Development of a chemiluminescence enzyme immunoassay (CLEIA) for quantitating L1 protein in HPV vaccines

The L1 protein serves as the principal capsid component of human papillomavirus (HPV). All globally commercialized HPV vaccines utilize virus-like particles (VLPs) formed through L1 protein self-assembly. Quantitative analysis of L1 protein concentration constitutes a critical parameter for evaluating the antigenic potency of HPV vaccines. In this study, we developed a chemiluminescent enzyme immunoassay (CLEIA) for precise quantification of L1 protein in bivalent HPV vaccines. Employing a sandwich immunoassay format, antigen-antibody complexes were immobilized on 96-well microplates using capture antibodies, followed by detection with HRP-conjugated secondary antibodies. Chemiluminescent signal amplification was achieved through enzymatic catalysis of luminol/hydrogen peroxide in the presence of enhancer molecules. Systematic optimization of experimental parameters yielded a validated methodology demonstrating excellent reproducibility (inter-assay CV < 4 %), accuracy (recovery rate 100.5 ± 2.8 % for 16L1 and 95.5 ± 6.4 % for 18L1), precision (inter-assay CV < 7 %) and sensitivity (limit of quantitation (LOQ) 0.0996 microg/mL for HPV16L1 and 0.1459 microg/mL for HPV18L1). This optimized assay provides a reliable analytical platform for quantitating L1 Protein in bivalent HPV vaccine.