[Histonet] Re: Hollande's fixative

Mon Mar 26 13:50:01 CDT 2007

Lynne A. Bell, HT (ASCP) at Central Vermont Medical Center in Barre VT asks:

 >>We are thinking of changing from Bouin's solution to Hollande's solution for our GI biopsies. I have looked at the MSDS and do not see any real issues. My biggest question is disposal of this solution. Can it safely go down the drain? Also, Freida Carson's book suggests washing the specimens before processing them. Does everyone do that?<<

 and Eric C. Kellar, Histology Laboratory Supervisor at Quest Diagnostics in South Florida replies that Hollande's fixative >>is stable and will decalcify small bone specimens. Tissue that is fixed with Hollande's can be stained successfully with most stains, and the cupric acetate in the solution stabilizes red blood cell membranes and eosinophil and endocrine cell granules so that less lysis occurs than with Bouin's solution.Thorough washing of the fixative prior to placing the specimen in a phosphate buffered formalin solution is necessary because the salts present in the solution will form an insoluble phosphate precipitate.<<
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 HOLLANDE'S FIXATIVE:

 Water 220 mL
 Cupric acetate 25 g
 Picric acid, saturated aqueous 780 mL
 Formalin 37% 100 mL
 Acetic acid, glacial 10 mL

 Dissolve the cupric acetate into the water.
 Add the picric acid and filter.
 Add the rest of the ingredients and mix.

 Original reference, cited at Stainsfile: Hollande: Comptes rendus hebdomadaires des séances et mémoires de la société de biologie [Paris] 1918;81:17. -- R.D. Lillie (3rd ed., 1965) cites Hartz, Am J Clin Path 1947;17:50 - the complete 1918 reference might be available here.

 present reference:
 http://stainsfile.info/StainsFile/prepare/fix/copper/hollande.htm

 Note that Bryan D. Llewellyn, compiler of Stainsfile, has recalculated the formula to use ready-made saturated picric acid in water rather than dry chemical, which because of its explosion hazard should not be in your laboratory!

 The formula is in Freida Carson's book, but I've just lent my copy to a couple of histotechs studying for the registry exam.

 Hollande's fixative cannot go down the drain because of the formaldehyde, the picric acid, and possibly the copper. (You should look for a better MSDS than the one you have.)

 Pre-packaged bottles of it are available commercially.

 In the many years I've done locum tenens pathology in many different laboratories, I have never seen Hollande's fixative in use. I'd be reluctant to handle it because of the picric acid, which is not only toxic and (when in anhydrous form) explosive, but which stains fingers, clothing, and anything else it can get to.

 If you were going to use Hollande's fixative, you would have to investigate its compatibility with your immunostains.

 Bob Richmond
 Samurai Pathologist
 Knoxville TN
 March 26th, 2007

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