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Viruses 2016, 8(4), 107;

Development of a Triple-Color Pseudovirion-Based Assay to Detect Neutralizing Antibodies against Human Papillomavirus

Division of HIV/AIDS and Sexually Transmitted Virus Vaccines, National Institutes for Food and Drug Control (NIFDC), No. 2 Tiantanxili, Beijing 100050, China
These authors contributed equally to this work.
Author to whom correspondence should be addressed.
Academic Editor: Joanna Parish
Received: 28 February 2016 / Revised: 15 April 2016 / Accepted: 15 April 2016 / Published: 25 April 2016
(This article belongs to the Special Issue Replication-Competent Reporter-Expressing Viruses)
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Pseudovirion-based neutralization assay is considered the gold standard method for evaluating the immune response to human papillomavirus (HPV) vaccines. In this study, we developed a multicolor neutralization assay to simultaneously detect the neutralizing antibodies against different HPV types. FluoroSpot was used to interpret the fluorescent protein expression instead of flow cytometry. The results of FluoroSpot and flow cytometry showed good consistency, with R2 > 0.98 for the log-transformed IC50 values. Regardless of the reporter color, the single-, dual-, and triple-color neutralization assays reported identical results for the same samples. In low-titer samples from naturally HPV-infected individuals, there was strong agreement between the single- and triple-color assays, with kappa scores of 0.92, 0.89, and 0.96 for HPV16, HPV18, and HPV58, respectively. Good reproducibility was observed for the triple-color assay, with coefficients of variation of 2.0%–41.5% within the assays and 8.3%–36.2% between the assays. Three triple-color systems, HPV16-18-58, HPV6-33-45, and HPV11-31-52, were developed that could evaluate the immunogenicity of a nonavalent vaccine in three rounds of the assay. With the advantages of an easy-to-use procedure and less sample consumption, the multiple-color assay is more suitable than classical assays for large sero-epidemiological studies and clinical trials and is more amenable to automation. View Full-Text
Keywords: human papillomavirus; HPV; vaccine; FluoroSpot; multiplex immunoassay human papillomavirus; HPV; vaccine; FluoroSpot; multiplex immunoassay

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Nie, J.; Liu, Y.; Huang, W.; Wang, Y. Development of a Triple-Color Pseudovirion-Based Assay to Detect Neutralizing Antibodies against Human Papillomavirus. Viruses 2016, 8, 107.

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