Xenograft B cell Lymphoma Model

Female, NOD.Cg-Prkdc^scid Il2rg^tm1Wjl/SzJ mice (NSG) mice (6 wks, ~15 g; Jackson Laboratories), were used for all in vivo studies involving re-stimulated 19BBz T cells. NSG mice were inoculated with 1×10⁵ luciferized Raji cells intravenously (day 0). On day 4, mice were randomized into treatment groups within each cage and intravenously administered vehicle (RPMI-1640) or 10⁷ 19BBz T cells re-stimulated with either APC-ms or Dynabeads for 7 or 14 days. The investigator performing the 19BBZ T cell infusions was blinded to the treatment group. Prior to cell infusion, cells were collected, washed twice with T cell media and with RPMI-1640, and then resuspended at the appropriate concentration in RPMI-1640. Dynabeads were magnetically removed (Dynal) and cell products were visually inspected for Dynabead contamination prior to injection. 19BBz T cells restimulated with APC-ms were used without additional purification steps. Tumor burden was evaluated over time by intraperitoneal Luciferin (Gold Biotechnology) injection and luminescence measurements 10 minutes post-injection via IVIS (Perkin Elmer). Total flux (p/s) per mouse was quantified in regions-of-interest (ROI) defined around individual mice. A single ROI with the same dimensions was placed below the mice as a measure of background luminescence in the image field, due to substantial bleed-over from mice with high tumor burden within the chamber, and used to normalize the signal of all mice imaged within the chamber. Luminescence signal of each individual mouse was reported as normalized total flux, calculated as (total flux)_{mouse ROI}/(total flux)_{background ROI}. One animal was excluded from analysis as it was accidently administered a different dose. Mice were monitored daily for signs of discomfort and euthanized upon losing more than 10% of initial body weight, the development of hind-limb paralysis, or when graft-versus-host disease was evident (hair loss, behavioral changes, decline in overall health).