We are developing thymocytes maturation co-cultures employing OP9-DL1 employing mice of 4-5 weeks-old. To isolate DN populations, we prepare thymus cell suspensions and we use this antibodies cocktail: CD8a, CD4, TCRγδ, TCRβ, Gr1, Ter119, CD122, NK1.1, CD5, CD11c, CD11b and CD45R(B220) for negative selection of the most immature populations (DN1 and DN2).

However, we do not manage to eliminate the DN3 population. I would be very grateful if somebody could give us a piece of advise to improve this purification.