Production of furin-cleaved Papillomavirus pseudovirions and their use for in vitro neutralization assays of L1-or L2-specific antibodies

Joshua W. Wang, Kihyuck Kwak, Shiwen Peng, Troy Kemp, Ligia Pinto, Richard B.S. Roden

Research output: Contribution to journalArticle

Abstract

Immunization with Human Papillomavirus (HPV) L1 virus-like particles or L2 capsid protein elicits neutralizing antibodies that mediate protection. A highthroughput and sensitive in vitro neutralization assay is therefore valuable for prophylactic HPV vaccine studies. Over several hours during infection of the genital tract, virions take on a distinct intermediate conformation, including a required furin cleavage of L2 at its N-terminus. This intermediate is an important target for neutralization by L2-specific antibody, but it is very transiently exposed during in vitro infection of most cell lines resulting in insensitive measurement for L2, but not L1-specific neutralizing antibodies. To model this intermediate, we describe a protocol to generate furin-cleaved HPV pseudovirions (fc-PsV), which deliver an encapsidated reporter plasmid to facilitate infectivity measurements. We also describe a protocol for use of fc-PsV in a high-throughput in vitro neutralization assay for the sensitive measurement of both L1 and L2-specific neutralizing antibodies.

Original languageEnglish (US)
Pages (from-to)14B.5.1-14B.5.26
JournalCurrent Protocols in Microbiology
Volume2015
DOIs
StatePublished - Jan 1 2015

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Keywords

  • Antibody
  • Furin-cleavage
  • HPV neutralization assay
  • Hpv l2
  • Human papillomavirus
  • Papillomavirus

ASJC Scopus subject areas

  • Parasitology
  • Microbiology
  • Virology

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