Isolation and Culture of Immune Cells

RPMI 1640 (Life Technologies, Carlsbad, CA, USA) supplemented with 10% fetal bovine serum (FBS) (Gemini Bio-Products, West Sacramento, CA, USA) was used for peripheral blood mononuclear cells (PBMCs), NK cells, T cells, monocytes, and oral squamous carcinoma stem cells (OSCSCs) cultures. Recombinant IL-2 was obtained from NIH- BRB. Antibodies to CD16 (clone 3G8) were purchased from Biolegend, San Diego, CA, USA. ImmunoCult-XF T cell expansion medium and Immunocult human CD3/CD28 T cell activator was purchased from Stem Cell Technologies (Vancouver, BC, Canada). OSCSCs were isolated from patients with tongue tumors at UCLA [22 (link),23 (link),24 (link),25 (link)]. Probiotic bacteria, sAJ2 is a combination of 8 different strains and is prepared as described previously [25 (link)], and RPMI 1640 supplemented with 10% FBS was used to re-suspend AJ2. Human ELISA kits for IFN-γ were purchased from Biolegend, San Diego, CA, USA. Chromium-51 radionucleotide was purchased from PerkinElmer, Richmond, CA, USA. The following antibodies with their respective clones were purchased from Biolegend, San Diego, CA, USA and used as suggested by the manufacturer for flow cytometric analysis; CD45 (H130) CD3/16/56: (UCHT1/3G8/MEM-188), CD3 (UCHT1), CD8 (HIT8a), CD14 (63D3) CD19 (HIB19).