Protocol for high-sensitivity western blot on murine hematopoietic stem cells
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Changzheng Li1, Jingjing Guan1, Yishan Li1
1Advanced Medical Technology Center, The First Affiliated Hospital, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, Guangdong, China; Key Laboratory of Stem Cells and Tissue Engineering (Ministry of Education), Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, Guangdong, China.
Abstract:
Hematopoietic stem cells (HSCs) sustain hematopoiesis during homeostasis and regeneration. However, their limited availability poses a challenge for protein analysis. Here, we present a protocol for performing high-sensitivity western blot on HSCs using two techniques that enhance HSC isolation from mice and boost sensitivity for low cell numbers. We describe steps for isolating murine bone marrow cells, antibody staining, and cell sorting and post-sort analysis. We then detail a western blot procedure suitable for low numbers of HSCs. For complete details on the use and execution of this protocol, please refer to Li et al (2022).1,2.