Direct Sequencing of Antibodies of the Influenza Immune Response

摘要 流感的年度流行导致大量的住院治疗， 据估计，全球有300万至500万例严重疾病和30万至50万例死亡。 在20世纪，发生了三次主要的流感大流行， 5000万到1亿人。最近，新的流感单克隆抗体（mAb）已被发现。 鉴定出能够中和多种病毒。在过去的八年里， 已经鉴定了几种广泛中和的茎反应性抗体。这些干 只有一种抗体识别所有甲型流感亚型，但这种识别范围 仅在体外诱变后才被引发。这表明，一个表位，以指导普遍的 疫苗战略尚未确定。干细胞抗体可以提供普遍的流感 因为它们可能能够中和甲型和B型流感病毒。一 鉴定中和B细胞特异性抗体的策略涉及来自 免疫个体。与H5和H7 HA亚型交叉反应的B细胞衍生mAb 已显示产生可结合所有甲型流感HA的高亲和力茎特异性mAb 亚型另一种不涉及B细胞克隆的策略是从抗原开始 然后可以直接将干细胞结合多克隆抗体（pAb）的特异性群体 通过蛋白质组串联质谱法（MS/MS）测序。这些pAb是优选的 mAb发现的来源，因为外周抗体群，而不是外周B细胞 群体，代表完整的体液免疫应答。直接测序 外周pAb可以快速挖掘这种免疫多样性，以产生针对新的和独特的 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.