Re: [Histonet] fixing sponge larvae for in situs

From:Rene J Buesa

  Larvae of Coelenterata and Platyhelmintes have to be narcotized before fixation, or they will explode. This is what is probably happening to the sponges planulae larvae.
  They could be narcotized either with menthol or a 2% hydrate chloral solution.
  Afterwards they could be fixed with a hot mercuric-acetic solution. Perhaps NBF could be used also.
  So, the kid of the thing is to narcotize the larvae before fixing them.
  Hope this will help you.
  René J.

Gary Radice  wrote:
  I have a colleague who wants to do in situ hybridizations on sponge 
larvae (yes, sponges are animals and they have larval forms!). She 
tried fixing some in 4% paraformaldehyde in sea water, and the larvae 
exploded. She also tried Carnoy's fix, and 4% PFA in Millonig's 
phosphate buffer, and the same thing happened.

The only thing I could think of to suggest was dropping to 1 or 2% 
PFA in sea water. I've seen references to using Bouins for light 
microscopy or osmium tetroxide for TEM, but I don't think either of 
these approaches would be good for in situs.

Any other ideas?

Gary P. Radice
Department of Biology 804-289-8107 (voice)
University of Richmond 804-289-8233 (FAX)
Richmond VA 23173

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