HPV Immunology Laboratory, SAIC-Frederick, Inc., NCI-Frederick, Frederick, MD 21702, USA.
Mol Cell Probes. 2012 Apr;26(2):73-80. doi: 10.1016/j.mcp.2012.01.002. Epub 2012 Jan 18.
Antibody responses against infectious agents are an important component in the prevention of disease. The avidity of antibodies for their antigens relates to their functional efficiency, and is a fundamental aspect in the investigation of humoral responses. Modified ELISAs are used to estimate avidity through the use of chaotropic agents and the measurement of the degree to which they disrupt the interaction between antibody and antigen. The theory behind the assay is the higher the avidity of an interaction the less susceptible it is to the effects of the chaotropic agent. The goal of this study was to generate a modified ELISA where a complex, multimeric coating-antigen, human papillomavirus (HPV) virus-like particles (VLP), was used to measure the avidity of anti-HPV antibodies generated following vaccination with HPV VLPs. A series of chaotropic agents were evaluated in the assay for their effectiveness in measuring avidity. Guanidine hydrochloride (GuHCl) was selected as a chaotropic reagent with the ability to disrupt antibody and antigen interactions, while not affecting the integrity of the plate-bound VLP. Two methods of determining the avidity index were assessed and shown to be comparable. This assay was then successfully applied to measure the avidity of anti-HPV VLP serum antibodies in samples from an HPV L1 VLP vaccine clinical trial. Overall, the assay was highly reproducible and captured a wide range of antibody avidities. Therefore, a GuHCl-modified ELISA is an acceptable method that can be used to determine HPV-specific antibody avidity indices within a clinical trial setting.
针对传染性病原体的抗体反应是预防疾病的重要组成部分。抗体对其抗原的亲和力与其功能效率有关，是体液免疫反应研究的一个基本方面。改良的 ELISA 通过使用离液剂和测量它们对抗体和抗原之间相互作用的破坏程度来估计亲和力。该测定背后的理论是，相互作用的亲和力越高，它对离液剂的影响就越不敏感。本研究的目的是生成一种改良的 ELISA，其中使用复杂的多聚体包被抗原，即人乳头瘤病毒 (HPV) 病毒样颗粒 (VLP)，来测量 HPV VLP 疫苗接种后产生的抗 HPV 抗体的亲和力。评估了一系列离液剂以确定其在测定亲和力方面的有效性。盐酸胍 (GuHCl) 被选为一种离液剂，具有破坏抗体和抗原相互作用的能力，同时不影响板结合 VLP 的完整性。评估了两种确定亲和力指数的方法，结果表明它们是可比的。该测定法随后成功地应用于测量 HPV L1 VLP 疫苗临床试验中 HPV VLP 血清抗体的亲和力。总体而言，该测定法具有高度的可重复性，并能捕捉到广泛的抗体亲和力范围。因此，GuHCl 改良的 ELISA 是一种可接受的方法，可用于在临床试验环境中确定 HPV 特异性抗体的亲和力指数。
