Hi Xuecai
I am new to IUE technique and I tried quite many trials following your protocol, using E14.5-E16.5 C57BL/6J mice and giving te pulse with 25V, 980msec off, 5 pulses, trying to target the prefrontal cortex. However, either the mothers loses 4-5g weight 1-day post-surgery (which probably indicate the death of embryos), or the embryos will die in P2-3. I am really frustrated and hope if you can give me some advice:
1) Do you think holding the embryo too tight might lead to such severe high mortality rate? I usually try to hold it using a cotton swab as one of my finger to help me push the embryo towards the uterine wall. I usually hold it as tight as when I insert the micropipette towards embryos head, I will use some force such that the insertion would not push the embryo away from uterine wall. Am I right or did I still enforce too much pressure on embryos?
2) I usually will rotate the embryos with cotton swabs to a position that I can see their head more clearly, do you rotate the embryos? I am thinking if the rotation itself might disrupt the embryos and kill them.
3) I usually will leave the embryos locating at the edge of the uterine horn not receiving IUE. May I ask for your criteria to choose which embryos to do/not do IUE that ensure higher embryo/newborn survival rate?
3) As I saw from your protocol, your insertion is nearly perpendicular to the lateral ventricles. I also see others insert from the area near the eye and in elevated angle for about 35 degree, 20degree angle to the left from the brain midline. Did you try this angle too and what will you comment on these 2 different angles for insertion?

Thank you so much for your protocol and everything!
Best regards
Beth
M.Phil student 6/2/2020 4:24:34 AM Reply