Department of Medicine, Division of Immunology and Rheumatology, Stanford University School of Medicine, Stanford, California, USA.
PLoS One. 2013 May 29;8(5):e64555. doi: 10.1371/journal.pone.0064555. Print 2013.
Existing methods to measure influenza vaccine immunogenicity prohibit detailed analysis of epitope determinants recognized by immunoglobulins. The development of highly multiplex proteomics platforms capable of capturing a high level of antibody binding information will enable researchers and clinicians to generate rapid and meaningful readouts of influenza-specific antibody reactivity.
We developed influenza hemagglutinin (HA) whole-protein and peptide microarrays and validated that the arrays allow detection of specific antibody reactivity across a broad dynamic range using commercially available antibodies targeted to linear and conformational HA epitopes. We derived serum from blood draws taken from 76 young and elderly subjects immediately before and 28±7 days post-vaccination with the 2008/2009 trivalent influenza vaccine and determined the antibody reactivity of these sera to influenza array antigens.
Using linear regression and correcting for multiple hypothesis testing by the Benjamini and Hochberg method of permutations over 1000 resamplings, we identified antibody reactivity to influenza whole-protein and peptide array features that correlated significantly with age, H1N1, and B-strain post-vaccine titer as assessed through a standard microneutralization assay (p<0.05, q <0.2). Notably, we identified several peptide epitopes that were inversely correlated with regard to age and seasonal H1N1 and B-strain neutralization titer (p<0.05, q <0.2), implicating reactivity to these epitopes in age-related defects in response to H1N1 influenza. We also employed multivariate linear regression with cross-validation to build models based on age and pre-vaccine peptide reactivity that predicted vaccine-induced neutralization of seasonal H1N1 and H3N2 influenza strains with a high level of accuracy (84.7% and 74.0%, respectively).
Our methods provide powerful tools for rapid and accurate measurement of broad antibody-based immune responses to influenza, and may be useful in measuring response to other vaccines and infectious agents.
现有的测量流感疫苗免疫原性的方法禁止对免疫球蛋白识别的表位决定簇进行详细分析。开发能够捕获高水平抗体结合信息的高度多重蛋白质组学平台，将使研究人员和临床医生能够快速生成有意义的流感特异性抗体反应读数。
我们开发了流感血凝素（HA）全蛋白和肽微阵列，并通过使用针对线性和构象 HA 表位的商业抗体进行验证，证明了该阵列可以在广泛的动态范围内检测到特定的抗体反应。我们从 76 名年轻和老年受试者的血液中提取血清，在接种 2008/2009 年三价流感疫苗前和接种后 28±7 天进行，并用流感阵列抗原测定这些血清的抗体反应。
我们使用线性回归，并通过对 1000 次重采样的 Benjamini 和 Hochberg 方法进行多次假设检验校正，鉴定出与年龄、H1N1 和 B 株疫苗后滴度显著相关的流感全蛋白和肽阵列特征的抗体反应（通过标准微量中和测定评估，p<0.05，q <0.2）。值得注意的是，我们确定了几个与年龄和季节性 H1N1 和 B 株中和滴度呈负相关的肽表位（p<0.05，q <0.2），表明这些表位的反应与 H1N1 流感相关的年龄相关缺陷有关。我们还使用交叉验证的多元线性回归构建了基于年龄和疫苗前肽反应的模型，这些模型能够高度准确地预测季节性 H1N1 和 H3N2 流感株的疫苗诱导中和作用（分别为 84.7%和 74.0%）。
我们的方法为快速准确地测量流感的广泛基于抗体的免疫反应提供了有力的工具，并且可能在测量对其他疫苗和传染病的反应方面有用。
