Hi, researcher.
It's a bit urgent that I have to inject tumor cells intravenously in 10 days. I am now practicing tail vein injection with a success rate of around 50%. I am still don't have confidence in this technique.
Here is the brief protocol. I heated the mice for 10 min under the UV that is commonly used for reptile and then fix the mice into the barrel. I can easily find the lateral tail vein but kept failing the injection. Problem is that my technique often results in the swelling of the tail and spread of the PBS that made worse the finding of the vein next time.
I know practice makes perfect, but in this case, can I use retro-orbital injection for single-cell suspension of tumor cells? Do you guys have any tips for tail vein injection? The current tip that I have is like when I poke the tail sometimes It felt relatively smooth. If I retracted the needle (25 G or 27 G), I can see the drop of blood. If I don't retract the needle, feeling "Oh, the needle seemed to be located in the vessel", I increased the angle of the needle and pushed the needle just a little bit, and then injected PBS. Sometimes, it worked with no resistance.
I want to succeed in this experiment.
Thank you.
For black mice with dark tail, make sure you don´t sit in direct light - that makes the vein disapear. Put a cloth or something over the tail and you will see how the vein sort of "pops" out.
For white mice with a light tail you will see the veins easier but still, direct light might fool you so be in a light room but away from direct light.
In both cases, use a 1ml insuline syringe 29G, have the bevel facing up and make sure to just go under the surface. Once in the vein you can "wiggle" the needle very gently and you can see the blood flowing around the tip.
As you mentioned, you need to heat up the animals first - but if the animals are to hot the i.v injection will be unsuccessful. Heat them up gently but for longer periods and you can also use a tourniquet (use your fingers) and press at the base of the tail to make the vein pop out more.
Also- always begin at the end of the tail. That way you can try more times up towards the base.
Last but not least - please do not use retro orbital.
Maria Valsjö Thank you. I appreciate it. I don't know how many times I watched this video. It's super-quick and meticulous, and mice did not resist against injection.
Myungchul Kim No, they rarely resist - other then of course I inject something that is irritating. The key is to be gently but firm and give the animal a comfortable quarter while doing this. They don't like open spaces so by putting a cloth over them gives them a safer place and also as a I mentioned, helps you so see the vein easier. I also prefer this type of restrainer - the animal is not severely restraint but can move around freely (except from the tail)
Good luck!
Maria Valsjö and guys, Thank you.
Could I have one more question? I will inject 100 ul of tumor cell suspension. If the tail vein injection is failed, it will result in the spread of PBS into the tissue, swelling, and make the vessel disappeared.
If you are not confident that the needle is correctly located in the vessel, do you retract the needle? sometimes, when I retracted the needle, there is blood coming out of the vessel. It means that I touched the vessel but failed to locate it. During these processes, I lost my samples too.
I appreciate your response.
Myungchul Kim You can try to aspirate and see if blood come into the syringe, otherwise my best tip is to have good light and try to see the tip of the needle - you can se the blood around the needle. Or, simply tro to inject - if you are in you are in you will feel it direct. If you are not you will feel resistance.
Bet tip is - wiggle the needle around slightly to see the blood around the tip. S.C you won´t see a thing.
How old are the mice and what strain do you use?
Maria Valsjö Thanks a million.
They are all 6 to 10 weeks old (C57BL/6J).
I have used the way "simple injection and no resistance detected". The disadvantage would be the injection of unwanted tumor cells into the tail, like a by accident. Now, I am using PBS for practice so it's fine.
Today I will try to test the "wiggle the needle" and will see the blood on the tip of the needle. To make sure, you mean when I inject the needle with bevel up, I should see blood on the bevel and move the needle slightly right or left side (not forward and backward or up and down) I could still see the blood on the bevel.
Nowadays, the success rate is being increased.
I appreciate it.
Maria Valsjö Now, I can understand what you said except for "wiggle". The good news is that I could inject 90% of mice, though I have to give it a shot several times. Today, I tried to evaluate the blood flow by doing needle back and forth or left to right and even using direct light at this moment. I am not familiar with this. I expected red blood on the bevel of the needle. Could you please explain the wiggling using other words?
Thank you so much.
Myungchul Kim What I mean with wiggle is just what you explained - to gently move the needle a bit from side to side in the vein, and also up and down (but not in and out) to see the blood moving around the tip of the needle.
Glad to hear about your success!
Maria Valsjö Again, thank you so much. I will keep practicing it.
Myungchul Kim
Sincerely,
Maria Valsjö I will do tail vein injection tomorrow. Do you raise the temperature of tumor cell suspension at least to room temperature?
Myungchul Kim, No - that way you might kill the cells. Keep the cells on ice.
Hi, Maria Valsjö. I ended up injecting tumor cells via tail today. Among 15 mice, 14 mice were taken the injection with no resistance. Although I couldn't achieve the 100% success rate, I think it could be sufficient to study a new phenotype.
Thank you. I appreciate it. I learned a lot from you.
Sincerely,