Reactivity of human sera in a sensitive, high-throughput pseudovirus-based papillomavirus neutralization assay for HPV16 and HPV18
Reactivity of human sera in a sensitive, high-throughput pseudovirus-based papillomavirus neutralization assay for HPV16 and HPV18
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DOI：
10.1016/j.virol.2003.12.027
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发表时间：
2004-04-10
期刊：
影响因子：
3.7
通讯作者：
Schiller, JT
中科院分区：
文献类型：
--
作者：
Pastrana, DV;Buck, CB;Schiller, JT
基于携带分泌碱性磷酸酶（SEAP）报告基因的假病毒，开发并验证了对人乳头状病毒（HPV）16、HPV18和牛乳头状瘤病毒1 （BPV1）的敏感的高通量中和试验。将密码子修饰的乳头状病毒结构基因瞬时转染到293细胞的SV40 T抗原表达系中产生SEAP假病毒，在一个烧瓶中产生足够的假病毒，可进行数千次滴定。在96孔板格式的初步表征中，该分析具有可重复性，并且似乎与标准的基于乳头状病毒样颗粒（VLP）的酶联免疫吸附试验（ELISA）一样敏感，但特异性更高。中和试验在感染相应HPV型别的大多数妇女血清中检测到HPV16或HPV18中和抗体（滴度为160-10240），但未在处女中检测到。HPV16 VLP疫苗接种者血清抗HPV16中和效价高（平均：45000；范围：5120-163840），但无抗hpv18中和活性。基于SEAP假病毒的中和试验应该是一种实用的方法，用于量化HPV自然史和预防性疫苗研究中潜在的保护性抗体反应。Elsevier Inc.出版。
Sensitive high-throughput neutralization assays, based upon pseudoviruses carrying a secreted alkaline phosphatase (SEAP) reporter gene, were developed and validated for human papillornavirus (HPV)16, HPV18, and bovine papillomavirus 1 (BPV1). SEAP pseudoviruses were produced by transient transfection of codon-modified papillornavirus structural genes into an SV40 T antigen expressing line derived from 293 cells, yielding sufficient pseudovirus from one flask for thousands of titrations. In a 96-well plate format, in this initial characterization, the assay was reproducible and appears to be as sensitive as, but more specific than, a standard papillornavirus-like particle (VLP)-based enzyme-linked immunosorbent assay (ELISA). The neutralization assay detected type-specific HPV16 or HPV18 neutralizing antibodies (titers of 160-10240) in sera of the majority of a group of women infected with the corresponding HPV type, but not in virgin women. Sera from HPV16 VLP vaccinees had high anti-HPV16 neutralizing titers (mean: 45000; range: 5120-163840), but no anti-HPV18 neutralizing activity. The SEAP pseudovirus-based neutralization assay should be a practical method for quantifying potentially protective antibody responses in HPV natural history and prophylactic vaccine studies. Published by Elsevier Inc.