I have seen a variety of methods that people use to rest after thawing PBMCs to inject them into mice including:
(1) Thaw, wash in HBSS, and inject right away (no resting)
(2) Thaw, and then culture in RPMI (5% human serum, 1% pen/strep, 1% LGlutamine, 1% NEAA, 1% sodium pyruvate, and 0.1% E-mercaptoethanol) at 37 overnight.
(3) Thaw, and rest overnight at 4C in RPMI with 20% FBS and 10 mM EDTA in Falcon tubes
(4) Thaw, rest in RPMI (1% pen/strep, 10% FBS) for 18h at 37
Has anyone humanized NSG mice using cyropreserved PBMCs. What methodology did you use for thawing/resting, and did you successfully humanize your mice?
Thanks in advance!!!
Option number 2 sounds reasonable to be. Thawed cells will repair its membranes during this time. Also Cryoprotectant will get diluted or washed away before injection.
