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Antibody Validation Vector Core

抗体验证载体核心

基本信息

批准号：
10231257
负责人：
Feng Qian
金额：
$ 18.2万
依托单位：
UNIVERSITY OF MARYLAND BALTIMORE
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-08-24 至 2025-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10231257
关键词：
3-Dimensional Affinity Animal Model Animals Antibodies Antibody Affinity Antigens Area Basic Science Binding Biochemistry Biotechnology Cell Culture System Cell surface Cells Clinical Trials Cloning Communities Complementary DNA Complex Critical Pathways Development Epitopes Gene Expression Gene Proteins Genes Goals Immunization Immunize Immunoprecipitation Length Libraries Lipids Mammalian Cell Maryland Membrane Proteins Monoclonal Antibodies Mus Mutate PKD2 protein Pathway interactions Phage Display Polycystic Kidney Diseases Positioning Attribute Principal Investigator Property Proprotein Convertase 1 Proprotein Convertase 2 Protein Fragment Proteins Protocols documentation Reagent Research Research Personnel Role Science Silent Mutation Site Specificity Surface Technical Expertise Translations Validation Western Blotting Work anti-PD1 antibodies experience expression vector in vivo innovation mutant nanobodies nanodisk next generation novel novel strategies polycystic kidney disease 1 protein programmed cell death protein 1 protein complex repository restriction enzyme tool vector

项目摘要

Project Summary A major obstacle to the development of protective therapies for polycystic kidney disease (PKD) has been our limited understanding of the cellular pathways that are altered when PKD genes are mutated. In general researchers have identified critical pathways in cell culture systems and then proceeded to validate them in animal models with the goal of moving on to clinical trials. This pipeline requires a robust tool kit of basic science reagents including high quality antibodies and expression constructs. This has been particularly challenging for PKD because polycystins in particular are multi-domain membrane proteins that are difficult to work with. Many of the available antibodies lack sufficient affinity and specificity and large cDNA constructs are difficult to manipulate in cloning applications. The overarching goal of the Antibody and Vector Core is not only to supply investigators with our existing reagents, but to apply recent advances in biotechnology to develop the most innovative tools that will advance the next generation of discoveries in PKD research. This application builds on the Principal Investigator’s decades long experience in the study of polycystin biochemistry that will allow him to provide technical expertise so that investigators can fully leverage these novel reagents. In order to accomplish this goal we propose the following specific aims: 1) Maintain and provide validated PKD antibody kits that detect full length endogenous proteins 2) Use a novel strategy of whole cell immunization with cell surface PC1/PC2 to develop mouse monoclonal antibodies directed against the polycystin complex in its native configuration 3) Develop the first PC1 and PC2 Camelid VHH nanobodies using purified and stabilized PC1/PC2 complex antigen 4) Develop the first PC2 VHH nanobodies using purified and functional PC2-tetramer embedded in lipid nanodiscs and 5) Provide and develop easy-to-use vectors for PKD gene expression, inactivation and manipulation. In summary the Antibody and Vector Core will provide the research community with a comprehensive array of validated reagents and unique expertise that will facilitate discoveries in the PKD field.

项目总结