Well-characterized HPV serology assays are required to evaluate performance of biosimilar candidate vaccines, reduced dosing schedules and novel administration methods. We report characterization of an expanded assay, M9ELISA, that detects antibodies to HPV virus-like particles (VLP) of nine types using direct IgG ELISA on the Meso Scale Discovery (MSD) electrochemiluminescence platform. The method is based on the previously published M4ELISA which detects antibodies to HPV6,11,16, and 18. It has been modified to add detection of antibodies to HPV31,33,45,52 and 58, and to streamline assay and reduce background.

The M9ELISA plates were prepared with purified type specific L1 + L2 VLPs coated on 10-spot/well standard MSD microplates. Results of ELISA on three serial dilutions of serum were read on MSD imager, and titers calculated using the parallel line method. Evaluations included dynamic range, assay reproducibility, and stability over time. We compared M9ELISA results to those from a pseudovirion-based neutralization assay in sera from a mixed cohort of unvaccinated and vaccinated individuals (n = ~116) and to competitive Luminex immunoassay (cLIA) results in sera from a predominantly unvaccinated cohort (n = 4426).

The linear range of the assay extended over 5 logs, with inter-assay reproducibility coefficient of variation ≤25% for all types. The pre-coated plates were stable for at least 2 years. Spearman correlation of antibody titers showed excellent correlation with PBNA (r = 0.86–0.97) and moderate correlation (r = 0.52–0.68) with cLIA. Thus, the M9ELISA can serve as a useful platform for high-throughput, sensitive and simultaneous quantitation of the antibody responses to nine HPV vaccine types.

需要充分表征的 HPV 血清学检测来评估生物仿制药候选疫苗的性能、减少的给药方案和新颖的给药方法。我们报告了扩展测定法 M9ELISA 的表征，该测定法在 Meso Scale Discovery (MSD) 电化学发光平台上使用直接 IgG ELISA 检测九种类型的 HPV 病毒样颗粒 (VLP) 抗体。该方法基于之前发布的可检测 HPV6、11、16 和 18 抗体的 M4ELISA。该方法经过修改，增加了对 HPV31、33、45、52 和 58 抗体的检测，并简化了检测并降低了背景。

M9ELISA 板是用包被在 10 点/孔标准 MSD 微孔板上的纯化类型特异性 L1 + L2 VLP 制备的。在 MSD 成像仪上读取三种系列血清稀释液的 ELISA 结果，并使用平行线法计算滴度。评估包括动态范围、测定重现性和随时间的稳定性。我们将 M9ELISA 结果与来自未接种疫苗和已接种疫苗的混合队列 (n = ~116) 的血清中基于假病毒粒子的中和测定结果以及来自主要未接种疫苗的队列 ( n = 4426) 的竞争性 Luminex 免疫测定 (cLIA) 结果进行了比较）。

测定的线性范围扩展超过 5 个对数，所有类型的测定间重现性变异系数≤25%。预涂板可稳定至少 2 年。抗体滴度的 Spearman 相关性显示与 PBNA 具有极好的相关性（ r = 0.86-0.97），与 cLIA 具有中等相关性（ r = 0.52-0.68）。因此，M9ELISA 可以作为一个有用的平台，对九种 HPV 疫苗类型的抗体反应进行高通量、灵敏和同步定量。