ACh challenge. Sixty days post RSV inoculation, all animals from each group underwent ACh challenge in a whole body plethysmograph and lung resistance (RL) was calculated as described elsewhere (29). Dose response curves were generated using RL values at baseline (no aerosol exposure) and after challenge with 0, 0. 5, 1. 5, 5, 15, 50, and 150 mg/mL of ACh in saline. After completion of the ACh challenge, animals underwent euthanasia with an overdose of sodium pentobarbital (120 mg/kg, intraperitoneally).
administration to Saucony Boston
cell culture supernatant prepared similarly Saucony Jazz Original Charcoal
Lungs were extracted under aseptic conditions, weighed, and processed for viral plaque assays, nucleic acid extraction, and quantitative histology. Viral plaque assays. To determine titers of replicating RSV within guinea pig lung specimens obtained at 60 d post RSV inoculation, one lung lobe was frozen at in 5 mL of minimal essential media (GIBCO, Burlington, Ontario, Canada). Subsequently, serial 10 fold dilutions of lung homogenates were subject to viral plaque assay on HEp 2 cell monolayers with an agarose overlay, as previously described (8,15). For each animal, results were reported as number of pfu/g of lung tissue. Semiquantitative RT PCR.
Lung tissue .
to viral stocks) delivered to the lungs by intranasal instillation, as described elsewhere (15). As in previous studies with this animal model (8), guinea pigs that received the same volume of a tracer substance (India ink) confirmed that the inocula were delivered to all lung lobes (not shown). CPG ODN immunoprophylaxis. CPG ODN, Saucony Camo Type B (single stranded, code 2007, sequence 5 was purchased from Coley Pharmaceuticals, Inc. (Wellesley, MA). Based on our previous work with CpG ODN Saucony Men's Running Shoes
guinea pigs (21), animals underwent anesthesia with 3% inhalational halothane and were intranasally inoculated with 100 of CpG ODN dissolved in 300 of sterile saline, given biweekly over 4 wk, for a total of three doses. CpG ODN treated animals were inoculated with RSV 7 d after receiving the final dose of CpG ODN (21).
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