all mice died after injecting 4% sodium thioglycollate?

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sxyan
sxyan's picture
all mice died after injecting 4% sodium thioglycollate?

I use 4% sodium thioglycollate to elicit peritoneal macrophage from mouse.
sodium thioglycollate was aged for 6 weeks.
i injected 1ml per mouse, but after 4hours, all the mice died.
did everybody have similar problem? Expect good answers, thanks so much!

genetika
genetika's picture
Hi,

Hi,
I have read that you had a problem of lethality when you injected thioglycolate in your mice. I have injected 2 ml of 4% of SODIUM THIOGLYCOLATE from Sigma (T0632) and in less than an hour mi mice have died. How did you resolve this problem? Could anybody help my??
Thanks!!

heehawmcduff
heehawmcduff's picture
Hi,

Hi,

If the mice died in less than an hour, is it possible that your intraperitoneal injection technique may be causing some damage to the liver or a blood vessel?  Are you able to successfully inject saline as a control? 

Additionally, how do you prepare your thioglycollate solution?

genetika
genetika's picture
First I made a 8% solution in

First I made a 8% solution in water and after autoclaving it I made I prepared a 4% solution in PBS.
Do you think that the problem is the solution?

Thanks

sxyan
sxyan's picture
Hi, my problem already solved

Hi, my problem already solved by changing new thiogllycollate solution.
I read many references and found that, the one that could stimulate peritoneal macrophage is not sodium thiogllycollate, but AGE (advanced glycation endproducts), and AGE is produced by sodium thioglycollate and agar. So I suggest you to change your reagent,  Thioglycollate Broth from Fluka can work well for me.
Good luck!

genetika
genetika's picture
Thank you for your help!!

Thank you for your help!!
Could you give me the reference of the thioglycolate you use and say me how do you prepare it exactly?
How much solution do you inject to the mice?
Thanks

sxyan
sxyan's picture
the cat no. is 70157, from

the cat no. is 70157, from Fluka, make 4% thioglycollate as the datasheet said, and inject 1ml to each 6-week-old mice.

genetika
genetika's picture
The datasheet says to suspend

The datasheet says to suspend 29 g of Alternative Thioglycollate Medium in 1000 ml of distilled water. I supposed that it is the 100% solution. How do you prepare the 4% solution? Do you make it in PBS1x or in distilled water?

Thanks 

sxyan
sxyan's picture
4% means the content of

4% means the content of sodium thioglycollate, I use distilled water.

genetika
genetika's picture
So if I understand well, you

So if I understand well, you make the solution suspending 232g of the Thioglycollate Broth, USP Alternative (#70157) in 100 ml of distilled water getting the 4% solution that you inject to the mice.
I am sorry for so many questions but I am afraid of doing something wrong and killing all my mice again.
Thanks
 

kunal H bhatt
kunal H bhatt's picture
Use thioglycollate broth not

Use thioglycollate broth not the sodium thioglycollate  Inject 1ml to mouse and use them after 3 day.

mkjl
mkjl's picture
Regarding Thioglycollate

Regarding Thioglycollate injections.

Use Brewers Thioglycollate medium B2551 from SigmaAldrich. Mix as in product sheet (then you get a 4% solution). The actual amount of Thioglycollate in your 4% solution is 0.5 g/l. If you mix T0632 from Sigma (as sxyan did) aldrich 4% w/v in PBS, you get 40g/l of thioglycollate (80x what you should).

In the product sheet on T0632 it is written that;
quote "Thioglycolate medium is frequently used in inflammation research to elicit a neutrophil and macrophage response in vivo." Notice "thioglycollate MEDIUM".

There is a reference to Potter, P. K., et al., J. Immunol., 170(6), 3223-3232 (2003).  Potter et al. uses Brewers Thioglycollate medium (B2551) and NOT thioglycollate T0632. I agree, that it seems from the product sheet of Sigma on T0632, that it can be used for macrophage recruitment. But it cannot. You kill your mice if you mix a 4% solution.

What can be used? Brewers Thioglycollate medium (B2551 from Sigma) and Thioglycollate Broth (USP alternative) (70157 from Sigma)

Brewers Thioglycollate medium (40.5g/1L = 4%)
Thioglycollate Broth (USP alternative)  (29g/L =3%)