Salmonella typhimurium flagellin fla st

Salmonella typhimurium flagellin (FLA-ST) was purchased from Invivogen. CBLB502 was a gift from Cleveland Biolabs, Inc. Monoclonal antibodies 9D9 (anti-CTLA-4) and RPM1-14 (anti-PD-1) were purchased from BioX Cell and maintained in 6.5 and 6.7 mg/mL stocks, respectively, and stored at 4 °C before use. d-luciferin (d-Luc) (BioGold), the substrate for firefly luciferase, was maintained in a 30 mg/mL solution of phosphate-buffered saline (PBS). Matrigel was obtain from Corning and maintained at −20 °C.

Collagenase-1, DNase-1, 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), Griess reagent, FITC-dextran, and Concanavalin-A were all purchased from Sigma Aldrich (St. Louis, MO). CD11c magnetic cell separation kit (MACS) were purchased from Miltenyi Biotec (Bergisch-Gladbach, Germany). All monoclonal antibodies (Abs) were purchased from eBioscience (San Diego, CA), while all the PRR agonists were purchased from InvivoGen (San Diego, CA). For all in vivo studies, ultrapure Salmonella typhimurium flagellin (FLA-ST) from InvivoGen (endotoxin level < 0.05 endotoxin units (EU)/mg) and was employed. FLA-ST is purified by acid hydrolysis, heating and ultrafiltration to obtain an estimated at 10% purity, before an additional purification step using monoclonal anti-flagellin affinity chromatography to obtain purity >95%.