Developing a sensitive and quantitative western blot kit

开发灵敏的定量蛋白质印迹试剂盒

• 批准号: 10256088
• 负责人: Hong Yan Liu
• 金额: $ 25.6万
• 依托单位: DOTQUANT, LLC
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2022-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10256088
• 关键词: Address; Affect; Animals; Antibodies; Attenuated; Biochemical Reaction; Biological; Biological Assay; Biology; Cells; Color; Communities; Complement; Data; Detection; Disadvantaged; Dyes; ERBB2 gene; Elastomers; Enzyme-Linked Immunosorbent Assay; Enzymes; Evaluation; Feedback; Fluorescence; Future; Gel; Goals; Gold; Horseradish Peroxidase; Hybrids; Image; Immunoblotting; Investigation; Kinetics; Label; Life; Light; Medicine; Membrane; Methods; Microscopy; Modeling; Modernization; Optics; Outcome; Phase; Photobleaching; Photons; Protein Analysis; Proteins; Protocols documentation; Quantum Dots; Reaction; Reproducibility; Research; Sampling; Signal Transduction; Source; Spottings; Standardization; Streptavidin; Techniques; Technology; Testing; Time; Tissues; Transillumination; Visualization; Western Blotting; absorption; base; design; detection method; detection sensitivity; flexibility; fluorescence microscope; fluorophore; imager; imaging probe; imaging system; improved; instrumentation; interest; member; migration; next generation; novel; particle; phase 2 study; polyvinylidene fluoride; technology development

ABSTRACT Western blotting is one of the most widely used bioanalytical techniques for the detection of proteins of interest in an unknown mixture such as cell/tissue homogenate or extract. This technology was invented in the late 1970s and has enabled numerous discoveries in biology and medicine in the past 40 years. Although it is not in the spotlight anymore, western blotting has been applied at least two times more often than other competing or complementing techniques, such as mass spec, ELISA, and IHC within the last 10 years. Despite this amazingly wide acceptance by the scientific community, western blotting, in general, is not a very sensitive technique and is only semi-quantitative. The lack of quantitation in Western blot has raised major concerns in the scientific community on data reproducibility. Building on two breakthroughs we made in recent years, monovalent Qdot probes and optically clear transfer membranes, we propose to develop the next generation Western blot kits with improved sensitivity, quantification ability, multiplexing capability and flexibility in readout (standard gel UV imager or microscopy-based particle counting). If successful, we expect this technology to quickly become a commercial product to power biology research for another 40 years. .

摘要 蛋白质印迹法是检测目标蛋白质的最广泛使用的生物分析技术之一 在未知混合物如细胞/组织匀浆或提取物中。这项技术发明于1980年代后期， 在过去的40年里，他在生物学和医学方面取得了许多发现。虽然不是 在聚光灯下，蛋白质印迹法的应用频率至少是其他竞争对手的两倍。 或补充技术，如质谱，ELISA和IHC在过去10年内。尽管如此 令人惊讶的是，被科学界广泛接受的蛋白质印迹法，一般来说，不是一个非常敏感的方法， 这是一种技术，只是半定量的。蛋白质印迹法定量的缺乏引起了人们的主要关注， 科学界对数据可重复性的看法。基于我们近年来取得的两项突破， 单价Qdot探针和光学透明的转移膜，我们建议开发下一代 具有改进的灵敏度、定量能力、多重能力和读出灵活性的蛋白质印迹试剂盒 （标准凝胶UV成像仪或基于显微镜的颗粒计数）。如果成功，我们预计这项技术将 迅速成为一种商业产品，为生物学研究再提供40年的动力。 .