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Aseptic techniques are used th
Significant breakthroughs in stem cell identification are continuously being made. To translate these discoveries, however, novel methods for cellular delivery must be devised. Here I report that the airways provide a safe route for stem cell transplantation to the lungs.
Cite this Article
Peter, Y. Tracheotomy: A Method for Transplantation of Stem Cells to the Lung. J. Vis. Exp. (2), e163, doi:10.3791/163 (2007).
- Recipient mouse and donor cells
- Operating platform with tape and suture thread
- 1cc syringe
- 23G needle
- Anesthetic: avertin 25 mg/ml (alternatively ketamine 9 mg/ml and xylazin 1 mg/ml)
- Hair clipper with #40 blade
- Sterile gloves
- Alcohol swabs
- Sterilized surgical instruments
- Suture thread
- 22G catheter
- 200 ul pipette (sterile)
- Sterilized pipette tips
- Antibiotic glue ointment
- Sharpie container
Aseptic techniques are used throughout this procedure.
- Weigh mouse.
- Intraperitoneal injection of anesthetic (250 mg/kg avertin or alternatively ketamine 90 mg/kg and xylazine 10 mg/kg) at doses sufficient to allow spontaneous breathing, but provide surgical-concentration anesthesia. For surgery to commence the toe pinch was used as an indicator of deep pain with no response from the anesthetized animal.
- Lay mouse with incisors tucked in suture thread and tape tail to stabilize.
- Clip hair using a #40 clipper blade.
- Change gloves (continue with sterile gloves).
- Disinfect twice with alcohol prior to skin incision.
- Sterilize area with an iodophore.
- Using sterile instruments make a small incision (<1 cm) parallel to the trachea.
- Carefully separate tissue from cartilage.
- A tie is inserted underneath the trachea.
- Gently raise tie upward and insert needle into exposed trachea.
- Remove needle and insert the tube smoothly, yet rapidly, until feeling slight resistance.
- Pipette cells (up to 50 microliters volume) directly into the catheter.
- Immediately remove tube and tie while liberating mouse from operating apparatus.
- If needed gently hold mouse upright (vertical) by clinging his anterior paws and move gently.
- Close the wound with one suture or antibiotic glue.
- Keep mouse warm.
- Buprenorphine (0.05 mg/kg) should be given as post-operative analgesia every 12 hours for the first 24 hours following this procedure.
|23 G needle|
|1 cc syringe|
|22 G catheter|
|antibiotic glue ointment|
|Anesthetic||Avertin 25 mg/ml. Alternatively ketamine 9 mg/ml and xylazin 1 mg/ml.|
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