Quantikines

The compound library, InhiTinib, and its related analogs were provided by the IRIC HTS platform. Because the analog ligands are proprietary, their structures are not shown here. CD3-CD28 dynabeads, CellTrace^TM, Hoechst 33342, nerve growth factor (NGF), and MitoSox^TM Red were purchased from Thermo Fisher. Human and murine interferon (IFN)-gamma Quantikines were purchased from R&D System. Annexin-V, propidium iodide (PI) and anti-TUBB3 antibody were purchased from Biolegend. Lipopolysaccharide, glial-derived neurotrophic factor (GDNF), cytosine arabinoside (AraC), and the immunosuppressive agent Cyclosporin A were purchased from Sigma. Collagenase A and dispase II were purchased from Roche Applied Sciences. T-cell isolation kit and lymphoprep^® were purchased from StemCell Technologies. Z-VAD-FMK pan-caspase inhibitor was purchased from APExBIO. Western blot antibodies against human and murine poly(ADP-Ribose) polymerase-1 (PARP-1) (Cat. #sc-8007 and 46D11) were purchased from Santa Cruz and Cell Signaling, respectively; γH2AX (Cat. #05-636) from EMD Millipore; and H4 and tubulin (Cat. #ab6161) from Abcam SPHERO^TM. AccuCount Particles were purchased from Spherotech, Inc.

TNFα, IFNγ, IL-6 and IL-1β were quantified with Multiplex technology using R&D System kit (Minneapolis, Minnesota, USA) (LXSAHM-05). Data were analyzed with Bioplex 2000™ device. TGF-β was quantified in cell culture supernatants by R&D System Quantikines.
Extracellular GRP94 was quantified using the ELISA kit from Cloud Clone.

Recombinant IL-7 (#217-17) and IL-21 (#210-21 for mouse and #200-21 for human) cytokines were purchased from PeproTech (Rocky Hill, NJ, USA). The flow cytometry antibodies CD3 (17A2), CD4 (GK1.5), CD5 (53-7.3), CD8 (53-6.7), NK1.1 (PK136), CD19 (1D3), CD25 (PC61), CD34 (RAM34), CD44 (1M7), CD45 (30-F11), CD62L (MEL-14), CD135 (A2F10.1), IL-10 (JES5-16E3), human CD3 (UCHT1), human CD4 (RPA-T4), human CD8 (SK1), human CD13 (WM15), human CD45 (H130), and Cytofix/Cytoperm Kits were purchased from BD Pharmingen (San Diego, CA, USA). FOXP3 (FJK-16S), granzyme B (NGZB), and human CD161 (HP-3G10) antibodies as well as monensin were purchased from eBioscence/Thermo Fisher (Waltham, MA, USA). Quantikines were purchased from R&D Systems (Minneapolis, MN). CD3-CD28 beads, CellTrace^TM, and TRIzol were purchased from Invitrogen (Burlington, ON, CA). Lipopolysaccharide (LPS), PMA, and ionomycin were purchased from Sigma (St. Louis, MO, USA). All cell isolation kits were purchased from STEMCELL Technologies (Vancouver, BC, Canada). RNA extraction kits were purchased from Qiagen (Toronto, ON, Canada).