I am currently injecting s.c LLC-1 cells or E077 cells into C57BL/6 mice.
I am injecting from 100 000 cells to 300 000 cells. The cell viability is around 80-90% before injection. The cells also are collecting from not very confluent plates (70-80%) and they grow very well in vitro. So they are heathy and not having any problem. I inject the cells in PBS.
However, many of mice does not develop the tumor and they are health after a month. On the other hand, when I am using 3 million cells, they grow in almost all the mice.
This problem also happens for my colleagues as well and we have no idea what would be the possible reason. I should mention that a renovation is going near our animal area so we are suspected that the noise, that sometimes coming to animals area, might be the possible reason.
Hello Sina,
Did you try injecting with Matrigel? What I usually do is injecting the cells with a mixture of DMEM/F12 media and Matrigel. Works well for me!
Hope this might help.
Noise is not a valid factor.
Quick injections could help.
Cell clumping may affect injections (clogged needle+sending out only PBS), don't keep the cells in ice for too long (the cells must be active to fight immune cells as soon as injected).
Because some mice form tumors I think it is not the issue of cell line despite matrigel could enhance tumorigenesis.
Take the help of colleagues so that you can speed up injection times.
Your cell numbers are ok.
Minimize processing time by quick cell count, viability check (both can be clubbed).
Check mice strain+age.
Minimize trypsinization time (very important).
Good luck
I agree with Stephen. Try to use matrix gel. It will help to consolidate all the cells and making the cell attachment even stronger.
Thank you guys.
I have used Matrigel, but did not work.
I have also have tried SC without anaesthesia and injecting in maximum one hour after collecting the cells...!
Where do you inject?
It takes time...yes, first because local defense response .... play classic music to mute the noise... it may help
Make sure you inject intraperitoneal, not going into an organ...then microtumors should grow on the surfaces of several organs (diaphragm, gut, liver, etc).
I have dissected the mice after 21 days (the group in which tumor did not grow), and did not see any sign of metastasise or tumor cells on any organs! Mice looked very healthy and dissection did not reveal any sign of tumor cells on any organ!
you may have to wait more time....5 weeks at least.
Otherwise you need to use thermographic tech to avoid killing animals before tumors grow
1. Rising cell viability >90%.
2. 1.0 X 106 to 2.0 X 106 viable cells are injected into the right flank of C57BL/6 mice. The tumors will growth up about one week.
Good luck.
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