Isolation and Quantification of Murine Immune Cells

The mice were sacrificed by cervical dislocation. Bone marrow, spleen and thymus were collected into tissue culture medium prepared as described previously [25 (link)]. Bone marrow cells were extracted by pressurized flow of buffer through dissected femurs and tibias. Single cell suspensions from spleen and thymus were prepared by passing the tissues through 70 μm nylon mesh filters (BD Biosciences). Red blood cells (RBC) in the spleen samples were removed by incubating splenocytes with RBC lysis buffer (150 mM NH₄Cl, 10 mM KHCO₃, 0.1 mM EDTA, pH 7.3). White blood cells were counted using the ViCELL cell counter (Beckham Coulter Inc.).