Ivis spectrum ct instrument

On day 4 (aerosol) or day 6 (sc) post challenge, mice were injected with 10 μg of Nano-Glo substrate sc in 500 μl PBS as previously described [59 (link)]. Four min after substrate injection, the mice were imaged using the IVIS Spectrum CT Instrument (PerkinElmer) using the autoexposure setting. The total flux (photons per second) in the head region was calculated for each animal using Living Image Software 4.5.1 with all images set to the same scale. Images of representative animals are shown from each LAV vaccine candidate.

NSG (NOD.Cg-Prkdcscid/Il2rgtm1Wjl/SzJ) mice were purchased from The Jackson Laboratory and housed in a specific-pathogen-free environment. For testing the in vivo effector functions of the OKT3-antiCD28/RPMI- and OKT3-RetroNectin/LymphoONE-stimulated HER2.CD28.z and HER2.41BB.z CAR T cells, seven-week-old female NSG mice were administered a subcutaneous (s.c.) injection in both flanks, each containing 3 × 10⁶ JIMT-1.ffLuc cells in 100 µL of PBS mixed with an equal volume of Matrigel (BD Biosciences, San Jose, CA, USA). Tumor growth was monitored with an IVIS Spectrum CT instrument (Perkin Elmer, Waltham, MA, USA). Before measurement, isoflurane-anesthetized animals were injected IP with D-luciferin (150 mg/kg). A bioluminescence image was obtained and analyzed after 10 min using Living Image software Version 4.0 (Caliper Life Sciences, Waltham, MA, USA). A region of interest of the same size was drawn over the tumor region, and the intensity of the signal measured as total photons per second per square centimeter per steradian (p/s/cm²/sr) was obtained. Mice received a single dose of 2.5 × 10⁶ HER2-CAR or unmodified T cells i.v. on day 14 post-tumor cell inoculation. To monitor the in vivo persistence of HER2.CD28.z and HER2.41BB.z CAR T cells, we labeled the effectors with firefly luciferase.