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Over time, you will notice additional tightening

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The cell pellets were centrifuged for 10 min at 600 g and 4 C, resuspended in fluorescence-activated cell sorting (FACS) buffer (PBS with 0.5% bovine serum albumin (BSA) and 5 mM EDTA), and filtered through a 70 m nylon mesh filter (Biosharp, Beijing, China