Gibco trypan blue solution 0.4

Spleen and tumor cell suspensions were prepared by grinding sterile excised tissue between the frosted ends of microscope slides and then passing the tissue through a sterile 60 μM mesh. Splenocytes were processed by lysing red blood cells and washing with sterile PBS. For plasmacytoid dendritic cell analysis, tumor lysate was washed with sterile PBS and processed by lysing red blood cells prior to staining. For tumor lymphocyte analysis, tumor lysate was washed with sterile PBS, and the mononuclear cell fraction (including TILs) was enriched by Lympholyte^®-M Cell Separation Media (Cedarlane^®, Burlington, NC) according to the manufacturer’s protocol. Prior to use all cells were counted by a Z1™ Coulter Counter^® (Beckman Coulter, Inc.; Brea, CA) and/or a hemocytometer with Gibco™ Trypan Blue solution 0.4% (Life Technologies, Carlsbad, CA) with cell viability routinely >90%.