HUMAN ANTIBODY REPERTOIRE TO H INFLUENZAE B

描述（改编自申请人的摘要）：研究 本提案中所述的措施是我们继续努力， 描述了可变区的遗传和结构基础 编码对B型特异的人抗体（Ab）库 (Hib)荚膜多糖（PS），并了解动力学 保护性抗体库不同的Hib PS结合疫苗 在婴儿中诱导Hib PS特异性抗体， 抗Hib的功能活性，这些差异与 特殊独特型/可变（V）区域的使用。此外，V 从婴儿开始， 而成人则不同地利用特定的抗Hib PS区域。的 研究人员的研究旨在确定分子机制， 这些现象背后。这一提议的一个核心假设是 由不同的Hib驱动的Ab V区的超突变 PS疫苗和暴露于天然存在的抗原作为 老化的结果，在决定V 区域/独特型表达和Ab功能质量。定义和 操纵婴儿和成人的抗Hib PS V区域库， 将使用两种实验方法，独特型分析和 人源性抗体Fab片段在噬菌体组合载体中的表达 图书馆.抗Hib PS V区cDNA序列分析 将在从接种疫苗的婴儿中获得的B淋巴细胞上进行， 成年人了这种方法将使我们能够确定V基因元件 编码婴儿抗Hib PS库，迄今为止， 无法达到分子定义。此外，该分析将 允许评估V区突变的程度， 由不同的疫苗和老化过程引起的。血清学 将制备与新独特型/V区反应的试剂， 将被用来扩展我们的V区表达的分析， 根据疫苗类型和年龄确定的人群。的关系 还将研究Hib PS的自身抗体反应性和特异性 由于V基因片段使用的抗Hib PS抗体也通常是 使用抗自身抗体。通过使用噬菌体操纵V基因组分， 组合文库，V区重组实验可以是 以评估单个V区的特异性潜力， 重链和轻链V区在 阻止抗体特异性和功能，以及抗体的结构决定因素 自我和非自我反应。将进行亲合力和特异性分析。 对序列定义的、噬菌体衍生的Fab进行，以关联V使用 和功能本周的长期目标是了解 形成人类抗体库的进化和躯体力量， PS抗原。在界定遗产继承规则时， 发育控制和表达的分子基础 这些重要的抗体特异性将帮助我们了解 不同的PS疫苗产生的抗体具有如此显著不同的 功能能力，从而使合理的设计， 更有效的未来PS疫苗用于婴儿以及 老人

DESCRIPTION (Adapted from the applicant"s abstract): The research described in this proposal represents a continuation of our efforts to delineate the genetic and structural bases of the variable (V) regions encoding the human antibody (Ab) repertoire specific for the type b (Hib) capsular polysaccharide (PS), and to understand the dynamics of this protective Ab repertoire. The different Hib PS conjugate vaccines induce Hib PS-specific Ab in infants that differ profoundly in functional activity against Hib, and these differences correlate with usage of particular idiotypes/variable (V) regions. In addition, V repertoire modification occurs as a consequence of aging since infants and adults differentially utilize particular anti-Hib PS regions. The investigator's research seeks to define the molecular mechanisms underlying these phenomena. A central hypothesis of this proposal is that hyper mutation of Ab V regions, variably driven by different Hib PS vaccines and by exposure to naturally-occurring antigens as a consequence of aging, plays a central role in determining both V region/idiotypic expression and Ab functional quality. To define and manipulate the anti-Hib PS V region repertoires of infants and adults, two experimental methodologies will be used, idiotypic analysis and expression of human Ab Fab fragments in bacteriophage combinatorial libraries. An extensive anti-Hib PS V region cDNA sequence analysis will be performed on B lymphocytes obtained from vaccinated infants and adults. This approach will allow us to determine the V gene elements encoding the infant anti-Hib PS repertoire, which heretofore have been inaccessible to molecular definition. In addition, this analysis will permit assessment of the extent to which V region mutation has been induced by different vaccines and by the aging process. Serological reagents reactive with new idiotypes/V regions will be prepared and they will be used to extend our analysis of V region expression in populations defined by vaccine type and age. The relationship between autoantibody reactivity and specificity for Hib PS will also be studied since the V gene segments used anti-Hib PS antibodies also are commonly used by anti-self Ab. By manipulating V gene components using phage combinatorial libraries, V region recombination experiments can be performed to assess the specificity potential of individual V regions, the relative roles that the heavy and light chain V regions play in deterring Ab specificity and function, and structural determinants of self and non-self reactivity. Avidity and specificity analyses will be performed on sequence-defined, phage-derived Fabs to correlate V usage and function. The long term objectives of this week are to understand the evolutionary and somatic forces that shape human Ab repertoires to PS antigens. Delineating the rules governing the inheritance, the developmental control and the molecular basis for the expression of these important Ab specificities will help us to understand how different PS vaccines generate antibodies with such markedly different functional capabilities, and thus will enable the rational design of more effective future PS vaccines for use in infants as well as the elderly.