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. ? ? ?
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