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用于蛋白质磷酸化定量的优化单分子下拉测定

JoVE Journal
Biochemistry

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JoVE Journal Biochemistry

An Optimized Single-Molecule Pull-Down Assay for Quantification of Protein Phosphorylation

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用于蛋白质磷酸化定量的优化单分子下拉测定

DOI:

10.3791/63665-v

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07:45 min

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June 06, 2022

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Elizabeth M. Bailey, Emanuel Salazar-Cavazos, Rachel M. Grattan, Michael J. Wester, David J. Schodt, Julian A. Rojo, Keith A. Lidke³, Diane S. Lidke³

¹Department of Pathology,University of New Mexico School of Medicine, ²Department of Physics and Astronomy,University of New Mexico, ³Comprehensive Cancer Center,University of New Mexico Health Sciences Center

Chapters

00:04Introduction
00:53Coverslip Preparation
03:18Functionalization of Array with Biotinylated Antibody
04:09SiMPull of Proteins of Interest from Whole-Cell Lysates
04:40Image Acquisition
05:45Results: Capture of EGFR-GFP from Protein Lysates Using SiMPull
07:19Conclusion

Summary

Automatic Translation

本方案描述了样品制备和数据分析，以使用改进的单分子下拉（SiMPull）测定来量化蛋白质磷酸化。

Tags

AI-powered Protein Phosphorylation Quantification Single-molecule Pull-down Assay SiMPull Protocol Antibody Labeling Autofluorescence Reduction Single-molecule Analysis Phosphorylation Status Of Individual Proteins Alternative To Western Blotting And Mass Spectrometry

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