I am newbie to immunology and trying to culture mouse spleen adherent cells to study cytokine production upon LPS stimulation.
I dissect spleen, mechanically dissociate using the end of plunger, and plate onto petridish/culture dish in RPMI1640/Heat Inactivated 10% New born Calf Serum/Pen/Strep for one hour.
After one hour incubation, I can see lots of cells adhere.
So I wash 3 times to remove non-adherent cells and incubate overnight. Then for some reason almost all cells come off the plate and not much left on the plate!
Please Immune expert here could you provide any solution/mistake I am making?
Thank you veery much in advance!