axolotl problems

Saul Leon Zackson zackson at acs.ucalgary.ca
Thu Jan 5 12:14:18 EST 1995


> 
> 
> 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.).

Saul Zackson
zackson at acs.ucalgary.ca




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