Withd luciferin

Infected mice were treated according to the indicated schedules.
Benznidazole (BNZ – Elea Phoenix,) was prepared by pulverization of
tablets followed by suspension in an aqueous solution of 1% sodium
carboxymethylcellulose with 0.1% Tween 80 and delivered orally by gavage at a
concentration dosage of 100, 250 or 500mg/kg body weight. Each mouse received
0.2 ml of this suspension. Luciferase-expressing parasites were quantified in
mice by bioluminescent detection. Mice were injected intraperitoneally with
D-luciferin (150 mg/kg; PerkinElmer,) and anesthetized using 2.5% (vol/vol)
gaseous isofluorane in oxygen prior to imaging on an IVIS Lumina II imager
(Xenogen,) as previously described (32 ).
Quantification of bioluminescence and data analysis was performed using Living
Image v4.3 software (Xenogen,). Exposure time was 5 minutes.

In total, 5 × 10⁶ A549, MDA-MB-468, or MDA-MB-468/Luc cells
(1:1 PBS:Matrigel) were injected subcutaneously into the hindflanks of nude mice (NCR
nu/nu, Taconic). Tumors were allowed to form for 2–3 weeks. Nanoparticles (8.3
× 10¹² NP · kg⁻¹, 5% glucose) were
injected via the tail vein into tumor-bearing nude or immunocompetent
mice (BALB/c, Taconic). Tumors were harvested after 48 or 72 h and processed by the
Swanson Biotechnology Histology Core Facility. Briefly, tumors were formalin-fixed,
paraffin-embedded, sectioned (5 μm), deparaffinized, antigen
retrieved, and stained using DAPI, biotinylated hyaluronic acid binding protein
(Calbiochem), anti-CD44-FITC (Life Technologies), and streptavidin-Alexa Fluor 546 (Life
Technologies) or DAPI and anti-HIF1α-fluorescein (R&D Systems). Slides were
mounted and imaged using a Nikon 1AR Ultra-Fast Spectral Scanning Confocal Microscope.
Whole-animal imaging was performed using a Xenogen IVIS Imaging System (Caliper) with
_D-luciferin (150 mg kg⁻¹ ip, PerkinElmer) as a
bioluminescent substrate. These experiments were approved by the Massachusetts Institute
of Technology Committee on Animal Care (CAC).