Introduction

Commercially available enzyme-linked immunosorbent assay (ELISA) kits designed for pertussis diagnostic purposes are frequently used to assess antibody responses to pertussis vaccines in clinical trials, but have limited accuracy and are not calibrated against international standards. We developed a new electrochemiluminescence (ECL)-based multiplexed assay and compared its performance to two commercial Bordetella pertussis ELISA kits and to historical in-house ELISAs.

Methods

The ECL assay quantifies serum concentrations of antibodies against four B. pertussis antigens: pertussis toxin (PT), filamentous hemagglutinin (FHA), pertactin (PRN), and fimbrial agglutinogen (FIM). The assay was validated for precision, accuracy, dilutability, lower limit of quantification, and specificity. Sera from a clinical trial (CTRI/2016/11/007434) were used to compare the ECL assay to two commercial ELISA kits available from GenWay BioTech and Demeditec Diagnostics for accuracy, linearity, specificity, and concordance to both internal (WWO-2-043) and international (NIBSC 06/140) references. Sera from four clinical trials (NCT02587520, NCT00255047, NCT00347958, NCT01346293) were used to compare the concordance to clinical ELISAs. Informed consent was ensured prior to using any sera.

Results

Precision, accuracy, dilutability, lower limit of quantification, and specificity were demonstrated for the ECL assay. Concordance between the ECL assay and established clinical ELISAs was met for antibody responses to PT, FIM, and PRN, but not for FHA. The ECL assay demonstrated higher accuracy and linearity than the ELISA kits. While concordance between the ECL and commercial kits was low, the ECL assay better distinguished between pre- and post-vaccination clinical samples.

Conclusion

The new ECL assay was validated for the quantitative evaluation of anti-PT, anti-FHA, anti-FIM, and anti-PRN IgG antibodies in samples from clinical trials, and demonstrated equivalent or better performance than two commercially available ELISA kits.

中文翻译：

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一种新的基于电化学发光的多重检测，用于评估人类对百日咳博德特氏菌疫苗的抗体反应

介绍

专为百日咳诊断目的而设计的市售酶联免疫吸附测定 (ELISA) 试剂盒经常用于评估临床试验中对百日咳疫苗的抗体反应，但准确性有限且未根据国际标准进行校准。我们开发了一种新的基于电化学发光 (ECL) 的多重检测，并将其性能与两种商业百日咳博德特氏菌ELISA 试剂盒和历史悠久的内部 ELISA 进行了比较。

方法

ECL 测定可量化针对四种百日咳杆菌抗原的抗体的血清浓度：百日咳毒素 (PT)、丝状血凝素 (FHA)、百日咳蛋白 (PRN) 和菌毛凝集原 (FIM)。该测定在精密度、准确度、稀释性、定量下限和特异性方面得到验证。来自临床试验 (CTRI/2016/11/007434) 的血清用于比较 ECL 检测与 GenWay BioTech 和 Demeditec Diagnostics 提供的两种商业 ELISA 试剂盒的准确性、线性、特异性以及与两者内部 (WWO-2- 043) 和国际 (NIBSC 06/140) 参考。来自四项临床试验（NCT02587520、NCT00255047、NCT00347958、NCT01346293）的血清用于比较与临床ELISA的一致性。在使用任何血清之前确保知情同意。

结果

ECL 分析证明了精密度、准确度、可稀释性、定量下限和特异性。对 PT、FIM 和 PRN 的抗体反应符合 ECL 检测和已建立的临床 ELISA 之间的一致性，但不符合 FHA。ECL 检测显示出比 ELISA 试剂盒更高的准确度和线性。虽然 ECL 和商业试剂盒之间的一致性较低，但 ECL 检测可以更好地区分疫苗接种前和疫苗接种后的临床样本。

结论

新的 ECL 检测方法经验证可用于定量评估临床试验样本中的抗 PT、抗 FHA、抗 FIM 和抗 PRN IgG 抗体，并证明其性能与两种市售 ELISA 试剂盒相当或更好。