Hi all,

I am working on a project which needs expansion of CD45+ selected cells from tumor and lymph nodes. My Tumor samples expand good but my samples from the lymph nodes are less and less between each passage. I use a combination of low (3000 IU/mL on day 0), high (6000 IU/mL on day 4)and very low (2000 IU/mL on day 7) doses of IL-2 after using CD28/CD3 beads (1 bead:1 cell ratio). I plate ~10^6 cells/well in 48 well plate and split them on days 4 and 7 (they don't seem confluent). For splitting the cells I collect the media and lift the cells using tripLE (0.5 ml per well for 6 minutes) and then use the cell scrapers gently to collect all cells. My model is C57 mouse.

My initial PD-1 analysis showed less exhaustion of lymph node cells compared to tumors. I don't know what causes this. I appreciate your suggestions/comments.