hESC FACS sorting protocol

1. wash hESC once with DMEM/F12 to remove most dead cells or cell debris.
2. add 1ml/well accutase, incubate at 37℃ for 8-10min.
3. Gently triturate cell aggregates with P1000 tips for 5-6 times to get homogeneous single cell suspension.
4. transfer cell suspension into 15ml tube, centrifuge at 200g for 5min.
5. Discard supernatant, resuspend cell pellets in 1-2ml mTeSR1 (Y27632+).
6. Filter cell suspension through 40um strainer into 15ml tube.
7. Count cell density, adjust density if needed.
8. Place cell suspension on ice before loading onto FACS instrument.
9. Load cell suspension into FACS sorter, collect target cells into mTeSR1 (Y27632+).
10. after FACS, collect cell pellets by centrifuge, resuspend cell pellets in mTeSR1 (Y27632+)