Fetal Liver infection

1. Lin- cells are resuspended with 48-72hr retroviral medium and seeded in 6 well plate.

2. 4-10ug/ml polybrene was added and cells were spin infected at 37 for 2 hours at 2000Xrpm.

3. Remove half retroviral medium and add 1/2 total volume of fresh medium with 10% FBS, 50uM of beta-ME, 20ng/ml SCF, 30ng/ml Flt3L, 10ng/ml of Il-6 culture o/n

4. Spin and remove medium and replace with 72-96hr retroviral medium with polybrene for another 1-2hrs at 2000 rpm.

5. Remove supernatant and resuspend cells in Fresh IMDM with 2% FBS at concentration of 1X10*6/100ul for injection. 100ul /mouse.