This article describes a method for infecting mice with Mycobacterium tuberculosis to study pulmonary tuberculosis. The process involves preparing a standardized aerosol inoculum and using a nebulizer to deliver the bacteria to the lungs of conscious mice.
Begin with a frozen stock of Mycobacterium tuberculosis, a pathogenic bacterium that causes pulmonary tuberculosis.
Thaw the vial and gently mix the suspension to disperse bacterial clumps and ensure a uniform concentration.
Dilute the required volume in sterile buffer to prepare a standardized aerosol inoculum.
Restrain a mouse in a mesh holder to limit movement, and place it inside the sealed aerosol exposure chamber.
Load the prepared inoculum into a syringe and inject it into the nebulizer unit attached to the chamber.
Activate the inhalation system, which uses compressed air to nebulize the suspension into fine, respirable droplets.
These droplets, containing viable M. tuberculosis, disperse into the chamber air.
The conscious mouse inhales the aerosol, allowing viable bacteria to penetrate deep into the lungs and initiate infection.
The mouse model of pulmonary tuberculosis is now ready for further studies.
Begin by warming Mycobacterium tuberculosis stocks of a known CFU titer. When the cells have thawed, use a 1 milliliter syringe fitted with a 27 gauge needle to carefully mix the suspension five times to disperse the bacterial clumps, avoiding the production of aerosols. Depending on the desired infection dose, transfer the required volume of the mycobacterial stock into a 50 milliliter tube containing sterile PBS to a final volume of 6 milliliters. Next, place the experimental animals individually into compartmented mesh baskets. Place the baskets into the circular aerosol chamber and close the lid of the chamber.
Attach the Venturi-nebulizer unit to the three stainless steel socket joints. Aspirate the mycobacterial suspension into a 10 milliliter syringe, fitted with an 18-gauge blunt needle, and carry the syringe to the aerosol chamber in a closed transport box. Now remove the nebulizer screw cap and carefully inject the mycobacteria suspension into the unit, taking care to avoid aerosol generation.
Discard the syringe into a sharps container containing 2% Buraton, and then seal the nebulizer.
Run the inhalation apparatus.
When the cycle is complete, confirm that the mycobacteria suspension has been nebulized completely.
繁體中文
