I have recently begun working with BrdU to label proliferating cells in the VZ of pigeons (thus, this is a 24 hr survival) -- the quality of the IHC has been less than impressive, but that's another concern for another day. Today's concern is with regards to the dissolving of BrdU and injections. I typically do 50-100 mg/kg IM injections dissolved in 1 mL of saline; three injections, 2 hrs apart. Because this is a fairly concentrated injection I dissolve by stirring over heat (manufacture's suggestion). Between injections I take the remaining BrdU off of stir and heat. It seems that these stringy/crystal-like things form once the BrdU cools and when I re-stir the solution turns a milky white unless I re-apply heat. Ok, questions: 1) is this normal; has anyone else experienced this? 2) is repeated cooling and re-heating somehow affecting the BrdU, perhaps destroying it/making it inactive (which would account for the fact that I feel as if I am not seeing as many positive labels as I should)? 3) I have had some BrdU/saline solution left over (I make a bit extra) and, because I am cheap, I have placed it in a freezer. Will I be able to thaw the BrdU/saline solution and use it at a later time? 4) Finally, any suggestions regarding the preparation of BrdU would be greatly appreciated.
Hi,
could you inject intraperitoneally? We did this for rats (I know far away from your experimental model) and we used 5 mg/ KG. So perhaps this would make it easier for you.
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