the assay range was >500-fold for all serotypes, and assay variability was <20% relative standard deviation (RSD) for all serotypes. IgG antibody measurements were shown to be serotype-specific (some cross-reactivity was observed only between the structurally related serotypes 6A and 6B as well as 19A and 19F), and no interference was observed between the serotypes when the assay was performed in the 13-plex format compared to the singleplex assays. The 13-plex dLIA platform developed by Pfizer Inc. generates up to 143 test results in a single 96-well plate and is a suitable replacement of the enzyme-linked immunosorbent assay (ELISA) platform for evaluating vaccine clinical trials. The pneumococcal enzyme-linked immunosorbent assay (ELISA) measures IgG antibodies in human serum, and it is an important assay that supports licensure of pneumococcal vaccines. The immune correlate of protection, 0.35 µg/ml of IgG antibodies, was determined by the ELISA method. Pfizer has developed a new Luminex-based assay platform to replace the ELISA. These papers describe the important work of (i) validating the Luminex-based assay and (ii) bridging the immune correlate of protection (0.35 µg/ml IgG) to equivalent values reported by the Luminex platform.
一种基于 Luminex 的直接免疫测定（dLIA）平台已经开发出来，以替代标准化的肺炎球菌酶联免疫吸附测定平台。该多重 dLIA 同时测量针对肺炎球菌荚膜多糖（PnPS）血清型 1、3、4、5、6A、6B、7F、9V、14、18C、19A、19F 和 23F 的血清免疫球蛋白 G（IgG）抗体的浓度。该测定使用多聚-L-赖氨酸（PLL）缀合的 PnPS，化学偶联到光谱上明显不同的 Luminex 微球上。使用从 13 价肺炎球菌结合疫苗（13vPnC）临床研究中获得的剩余人血清样本进行了测定验证实验。测定结果以国际参考血清 007sp 为单位表示为每毫升血清 IgG 的微克数。13 重 dLIA 中涵盖的所有血清型的定量下限（LLOQ）均在 0.002 至 0.038 µg/ml 血清 IgG 范围内。所有血清型的测定范围下限和上限之间的差异均超过 500 倍，所有血清型的测定变异性均<20%相对标准偏差（RSD）。IgG 抗体测量结果表明具有血清型特异性（仅在结构相关的血清型 6A 和 6B 以及 19A 和 19F 之间观察到一些交叉反应），与单重测定相比，当在 13 重格式下进行测定时，各血清型之间没有观察到干扰。辉瑞公司开发的 13 重 dLIA 平台在单个 96 孔板中可产生多达 143 个测试结果，是评估疫苗临床试验的酶联免疫吸附测定（ELISA）平台的合适替代品。肺炎球菌酶联免疫吸附测定（ELISA）测量人血清中的 IgG 抗体，是支持肺炎球菌疫苗许可的重要测定。保护的免疫相关性为 0.35 µg/ml IgG 抗体，由 ELISA 方法确定。辉瑞公司开发了一种新的基于 Luminex 的测定平台来替代 ELISA。这些论文描述了重要的工作：（i）验证基于 Luminex 的测定和（ii）将保护免疫相关性（0.35 µg/ml IgG）桥接到 Luminex 平台报告的等效值。
