Specific pathogen-free (SPF) 7-9 weeks old female C57BL/6 mice were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. (licensed by Charles River). All mice used in this study are in good health and are not involved in other experimental procedure. Mice were housed with 5 companions per cage. All animals were allowed free access to water and standard chow diet and provided with a 12-hour light and dark cycle (temperature: 20-25°C).
To prepare inocula for infections, P. aeruginosa strains were grown in LB (Δ3, Δ5, Δ8) or LB with10 mM D-Glu (Δ6, Δ9) overnight, and then subcultured in fresh medium, grown at 37°C to an OD600 of 1.0. The cells were harvested by centrifugation and pellets washed twice were suspended in sterile 0.9% NaCl, adjusted to 5 × 108 CFU/ml. For vaccination, mice were immunized with indicated aPA strains [with a total volume of 20 μl (5 × 107 CFU bacteria) per mouse] via intranasal route at biweekly intervals. As an intranasal vaccination control, recombinant RBD protein (GenScript, Z03483) was diluted with PBS, and mixed with an equal volume of curdlan adjuvant (20 mg/mL) (24 (link)). Serum samples were collected after vaccination as indicated in figures legends.
To prepare inocula for infections, P. aeruginosa strains were grown in LB (Δ3, Δ5, Δ8) or LB with10 mM D-Glu (Δ6, Δ9) overnight, and then subcultured in fresh medium, grown at 37°C to an OD600 of 1.0. The cells were harvested by centrifugation and pellets washed twice were suspended in sterile 0.9% NaCl, adjusted to 5 × 108 CFU/ml. For vaccination, mice were immunized with indicated aPA strains [with a total volume of 20 μl (5 × 107 CFU bacteria) per mouse] via intranasal route at biweekly intervals. As an intranasal vaccination control, recombinant RBD protein (GenScript, Z03483) was diluted with PBS, and mixed with an equal volume of curdlan adjuvant (20 mg/mL) (24 (link)). Serum samples were collected after vaccination as indicated in figures legends.
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