Nutrifreez d10 cryopreservation medium

XF hEPS cells were dissociated and centrifuged at 450–600 x g for 3–5 min. Then supernatant was discarded and 1 ml of cold cryopreservation solution (NutriFreez® D10 Cryopreservation Medium, Biological Industries, 05–713) was added to suspend cells, which were transferred to cryopreservation tubes. The tubes were immediately stored in Gradient cooling box at −80 °C Celsius for 24 h, then transferred to liquid nitrogen tank for long-term preservation.

Donor leukemic CD45.2 cells or control cells were transplanted into new hosts (chimeric F1 type mice presenting CD45.1 + CD45.2) accompanied by competitor CD45.1 cells. Cultured donor cells and WBM of competitor cells were mixed and intravenously (iv) injected into the tails of lethally irradiated host mice. BM or spleen cells were extracted from ML23 and ML21 mice, 1 million fresh cells were used to passage the leukemia line to secondary and tertiary recipient. For cryopreservation, BM or spleen cells from leukemic ML23 or ML21 mice were aliquted to 40 million cells per tube in NutriFreez D10 Cryopreservation medium (Biological Industries, Israel). The cells were kept frozen in −80 °C. Frozen cells were defrost and counted with trypen blue, 1 million viable cells were injected per mouse.