Direct Sequencing of Antibodies of the Influenza Immune Response

摘要 每年的流感大流行导致相当多的人因流感而住院 据估计，全球有300万至500万严重疾病病例，30万至50万人死亡。 在20世纪期间，发生了三次主要的流感大流行，总死亡率为 五千万到一亿人。最近，新的流感单抗(MAbs)已经被发现 已确定的能够中和多种病毒的药物。在过去的八年里， 已鉴定出几种广泛中和的干细胞反应性抗体。这些茎中的 抗体，只有一种识别所有的甲型流感亚型，然而识别的广度 只有在体外诱变后才能诱发。这表明，引导宇宙的表位 疫苗战略尚未确定。干细胞抗体可以提供普遍的流感 覆盖范围，因为它们有可能中和甲型和乙型流感病毒。一 鉴定中和干细胞特异性抗体的策略涉及从 已接种疫苗的个体。与H5和H7 HA亚型交叉反应的B细胞来源的单抗 已被证明可产生高亲和力的干细胞特异性单抗，可与所有甲型流感病毒HA结合 子类型。另一种不涉及B细胞克隆的策略是从抗原开始 干细胞结合多克隆抗体(PAB)的特定群体，然后可以直接 用蛋白质组级联质谱(MS/MS)进行测序。这些Pabs是首选 发现mAb的来源是因为外周抗体群体，而不是外周B细胞 群体中，是体液免疫反应充分的代表。直接测序 外周抗体可以快速挖掘这种免疫多样性，以产生针对新的和独特的单抗 表位。

Abstract The annual epidemics of influenza result in a substantial number of hospitalizations with an estimated 3 to 5 million cases of severe disease, and 300,000 to 500,000 deaths globally. During the 20th century, three major influenza pandemics have occurred with a total mortality of 50–100 million people. Recently, new influenza monoclonal antibodies (mAbs) have been identified that are capable of neutralizing a wide range of viruses. In the last eight years, several broadly neutralizing, stem-reactive antibodies have been identified. Of these stem antibodies, only one recognized all influenza A subtypes however this breadth of recognition was only elicited after in vitro mutagenesis. This suggests that an epitope to guide a universal vaccine strategy has yet be defined. Stem antibodies could provide universal influenza coverage since they are potentially capable of neutralizing both influenza A and B viruses. One strategy to identify neutralizing stem-specific antibodies involves immortalized B cells from immunized individuals. B cell-derived mAbs that cross react with the H5 and H7 HA subtypes have been shown to yield high affinity stem-specific mAbs that can bind to all influenza A HA subtypes. An alternative strategy, that doesn’t involve B cell cloning, starts with an antigen specific population of stem binding polyclonal antibodies (pAbs) that can then be directly sequenced by proteomic tandem mass spectrometry (MS/MS). These pAbs are a preferred source for mAb discovery since the peripheral antibody population, not the peripheral B cell population, is representative of the full humoral immune response. Direct sequencing of peripheral pAbs can rapidly mine this immune diversity to yield mAbs against novel and unique epitopes.