Xenogen ivis 200 system

Mice were first shaved then injected i.p. with 200 μl of 15 mg/ml of the luciferase substrate, d-luciferin (#LUCK; Goldbio) in PBS and imaged after 8 min with a Xenogen IVIS-200 system (PerkinElmer). Photon emission was detected with acquisition times ranging from 5 s to 3 min. Analysis of the images was performed using Living Image software (PerkinElmer) by obtaining average radiance per second per cm² of specified regions of interest.

After MR imaging, the animals were sacrificed and immediately, NIR imaging of the open abdomen was performed. Tumors and organs (heart, lungs, liver, spleen, kidney, stomach, and intestine) were then dissected and excised, and fluorescence imaging of excised tumor and organs was performed. For NIR imaging, a Xenogen IVIS 200 system (Perkin Elmer Inc., Waltham, MA, USA) was used. ICG fluorophore excitation (λ_excitation = 705–780 nm) and emission (λ_emission = 810–885 nm) filter sets were used. Using Living Image 2.5 software, regions of interest (ROI) were drawn for each organ/tumor and total radiant efficiencies (p/s)/(μW/cm²) were measured.

We used in vivo bioluminescence imaging to quantify tumor growth using the Xenogen IVIS-200 System (Perkin Elmer, Waltham, MA). Prior to each imaging session, mice were IP injected with sterile d-luciferin at 10μl per gram body weight. D-luciferin was prepared in PBS at 15mg/ml. After mice were injected, they were placed inside an oxygen rich induction chamber consisting of 2.5–3% isoflurane. All mice, regardless of anesthesia treatment, received the isoflurane as a necessity for performing imaging. Any effects of these repeated exposures would indiscernibly manifest across all treatment groups. The mice were imaged for 5 minutes post injection of d-luciferin. Mice were placed on their backs with ventral side up on the imaging platform. Anesthesia was maintained using nose cones with a 2% isoflurane flow rate. The IVIS imaging chamber consisted of a warming platform and a cryogenically cooled CCD camera to capture both a visible light photograph and a bioluminescent image. Imaging time points included at the time of injection (time 0) and on days 3, 10, and 14 post injection.

Tongue xenografting and experimental lung metastasis experiments were performed as previously described (Goldie et al., 2012 (link)). Animal studies were subject to Cancer Research UK and King’s College London ethical review and performed in accordance with an approved UK Government Home Office license. A dox-rich diet (Harlan) was used to induce miR-203 expression in vivo. Bioluminescent imaging was conducted using a Xenogen IVIS 200 system (Perkin Elmer).

Breast cancer tumors were established in BALB/c mice by intramammary injection of 1×10⁵ 4T1 cells. On day 10–14, mice were inoculated weekly by intravenous tail vein injection as follows: PBS control, DOX-NP (2.5 or 5.0 mg/kg); MPL (10 µg); MPL-pSi microparticles (5×10⁸); or the hybrid combination of DOX-NP/MPL-pSi particles. Alternatively, mice were injected intratumorally with PBS, MPL (10 µg), or MPL-liposomes (1000 µg lipid/6.25 µg MPL; 50 µl). Kineret (anakinra, 30 mg/dose; Amgen, Thousand Oaks, CA, USA) was administered 3 times per week by intraperitoneal injection. Tumor growth was monitored by caliper measurements 3× per week and based on luciferase expression weekly with the Xenogen IVIS-200 System (Perkin Elmer Inc., Waltham, MA, USA) following intra-peritoneal injection of 150 mg/kg RediJect D-Luciferin (Perkin Elmer Inc.). Mice were sacrificed 24–26 days after initiation of tumor growth, blood was collected by cardiac puncture, and tumor and spleen were collected for weight, size and immunohistochemical staining.