Vivoglo d luciferin potassium salt

Isoflurane anesthesia was performed as described above, following a subcutaneous injection of VivoGlo D‐luciferin potassium salt (Promega, Fitchburg, MA, USA) from a 40 mg·mL⁻¹ stock in PBS. The dosage was calculated after weighting the animals prior to imaging (150 mg luciferin per kg animal weight). On the indicated days after injection (P3, P7, P10, P14, P28, P42), the anesthetized animals were imaged under maintenance of anesthesia using a Caliper Lumina II system (PerkinElmer, Waltham, MA, USA). Animals were imaged with their right side up, three to five at a time using full‐frame camera height. Images were gathered over 45 s each. Image visualization and quantification were performed using the living image analysis software 4.7.4 (PerkinElmer).

Under isoflurane anesthesia performed as described above, animals (n = 6, 10⁴ cells injected per eye) were injected subcutaneously (s.c.) with VivoGlo d-luciferin potassium salt (Promega), from a 20 mg/mL stock in PBS. Dosage was 150 mg luciferin per kg animal weight; animals were weighed immediately prior to each imaging session. Under maintenance anesthesia, animals were imaged on the indicated days after injection (P3, P7, P10, P14) on a NightOwl LB981 (Berthold, Oak Ridge, TN, USA) system. Animals were imaged on their left sides, 3 at a time until too large for the chamber, then 2 at a time using full-frame camera height. Images were gathered over 6 intervals of 5 minutes each, for a total of 30 minutes’ imaging. In all cases, image import, visualization, segmentation and quantification were performed using custom, in-house developed bioluminescence image analysis software.