The membrane was incubated for 3 minutes with the substrate (Lumi-Phos WB; Fisher Scientific, Pittsburgh, PA, USA) at a volume of 0.125 ml for every cm2 of membrane.
Spatiotemporal analysis of the flow field associated with individual isolated cells was achieved through seeding the fluid with 0.5 μm polystyrene microspheres (Invitrogen, USA) at a volume fraction of 2 × 10−4.
All compounds were dosed orally in a vehicle of aqueous 0.5% Natrosol™ (Ashland, Covington, KY, USA) weight/volume at a volume of 3 mL/kg.
Animals were mechanically ventilated (Ventilator Model 683, Harvard Apparatus, Holliston, MA, USA) at a tidal volume (TV) of 8 mL/kg and a respiratory rate of 40 to 45 breaths/min, which was adjusted to maintain an EtCO2 between 35 and 45 mmHg (Micro-Capnometer, Columbus Instruments, Columbus, OH, USA).
Triplicate cultures were grown in 96-well plates (Becton Dickinson, Franklin Lakes, NJ, USA) at a final volume of 200 µl/well.
The RT-PCR reaction was carried out at 37°C for 30 min on a C1000™ Thermal Cycler (Bio-Rad, USA) at a final volume of 20 μl, and inactivation of the enzyme was completed at 85°C for 5 min.
Seven hours after the cecal ligation and puncture (CLP), we reanesthetized each animal and intubated it with a beveled, 16-gauge angiocatheter and ventilated it with room air using a Harvard rodent ventilator (Holliston, MA, USA) at a tidal volume of 10 ml/kg and a frequency sufficient to maintain an arterial PCO2 between 35 and 45 mmHg.
Thermal hyperalgesia was induced by administering LPS (Sigma-Aldrich, St Louis, MO, USA) at 1 mg/kg at a volume of 5 μl/g body mass.
PG-based nanofluids were prepared by Nanophase Technologies (Romeoville, IL, USA) at 7% volume fraction without any dispersants, and were diluted to 1, 3, and 5% for all particle sizes.
Huh-7 cells were seeded in 24-well plates at a density of 4 × 10/well in quadruplicate and WST-1 reagent (Nalgene, Rochester, NY, USA) was added to each well at a volume up to 5% that of the medium.
PBMCs were plated in a 96-well plate (Corning, NY, USA) at a final concentration of 2.5×10/ml in an end volume of 200 μl per well.
