>>> Well, to all the fans of the axolotl, I must admit that this is indeed an
> interesting breed. Unfortunately, however, I am experiencing
> difficulties in handling the embryos.
> I am looking for some advice on handling the embryos after I have removed
> the vitelline membrane. I am working in different concentrations of NAM
> and have found that by using this medium, I am having more success in
> removing the membrane without puncturing the actual embryo. This was a
> great improvement over working in the solution that the embryos came
> in. One problem still exists. The ectoderm of the embryo in contact
> with the plastic petri dish is deteriorating, leaving an embryo with a
> yolky bottom and a perfectly good animal hemisphere. This has happened
> over and over again, and I'm wondering whether it's the medium that I'm
> working in. I've tried using antibiotics (gentamicin), thinking that the
> problem might lie bacteria growing on the plastic petri dish. But this
> proved to not be the problem either.
> My suspicion is that the embryo is faced with the problem of becoming a
> flat glob of cells without the protective vitelline membrane. I am
> working on lineage studies of cilia and would like to see my embryos
> making it past stage 26-28 (according to Detlaff's table). Please give
> feel free to give me some advice. It would be greatly appreciated, as I
> am feeling really bad about these wasted embryos.
>> Steve Hua (the guy who switched from Necturus to the ever popular
> Ambystoma mexicanum.)
>> <u9114317 at muss.cis.mcmaster.ca>
Here's the solution:
Dissolve agarose to about 2% in 15% HBSt by boiling. pour a layer
into a plastic petri dish (after the agarose has cooled to about
55-60 C). Allow agarose to harden, then Fill dish with 15% HBSt, and put demembranated embryos
in there. Epidermis won't stick to the agarose. (Concentrations
are very approximate, eyeball guesses are good enough.).
zackson at acs.ucalgary.ca