Luminex Technology for the Quantification of Cytokines in Non-Human Primates

描述（由申请人提供）：非人灵长类动物（NHP）是临床前研究的首选动物模型，因为它们在生理和遗传学上比任何其他动物更接近人类。通过使用NHP模型，免疫学研究取得了重大进展，特别是在艾滋病发病机制、治疗和疫苗开发方面。细胞因子和趋化因子是免疫系统的可溶性介质，在细胞间信号传导和细胞募集到炎症部位中起着至关重要的作用。鉴定NHP中的这些分子对于理解这些物种中发生的复杂生理和病理机制以及证明这些机制是否在人类中起类似作用至关重要。Luminex100系统是一种台式流式细胞仪，允许用户在单个管中同时定量多达100个分子。最近，我们发现了几种针对人类细胞因子的抗体，这些抗体具有识别同源趋化因子和NHP来源的细胞因子的能力。目前，我们有一组试剂，可以同时鉴定23种细胞因子和趋化因子，这些细胞因子和趋化因子来自黑猩猩和旧大陆猴，恒河猴，狒狒，食蟹猴，长尾猴和非洲绿猴。在这项R-24申请中，我们建议通过完成以下具体目标来促进和扩大Luminex技术在NHP中的使用：1)通过增加旧大陆猴子物种中可检测分子的数量来提高我们23-plex NHP Luminex面板的广度和分析能力；2)通过提高目前用于旧大陆猴子物种细胞因子定量的一些次优试剂的灵敏度，提高我们23层NHP Luminex面板的质量和深度；3)开发用于同时定量狨猴细胞因子的重组抗体，狨猴是一种新的世界猴子物种，越来越多地用于疫苗开发、衰老和肥胖研究。我们将对现有的针对旧大陆猴细胞因子的单克隆抗体进行广泛的测试，并使用最先进的重组抗体技术来创建针对狨猴细胞因子的抗体。这些研究产生的信息和方案将在SNPRC的流式细胞术网站上列出；新型抗体将分发给那些在成本回收基础上证明其需要的科学家。

DESCRIPTION (provided by applicant): Nonhuman primates (NHP) are the preferred animal models for pre-clinical research because they approximate humans in physiology and genetics more closely than any other animal. Critical advances in immunologic research have been made through the use of the NHP model, most notably in AIDS pathogenesis, treatment, and vaccine development. Cytokines and chemokines are soluble mediators of the immune system that play a crucial role in intercellular signaling, and in the recruitment of cells to inflammation sites. Identification of these molecules in NHP is crucial for the understanding of complex physiological and pathological mechanisms that occur in these species, and to demonstrate whether these mechanisms function similarly in humans. The Luminex100 system is a bench-top flow cytometer that allows the user to quantify up to 100 molecules simultaneously in a single tube. Recently, we identified several antibodies specific for human cytokines that have the capacity to recognize homologous chemokines and cytokines of NHP origin. Currently, we have a panel of reagents that allow for the simultaneous identification of 23 cytokines and chemokines from chimpanzees and the Old World monkeys rhesus macaques, baboons, cynomolgus macaques, pig-tailed macaques, and African green monkeys. In this R-24 application we propose to facilitate and expand the use of the Luminex technology in NHP by completing the following Specific Aims: 1) to improve the breadth and analytical power of our 23-plex NHP Luminex panel by increasing the number of detectable molecules in Old World monkey species; 2) to improve the quality and depth of our 23-plex NHP Luminex panel by increasing the sensitivity of some suboptimal reagents currently used for quantification of cytokines from Old World monkey species; and 3) to develop recombinant antibodies for the simultaneous quantification of cytokines in the marmoset, a New World monkey specie increasingly used in vaccine development, aging, and obesity research. We will perform extensive testing of available monoclonal antibodies against Old World monkey cytokines, and state-of-the-art recombinant antibody technology for the creation of antibodies against marmoset cytokines. The information and protocols generated from these studies will be listed in the Flow Cytometry website of the SNPRC; novel antibodies will be distributed to scientists who justify their need on cost-recovery bases.