[Coral-List] RE: Optimal fixatives for coral tissues

[James Rolfe Guest]

Hi Shelley,

You're right when you say there is no standardised method for histological processing of coral tissue. I'm not sure if there is one optimal method that can be applied in any situation. Personally, I think it is better to apply the most appropriate method for the job in hand. 

The most common method is to use 10% seawater formalin, embed in parafin wax and stain with Mayer's Haemotoxilin and Eosin. I have been looking at the gametogenic cycles of certain corals and got better results from using a 5% seawater-formalin-Zenker's fixative (add 50g zinc chloride, 25g potassium dichromate per litre of solution) and stain with Aniline Blue/Azocarmine G as used by Glynn et al (1994), Marine Biology 118 p 191-208. A really useful book with many fixation and staining techniques is Luna (1968) Manual of histological staining methods of the armed forces institute of pathology, 3rd Ed, McGraw Hill, New York. 

If you plan to do histopathology combined with molecular techniques (e.g. fluorescent in situ hybridisation) you might fix with 4% paraformaldehyde. Gluteraldehyde/Osmium gives very good preservation and is generally used for doing EM work, but has been used for examination of coral diseases by light microscopy, see Bythell et al (2002) Histopathological methods for the investigation of microbial communities associated with disease lesions in reef corals LETT APPL MICROBIOL 34 (5): 359-364.

I hope these comments are useful. Please let me know if you have any other questions.

Cheers,

James    



Message: 2
Date: Tue, 13 Jan 2004 10:06:09 +1000
From: Shelley Anthony <s.anthony at gbrmpa.gov.au>
Subject: [Coral-List] Optimal fixatives for coral tissues??
To: coral-list at coral.aoml.noaa.gov
Message-ID: <40033671.6070301 at gbrmpa.gov.au>
Content-Type: text/plain; charset=us-ascii; format=flowed

Dear Coral-Listers,
  I would very much like to hear from scientists who are dealing with 
different fixation techniques for histological work on hard corals.  I 
have been using standard 5% seawater-buffered formalin, per 
recommendation from my supervisor.  However, I have had numerous 
conflicting suggestions from histologists, lab technicians and other 
sources that the following fixatives give better results: 4% formalin, 
10% formalin, 70% ethanol, and 73% ethanol, glutaraldehyde, Helley's 
fixative, etc.  (However, most of these people were not necessarily 
working on corals.)   The publications I have read also do not seem to 
have a standard procedure, or the fixative may be dependent on the 
intended usage.
      My questions are:  Is there an optimal fixative specifically for 
hard corals that can be used for several different kinds of 
histological/histopathological/microscopic procedures?  What kind of 
results have you gotten when using different fixatives?  I would 
appreciate any information you can share.  Please reply directly to my 
email address.  Thanks!
Cheers,
Shelley

-- 
Shelley L. Anthony <shelleya at gbrmpa.gov.au>

PhD Student, CRC Reef Research Center
School of Marine Biology & Aquaculture
James Cook University
Townsville, QLD 4811  AUSTRALIA

ReefHQ/GBRMPA
Townsville, QLD 4810  AUSTRALIA
(07)4750-0898

James R. Guest
PhD Research Scholar
Marine Biology Laboratory
Department of Biological Sciences
National University of Singapore
Blk S2, 14 Science Drive 4
Singapore
117543
Tel: (65) 68746867
E-mail: scip9051 at nus.edu.sg