Hi, i am planning to measure some cytokines in mouse serum at different time points in my model and was wondering if anyone has done these retro-orbital blood collection and the amount that one can safely withdraw (3 times over the course of 30 days) without any adverse effects to the animal. thanks
0,5 ml can be withdrawn safely by an experienced handler using the hematocrit tube method, time spacing should be at least 1 week and the other side should be used for the subsequent sampling. Please be aware that in many cases pathological alterations can be caused (retro-bulbar oedema etc) even with very skilled sampling. So if you need pathological evaluation of the eye/optic nerve at the end of your study, better use satellite mice for blood sampling. So in your case, I would suggest, 3 x sampling per animal with 10 days spacing in between alternating the sampled eyes.
I used to collect 250ul every 2 weeks, and they suffered significant weight loss in between. Mice have only about 1.5ml of blood total so our protocol specified we weren't allowed to take more than 25% of that in any one experiment. Even then I thought it was a bit much. The mice had barely recovered their body weight after 2 weeks.
Here's a fairly conservative guide: https://www.nc3rs.org.uk/mouse-decision-tree-blood-sampling
Also, retro-orbital blood collection is very stressful for the mouse. It is much better to use sub-mandibular/facial vein puncture. See video: https://www.youtube.com/watch?v=niTVnEAHOko