I am going to prepare a suspension of OVA and alum in pbs for I.P administration in mice. what could be the challenges i will be facing and what precautions should i takeWhile preparing an ova and alum suspension, should the solvent be 10x pbs or do we need to reconstitute to 1x pbs
The typical protocol to make OVA/Alum that I use is:
1) (IN THE HOOD!) Prepare the OVA-Alum by adding 950ul PBS + 50ul OVA (stock in aliquots kept in -20℃ freezer, 10mg/ml in PBS) + 500ul Alum (Imject® Alum, Catlog# 77161, stored at RT, vortex before use, pipette VERY slowly to get accurate volume because it’s sticky) in an 2ml eppendorf tube. NOTE: This is for 10 mice as an example, and we need to
inject 150ul/mouse, 50ug/mouse.
2) Vortex for ~1min after adding everything. NOTE: This is the step where OVA binds alum, VERY IMPORTANT!
3) Then put it on the rotator (rotate slowly!) in the cold room for 30min.
4) Do i.p. injection of 75ul to each site of belly, and total two sites either left or right to the middle axis of the belly. NOTE: Do this way because we want to make sure the mice gets OVA immunized.
Hope this is helpful to you.
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