Rezapkin Gennady, Martin Javier, Chumakov Konstantin
Center for Biologics Evaluation and Research, Food and Drug Administration, 1401 Rockville Pike, HFM 470, Rockville, MD 20852, USA.
Biologicals. 2005 Mar;33(1):29-39. doi: 10.1016/j.biologicals.2004.11.001. Epub 2005 Jan 27.
A new block-ELISA test for quantitative evaluation of relative reactivity of antigenic sites was developed and used to reveal the detailed epitope structure of inactivated poliovirus vaccines (IPV) and live poliovirus strains. Poliovirus was captured on ELISA plates coated with rabbit anti-poliovirus IgG and blocked by monoclonal antibodies (Mabs) specific to individual epitopes before the remaining reactive antigenic sites were quantified by polyclonal anti-poliovirus IgG conjugate. The decrease of conjugate binding by the pre-treatment with a Mab reflects its contribution to the overall reactivity of poliovirus antigen. The level of block activity of Mabs for a given antigen can be expressed as a percent of reduction of antigenic reactivity as determined by ELISA test. It can be normalized by expressing this value as a ratio to the block activity of a reference sample. The data on the blocking-activity of a panel of monoclonal antibodies specific to different antigenic sites represents the epitope composition (antigenic profile) of a sample. Quantitative differences in epitope composition were determined for nine samples of inactivated poliovirus vaccine (IPV) and compared with the International Reference Reagent. This method could be used for monitoring consistency of IPV production, comparison of vaccines made by different manufacturers, and for the analysis of antigenically modified strains of attenuated poliovirus. Antigenic structures of two isolates of type 1 vaccine-derived poliovirus (VDPV) were compared with the structures of parental Sabin 1 and wild-type Mahoney strains using 17 monoclonal antibodies and revealed significant differences, suggesting that the method can be used for screening of field isolates and rapid identification of antigenically divergent VDPV strains.
开发了一种用于定量评估抗原位点相对反应性的新型阻断酶联免疫吸附测定（block-ELISA）试验，并用于揭示灭活脊髓灰质炎病毒疫苗（IPV）和脊髓灰质炎病毒活毒株的详细表位结构。脊髓灰质炎病毒被捕获在包被有兔抗脊髓灰质炎病毒IgG的酶联免疫吸附测定板上，并在通过多克隆抗脊髓灰质炎病毒IgG缀合物对其余反应性抗原位点进行定量之前，用针对各个表位特异的单克隆抗体（Mab）进行封闭。用一种单克隆抗体进行预处理后缀合物结合的减少反映了其对脊髓灰质炎病毒抗原总体反应性的贡献。给定抗原的单克隆抗体的封闭活性水平可以表示为通过酶联免疫吸附测定试验确定的抗原反应性降低的百分比。通过将该值表示为与参考样品的封闭活性的比率，可以对其进行标准化。一组针对不同抗原位点的单克隆抗体的封闭活性数据代表了样品的表位组成（抗原谱）。确定了九份灭活脊髓灰质炎病毒疫苗（IPV）样品表位组成的定量差异，并与国际参考试剂进行了比较。该方法可用于监测IPV生产的一致性、比较不同制造商生产的疫苗，以及分析减毒脊髓灰质炎病毒的抗原修饰毒株。使用17种单克隆抗体比较了两株1型疫苗衍生脊髓灰质炎病毒（VDPV）分离株的抗原结构与亲本Sabin 1和野生型Mahoney毒株的结构，结果显示存在显著差异，这表明该方法可用于筛选现场分离株并快速鉴定抗原性不同的VDPV毒株。
