Ivis lumina 2 imaging system

Parasite liver loads in live mice were quantified by real time in vivo imaging as previously described^{101 (link), 102 (link)}. Liver stages were visualized and liver loads quantified by measuring luciferase activity of parasites in whole bodies of mice at 44 h after injection of sporozoites using the IVIS Lumina II Imaging System (Perkin Elmer Life Sciences, Waltham, USA). Before measurements the fur from bellies of the mice were shaved and during measurements mice were anesthetized using the isofluorane-anesthesia system (XGI-8, Caliper Life Sciences, Hopkinton, USA). D-luciferin was dissolved in PBS (100 mg/kg; Caliper Life Sciences, USA) and injected subcutaneously in the neck. Measurements were performed within 8 minutes after the injection of D-luciferin. Quantitative analysis of bioluminescence of whole bodies was performed by measuring the luminescence signal intensity using the ROI (region of interest) settings of the Living Image® 4.4 software.

Mice were injected i.p. with 150 mg/kg XenoLight d-luciferin – K⁺ Salt (PerkinElmer, U.K.). Mice were left for 10 min prior to the induction of anesthesia using isoflurane and bioluminescence measurement using an IVIS Lumina II Imaging System (PerkinElmer). Average radiance for areas of interest in bioluminescence images were calculated using Living Image 4.2 Software (PerkinElmer). For an in vitro measurement of bioluminescence using cultured cells, cells were transferred to black 96-well tissue culture plates before the addition of d-luciferin at a final concentration of 150 μg/ml. Cells were incubated for 10 min before imaging.

For MCF‐7 tumor model establishment, 6–8‐week old female BALB/c nude mice purchased from Xiamen University Laboratory Animal Center were first subcutaneously implanted with 60‐day release 17β‐estradiol pellets (Innovative Research of America) in the right side of the neck. Three days later, 2 × 10⁶ MCF‐7 cells were injected subcutaneously in 1:1 PBS:Matrigel (BioCoat) into the right leg of the nude mice. Tumors were measured by caliper two or three times per week.
When MCF‐7 tumors grew to a volume of 75–100 mm³, the mice were randomly divided into two groups (n = 6) and given palbociclib (150 mg/kg diluted in 50 nM sodium D‐lactate) or sterile water daily by oral gavage. Magnetic resonance imaging was performed before (0 days) and 7 days after treatment using a 9.4T BioSpec MicroMRI (Bruker), and fluorescence imaging was also performed using the IVIS Lumina II imaging system (PerkinElmer) at the observation time points after intravenous injection with CPP30‐Lipo/CDKACT4 (10 μl/g body weight, containing 30 μM CDKACT4). After imaging, tumor tissue was collected for IHC staining.