I did a competitive bone marrow transplantation using sorted 250 of HSCs (Lin-, cKit+, Sca1+, CD48-, CD150+) from CD-1 background mice mixed with 5*10^5 helper bone marrow cells from CD45.1 mice (C57BL/6 background). The recipient mice (CD45.1 B6.SJL) were sub-lethally irradiated before transplant. I checked donor reconstitution 4 weeks after transplant and I found no CD45.2+ population in all the recipient mice. I believe the irradiation worked because one recipient mouse died few days after transplant. I was wondering if the graft rejection occurred in this transplant experiment so that only the helper bone marrow cells successfully engrafted.
Dear Wei-Chieh,
I cannot directly reply to your question, but I can tell you that in similar BM transplantations I also had cases of complete rejection of cells by the irradiated hosts, and I attributed that to the fact that donnor mice were not in full b6 backgroung as hosts and as 3rd party cells (or irradiation protective cells). I overcame the problem using animals lacking lymphocytes, ex: Rag deficient animal.
You can confirm this "quickly" by repeating chimeras in a small group of animals with total BM (or T cell depleated) transplant instead of HSCs. Don't forget to sex match animals in these type of experiments.
Neverthless, since you are checking HSC's progeny bleed them again at 6 or 8 weeks just to double check.
Good luck!
Silvia
PS-When doing these type of experiments with few sorted cells to use in vivo, I used the 100mm nozzle in the sorter to maximize cell viability.