HUMAN ANTIBODY REPERTOIRE TO H INFLUENZAE B

乙型 H 型流感病毒的人类抗体库

• 批准号: 3138295
• 负责人: Alexander H. Lucas
• 金额: $ 28.27万
• 依托单位: CHILDREN'S HOSPITAL & RES CTR AT OAKLAND
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-09-01 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3138295
• 关键词: Epstein Barr virus; Haemophilus influenzae; Haemophilus influenzae vaccines; antibacterial antibody; antibody specificity; antiidiotype antibody; antiserum; bacterial disease; bacterial genetics; bactericidal immunity; cell transformation; cross immunity; gene expression; genetic mapping; human subject; hybridomas; immunoglobulin genes; immunoglobulin idiotypes; immunological substance; laboratory mouse; monoclonal antibody; monocyte; nucleic acid sequence; passive immunization; polymerase chain reaction; synthetic peptide

Despite the availability of antibiotics, Haemophilus influenzae b (Hib) remains a significant human pathogen: it is the leading cause of meningitis in children in North America, and is the etiologic agent in other serious invasive diseases such as epiglottitis, septic arthritis, cellulitis and pneumonia. The Hib capsular polysaccharide (Hib PS) is the primary virulence factor, and antibodies to Hib PS confer protection from invasive Hib disease. Children less than two years of age synthesize little or no antibody to Hib PS and this accounts for this group having the highest susceptibility to infection. The experiments described here will examine the diversity of the neonatal and adult Hib PS antibody variable (V) region repertoire. Our objective is to determine the extent to which particular, germ-line encoded V domains dominate the antibody response to Hib PS, and to determine whether the age-dependent development of immunocompetence, is explicable in terms of limited and/or differential V region utilization. Idiotypic and sequencing analyses will be used to delineate V region expression. Hybridomas, secreting Hib-PS specific monoclonal antibody (mAb) will be prepared from adult peripheral blood mononuclear cells (MNC) and from hyperimmunized scid mice engrafted with cord blood MNC. This panel of human mAb's will be tested for expression of cross-reactive idiotype (CRI) using polyclonal and monoclonal antibodies specific for a recurrent and predominant CRI associated with antibodies to Hib PS. V region sequences of these mAbs will be determined by sequencing cloned, polymerase chain reaction amplified, hybridoma cDNA. Comparison between V region sequence and CRI expression should help elucidate the structural correlates of CRI and will delineate the molecular diversity of the neonatal and adult V region repertoires to Hib PS. To further localize CRI determinants, antibodies will be prepared against synthetic peptides which encode conserved hypervariable regions of mAb anti-Hib PS heavy (H) and light (L) chains. These anti-peptide antibodies will be tested for idiotypic specificity against the panel of human Hib PS-specific mAb's and their isolated H and L chains. This approach should allow for identification of particular hypervariable regions which encode CRI determinants, as well as delineate the individual roles of H and L chain V regions in CRI expression. The relationship between CRI/V region expression and Hib PS antibody affinity and functional activity will also be examined. MAb's reactive with the major CRI of Hib PS antibodies will be administered to scid-hu mice to determine whether they might function as a surrogate Hib vaccine. These studies should provide insight into the ontogeny of V region expression and the relationship between V region structure, idiotypes and antibody function.

尽管有抗生素可用，但流感嗜血杆菌B（HIB） 仍然是人类病原体：它是脑膜炎的主要原因 在北美的儿童中，是其他严重的病因学药 侵入性疾病，例如上皮炎，化脓性关节炎，纤维炎和 肺炎。 Hib帽囊多糖（HIB PS）是主要的 毒力因子和HIB PS抗体抗体赋予侵入性保护 HIB病。年龄不到两岁的儿童综合几乎没有或没有 HIB PS的抗体，这说明了该组的抗体 感染的敏感性。这里描述的实验将检查 新生儿和成人HIB PS抗体变量（V）区域的多样性 曲目。我们的目标是确定特定的程度， 种系编码的V结构域主导了对HIB PS的抗体响应，并且 确定免疫能力的年龄依赖性发展是 可从有限和/或差异V区利用率方面解释。 白痴型和测序分析将用于描述V区域 表达。杂交瘤，分泌HIB-PS特异性单克隆抗体（MAB） 将从成人外周血单核细胞（MNC）和 来自植入脐带血跨度的高免疫的SCID小鼠。这个面板 人MAB的表达将被测试 使用针对复发的多克隆和单克隆抗体， 与HIB PS抗体相关的主要CRI。 V区域序列 这些mAB将通过测序克隆的聚合酶链确定 反应扩增，杂交瘤cDNA。 V区域序列之间的比较 CRI表达应有助于阐明CRI的结构相关性 并描绘新生儿和成人V的分子多样性 hib ps的区域曲目。为了进一步定位CRI决定因素， 将制备针对编码的合成肽的抗体 保守的MAB抗Hib PS PS（H）和光（L）的高变量区域 链。这些抗肽抗体将被测试 针对人Hib PS特异性mab及其面板的特异性 孤立的H和L链。这种方法应允许识别 编码CRI决定因素以及 描述H和L链V区域在CRI中的各个角色 表达。 CRI/V区域表达与HIB PS之间的关系 还将检查抗体亲和力和功能活性。 mab的 用HIB PS抗体的主要CRI反应性 SCID-HU小鼠以确定它们是否可以充当替代HIB 疫苗。这些研究应洞悉V区域的个体发育 表达和V区域结构，白痴和 抗体功能。