I am trying to collect Bronchoalveolar lavage from mice. I learned it from another lab where they used needles to insert into the trachea. But when I was trying, the liquid could leak and the needles sometimes got out of the trachea. I checked the protocols online and I found lavage tubes were used in some protocols.
Could anyone who is using lavage tubes to collect BAL from mice, please provide me wit further details (supplier etc.)?
We use a 22ga x 3/4" Luer adapter as a tracheal cannula for our BALs. It's essentially a 22ga blunt stainless steel needle with a female Luer hub that allows you to attach a syringe for injecting the lavage fluid. Here is our BAL protocol for mice:
Make a 1.5-2cm longitudinal incision on the ventral side of the neck.
Expose the trachea by blunt dissection a work curved forceps underneath the trachea.
Use the forceps to pull a 10-12cm piece of suture thread underneath the trachea.
Pull on the ends of the suture, cranially, to "stretch" the trachea in front of you.
Use a 20ga needle to poke a hole in the trachea as close to the larynx as possible.
Insert the 22ga cannula into the hole and advance approximately 5-8mm (stop short of the carina to avoid puncturing or tearing the bronchi).
Secure the cannula in the trachea by tying the suture around it with an overhand knot (the reason we use the steel cannula is so we can tighten the suture knot snugly to prevent leakage).
Attach a 4-way stopcock to the cannula with 2, 5cc syringes attached to it (one contains 5ml of lavage fluid and the other is empty, ready to collect the recovered BAL fluid).
Alternate injecting 1ml of fresh fluid and collecting 5X. Obviously, you can adjust the volumes based on your particular needs.
Dear Jacky,
use PE tubing to go over your needle (25 to 21 gauge), so that the edge of the needle does not puncture the trachea. This becomes a "catheter". Once inserted in the trachea after an incision (don't cut the whole trachea!), tie it with a thread (suture thread is great) that you passed under the trachea before inserting your catheter.
Good luck!
We use a 22ga x 3/4" Luer adapter as a tracheal cannula for our BALs. It's essentially a 22ga blunt stainless steel needle with a female Luer hub that allows you to attach a syringe for injecting the lavage fluid. Here is our BAL protocol for mice:
Make a 1.5-2cm longitudinal incision on the ventral side of the neck.
Expose the trachea by blunt dissection a work curved forceps underneath the trachea.
Use the forceps to pull a 10-12cm piece of suture thread underneath the trachea.
Pull on the ends of the suture, cranially, to "stretch" the trachea in front of you.
Use a 20ga needle to poke a hole in the trachea as close to the larynx as possible.
Insert the 22ga cannula into the hole and advance approximately 5-8mm (stop short of the carina to avoid puncturing or tearing the bronchi).
Secure the cannula in the trachea by tying the suture around it with an overhand knot (the reason we use the steel cannula is so we can tighten the suture knot snugly to prevent leakage).
Attach a 4-way stopcock to the cannula with 2, 5cc syringes attached to it (one contains 5ml of lavage fluid and the other is empty, ready to collect the recovered BAL fluid).
Alternate injecting 1ml of fresh fluid and collecting 5X. Obviously, you can adjust the volumes based on your particular needs.
Thanks, Philippe, So you insert the PE tube into the trachea? Actually I blunted the needles before I insert into the trachea. Someone also told me I should try to use thread, and I guess it is a good idea.
Thank you very much for the details, Bruce. I will practice more mice and I think I am doing better now.
Could be useful if you can access it (which I can't at the moment):
http://www.jove.com/video/1720/a-reversible-non-invasive-method-for-airway-resistance-measurements
Dear Jacky,
similar to Bruce Davidson I also used a luer cannula (IV cannula, in german "Venenverweilkanüle", e.g. Introcan from company Braun). 20 G worked fine in my hands for adult but not to old mice (around 8-12 weeks of age). If the mice are smaller you need a thinner cannula. I usally make a small incision with a scissor into the trachea or carefully puncture it with a needle, then insert the IV cannula plastic tube which can be directly attached to a 1 ml syringe and flush with PBS/EDTA several times. Don't forget to weigh your collecting tubes before and after if you want to calculate your BALF cells/ml.
Best regards and good luck for your experiments! Ann-Kathrin
Thank you pretty much for a perfect job attending to this issue. It's been a little bit difficult for me recovering enough PBS due to rupture of the trachea and I hardly collect the BALF from a single mouse up to three times. I will follow the protocols to the letter; hoping for a better result.
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