Generation of monoclonal and polyclonal antibodies to neolacto-series ganglioside

描述（由申请人提供）：神经节苷是存在于浆细胞膜中的唾液化鞘糖脂。它们尤其集中在神经系统中。神经节苷类在细胞相互作用和信号转导中起着重要作用。根据寡糖结构的不同，神经节苷类可分为神经节系列和新乳系列。神经节系列神经节苷脂存在于中枢神经系统（CNS）和周围神经系统（PNS），而新乳酸系列神经节苷脂主要存在于PNS。人PNS主要的新乳酸系列神经节苷脂是LM1和Hex-LM1。神经节苷类在多种自身免疫性神经病变中作为抗原存在。例如，抗gm1抗体与运动神经病变有关，抗gd1a抗体与急性运动轴索神经病变有关，抗lm1抗体与格林-巴利综合征（GBS）有关，抗gq1b抗体与米勒-费雪综合征（GBS的一种变体）有关。新乳酸系列神经节苷类抗体在自身免疫性周围神经病变中的作用以及这些神经节苷类的定位和功能尚不清楚。这部分是由于缺乏针对这些神经节苷脂的特异性探针。缺乏特异性抗体等探针的一个原因是很难从周围神经中分离出大量的神经节苷脂。有趣的是，LM1神经节苷脂也存在于人类红细胞中。我们已经用人血分离和纯化了大量的LM1。本研究的总体目标是开发针对新乳酸系列神经节苷脂的高亲和力特异性抗体。我们建议通过两个具体目标来实现我们的目标。在Specific Aim 1中，我们将使用纯化的LM1偶联keyhole帽贝血青素（KLH）作为免疫原，开发小鼠单克隆抗体。在Specific Aim 2中，我们将用LM1混合KLH免疫新西兰大白兔，并用弗氏佐剂乳化产生多克隆抗体。我们的研究将有助于开发针对LM1和Hex-LM1的高亲和力IgG抗体。新乳酸系列神经节苷脂的多克隆抗体和单克隆抗体的可用性将为阐明它们在周围神经中的定位和生物学功能提供非常有力的工具。这些抗体也将是非常有用的描述他们的致病潜力自身免疫性疾病的周围神经系统。公共卫生相关性：目前还没有针对周围神经系统新乳酸系列神经节苷的特异性试剂，如单克隆抗体。开发针对这些重要神经节苷脂的高亲和力抗体将为探索它们在健康和疾病中的生理功能提供有力的工具。

DESCRIPTION (provided by applicant): Gangliosides are sialylated glycosphingolipids present in plasma cell membranes. They are particularly concentrated in the nervous system. Gangliosides have been implicated as playing major roles in cellular interactions and signal transduction. Based on the oligosaccharide structures, gangliosides can be categorized into several major families, including ganglio-series and neolacto-series. While ganglio- series gangliosides are present in both the central nervous system (CNS) and peripheral nervous system (PNS), the neolacto-series gangliosides are present mainly in the PNS. The major neolacto- series gangliosides of human PNS are LM1 and Hex-LM1. Gangliosides have been implicated as antigens in various autoimmune neuropathies. For example, anti-GM1 antibodies are associated with motor neuropathy, anti-GD1a antibodies are associated with acute motor axonal neuropathy, anti-LM1 antibodies are associated with the Guillain-Barre syndrome (GBS) and anti-GQ1b antibodies are associated with Miller Fisher syndrome, a variant of GBS. The role of antibodies to neolacto-series gangliosides in autoimmune peripheral neuropathies and the localization and function of these gangliosides remain unknown. This is partly due to the lack of specific probes for these gangliosides. One reason for the lack of probes such as specific antibodies is that it is difficult to isolate large quantities of these gangliosides from peripheral nerves. Interestingly, LM1 ganglioside is also present in human red blood cells. We have used human blood to isolate and purify large quantities of LM1. The overall objective of this study is to develop high-affinity specific antibodies against neolacto-series gangliosides. We propose to achieve our objective in two specific aims. In Specific Aim 1, we will develop mouse monoclonal antibodies using purified LM1 conjugated to keyhole limpet hemocyanin (KLH) as an immunogen. In Specific Aim 2, we will generate polyclonal antibodies in New Zealand white rabbits by immunizing them with LM1 mixed with KLH and emulsified in Freund's adjuvant. Our study should lead to the development of high-affinity IgG antibodies to LM1 and Hex-LM1. The availability of polyclonal and a panel of monoclonal antibodies to the neolacto-series gangliosides will provide very powerful tools to elucidate their localization and biological function in the peripheral nerves. These antibodies will also be very useful to delineate their pathogenic potential in autoimmune diseases of the peripheral nervous system. PUBLIC HEALTH RELEVANCE: Specific reagents such as monoclonal antibodies to peripheral nervous system neolacto- series gangliosides are currently not available. Development of high-affinity antibodies to these important gangliosides would provide powerful tools to probe their physiological functions in health and disease.