LPS-specific Aptamers for Microbial Detection

性状（由申请方提供）：脂多糖是革兰氏阴性菌表面的显著特征。识别和区分不同O-特异性寡糖单元的抗体用于鉴定在污染食品中发现的致病细菌菌株，这些菌株已导致人类死亡和疾病。LPS分子通常密切参与发病机制，并参与细菌粘附和运动等功能，以及引发自身免疫反应。脓毒性休克综合征是一种严重的炎症反应，通常由LPS引起，是致命的，40%的有症状的人死亡。认识到病原菌是致命的，容易传播，并可用于生物恐怖主义，强调需要随时发现病原菌和有解毒剂，以防止他们。 抗体是检测特异性LPS分子的非常重要的手段。然而，抗体的生产和特异性是昂贵的，结果可能是可变的。有时，抗体也在两种LPS结构之间或与人体组织中的碳水化合物交叉反应，因此使用有限。在这里，我们建议开发识别特定LPS结构的适体。适体是与抗体一样以高亲和力特异性识别它们所选择的靶大分子的核酸。与抗体不同，适体是离体选择的，因此比抗体更容易修饰，从而具有特定的所需特征。适体还具有许多有利于诊断和现场工作的特征。我们对抗LPS适体的长期目标是，它们将为从细菌表面检测LPS的能力增加一个新的维度，并且这些适体也可能在被动免疫或其他医学应用中找到用途。我们预见到适体在均相测定和微阵列中用于病原体的原位检测，作为未来公共场所中用于早期检测生物恐怖活动的原位检测系统的一部分。 本申请的具体目的是：1）选择以高特异性和高亲和力结合LPS的适体，2）表征所选择的适体，3）开发检测特异性LPS寡糖的变构适体CLAMP。

DESCRIPTION (provided by applicant): Lipopolysaccharides are a distinguishing feature of gram negative bacterial surfaces. Antibodies that recognize and distinguish between different O-specific oligosaccharide units are used to identify pathogenic bacterial strains that are been found in contaminated foods and have caused human death and disease. LPS molecules are often intimately involved in pathogenesis and participate in functions such as bacterial adhesion and movement, and in eliciting autoimmune responses. Septic shock syndrome, a severe inflammatory response that is often elicited by LPS, is deadly, killing 40% of individuals with symptoms. Recognition that pathogenic bacteria are deadly, easily dispersed and could be used for bioterrorism emphasizes the need to readily detect pathogenic bacteria and of having antidotes to protect against them. Antibodies are a very important means of detecting specific LPS molecules. However, the production and specificity of antibodies is expense and results can be variable. On occasion, antibodies are also cross-reactive between two LPS structures or with carbohydrates in human tissues and thus have limited use. Here we propose to develop aptamers that recognize specific LPS structures. Aptamers are nucleic acids that, like antibodies, specifically recognize with high affinity the target macromolecules against which they were selected. Unlike antibodies, aptamers are selected ex-vivo and consequently can be more readily modified than antibodies so as to possess particular desired characteristics. Aptamers also possess many characteristics that are advantageous for diagnostics and field work. Our long term goal for anti-LPS aptamers is that they will add a new dimension to the ability to detect LPS from bacterial surfaces and that these aptamers also might find use in passive immunity or in other medical applications. We foresee the use of aptamers in homogeneous assays and in microarrays for on location detection of pathogens as a part of future in-place detection systems in public spaces for early detection of bioterrorist activities. The specific aims for this application are to: 1) Select for aptamers that bind LPS with high specificity and high affinity, 2) Characterize the selected aptamers, 3) Develop an allosteric aptamer CLAMP that detects a specific LPS oligosaccharide.