2.4. In vivo measurement of tumor growth and metastasis by IVIS imaging

Bioluminescent imaging was performed using a highly sensitive, IVIS Spectrum in vivo imaging system (PerkinElmer). D‐luciferin (Gold Biotechnology) was prepared per manufacturer's instructions in DPBS and filter sterilized with a 0.2 μm filter for a concentration of 15 mg/ml. For all in vivo imaging, D‐luciferin solution was intraperitoneally injected (150 mg/kg) in all mice for 10 min. Mice were then anesthetized with 2% isoflurane prior and during imaging with IVIS spectrum. Bioluminescence of D‐luciferin carried 4 T1‐Luc2‐RFP tumor cells was quantified using Living Image, in which regions of interest from displayed images were identified and quantified as the total flux which is the radiance (photons/sec) in each pixel summed or integrated over the ROI area.