Balb c

BALB/c and NOD/SCID female mice, 6–8 weeks of age, were purchased from SLAC Laboratory Animal (Shanghai, China). The Peabody mjr strain of B. microti was obtained from ATCC and maintained through infection of BALB/c by intraperitoneal infection of parasite-erythrocytes. Briefly, mice were intraperitoneally administered 1 × 10⁷ erythrocytes infected with B. microti.
Parasitemia was detected in thin blood smears stained with Giemsa using a light microscope. Parasitemia was further evaluated using flow cytometry with SYBR Green I dye (Somsak et al., 2012 (link)). Briefly, 2 μl of blood obtained from the tail of mice infected with B. microti was collected into a 1.5 ml tube containing 500 μl of a premixed stock of glucose-PBS-EDTA (pH 7.4). SYBR Green I was added to the tube at a 6 × concentration and the mixture was incubated at 37°C in the dark for 30 min. Parasitemia was determined using a FACSCalibur (BD).

Female BALB/c and SCID CB17-Prkdcscid/J mice (age 6–8 weeks) were purchased from Jackson Laboratories, and maintained according to the IUCAC Guidelines of Emory. 4T1, a BALB/c mouse breast cancer cell line was obtained from ATCC (Manassas, VA). The protocol was approved by the committee on the Ethics of animal Experiments of the University of Emory (Permit number: A3180-01). Stably transfected luciferase-expressing 4T1 was a gift of Dr. Lili Yang. 4T1 cell lines were cultured in RPMI 1640 (Life Technologies, Grand Island, NY) and Dulbecco's modified Eagle's medium (DMEM/hi glucose) supplemented with 10% fetal bovine serum enriched with 0.4 mmol/L of sodium pyruvate, and antibiotics (penicillin, streptomycin), respectively. Female BALB/c or SCID mice were injected subcutaneously in the left 1rd mammary fat pad with 1 × 10⁶ cells viable 4T1 tumor cells in 100 μL phosphate-buffered saline (PBS). Mice were monitored every other day to evaluate tumor growth. Treatment of tumor-bearing mice began when tumor growth was confirmed with BLI.

All animal studies were approved by the MIT Institutional Animal Care and Use Committee (protocol 0619-032-44) and were conducted in compliance with institutional and national policies. The 7- to 9-wk-old female mice (BALB/c, Taconic) were dosed with either SP (NCTC 7466), KP (ATCC 43816), HI (ATCC 33391), PVM (ATCC VR-1819), or influenza (Influenza A/PR/8/34 (H1N1), Charles River). Nanosensors were synthesized by CPC Scientific. ABNs were dosed in mannitol buffer (0.28 M mannitol, 5 mM sodium phosphate monobasic, 15 mM sodium phosphate dibasic, pH 7.0 to 7.5) and deposited into the lungs by intratracheal instillation (50 µL total volume, 20 µM per ABN). After 1 h, the bladder was manually voided, the urine was discarded, and the mice were put into a collection chamber for the next hour. Two hours after ABN administration, the bladder was manually voided and the urine was collected, along with any urine that was produced in the collection chamber. These urine samples were then sent to Syneos Health for liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. Reporter quantification by LC-MS/MS was performed as previously described (26 (link)).

The 4T1 (ATCC^® CRL-2539™) cell line, initially derived from spontaneous breast tumor growth in a BALB/c, was purchased from American Type Culture Collection (ATCC, USA). 4T1-R was derived from 4T1 (ATCC^® CRL-2539™). They were grown in RPMI 1640 medium supplemented with 10% fetal bovine serum (FBS), 100 U/mL Penicillin with 100 μg/ml Streptomycin, and 4 mM glutamine. Cells were cultured in 5% CO₂/95% air at 37°C in a humidified chamber and were split every 2 to 3 days. All cells tested negative for mycoplasma using the MycoAlert™ Mycoplasma Detection Kit. All materials were purchased from Lonza (Basel, Switzerland).