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Flow cytometry of SirNeoblasts

KN Kaimeng Niu
HX Hao Xu
YX Yuanyi Zhou Xiong
YZ Yun Zhao
CG Chong Gao
CS Chris W. Seidel
XP Xue Pan
YY Yuqing Ying
KL Kai Lei
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In order to obtain isolated SirNeoblasts, the tails of the planarians (> 8 mm in length) were amputated, then pooled and rinsed in calcium and magnesium free buffer with 1% bovine serum albumin (CMFB). Cells were macerated by rocking in the tube on a rotating platform for 20 min with agitation every 3 min. After filtering the macerated cells through a 70 μm cell-strainer cap, the dispersed cells were centrifuged at 290 x g for 10 min. Cells were then resuspended in isotonic planarian medium (IPM) with 10% Fetal Bovine Serum (FBS, CellMax SA211.02) for SiR-DNA staining by incubation in SiR-DNA (1 μM, Cytoskeleton Inc., CY SC007) for 1 h and Cell Tracker green CMFDA stains (2.5 μg/ml, Thermo Fisher Technologies, C7025) for 10 min. Target cells were sorted using a BD Influx cell sorter equipped with a 100 tip and purity sort mode.

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