Re: bone marrow smears

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From:"J. A. Kiernan" <>
To:lizellis <>
Content-Type:TEXT/PLAIN; charset=US-ASCII

On Thu, 9 Mar 2000, lizellis wrote:

> I have recently been told that bone marrow smears exposed to formalin vapors
> may interfere with the quality
> of the Wright's stain. Has anyone else out there ever heard of this or know
> the theory behind it ?

  With Wright's or any other stain of the Romanovsky-Giemsa type you
  are appying a mixture of blue cations and red anions. The former are
  attracted to negatively charged groups in the tissue, such as phosphate
  (DNA, RNA) and carboxylate (mostly of protein) ions. The anions are
  attracted to positive ions, mostly protonated amino groups of proteins.

  Formaldehyde combines with amino groups and reduces the general
  affinity of the tissue for anionic dyes. Consequently, after
  formaldehyde the blue components of a blood stain will predominate
  over the red (eosinophilic) components. This can be corrected by
  using a lower pH for staining and rinsing. You may have to try a
  series of buffers, from about 3.5 to 6.5. (Somewhere between 4 and
  5.5 is often good for paraffin sections of formaldehyde-fixed
  material.) The lowered pH acts by protonating more of the weakly
  basic amino groups, which are not present as cations when the
  ambient pH is closer to neutral - as in the usual pH 6.8 for
  Romanowsky-Giemsa staining of alcohol-fixed blood smears.

  The above is a simplified explanation and does not include all
  the reasons why blood stains provide their characteristic colours
  with cell nuclei, leukocyte granules, red cells etc. For more
  detailed information you will need to look in a textbook of
  haematology or histochemistry written since about 1980. Older
  books have little to add to what's in the previous paragraph, and
  they contain outdated (and largely wrong) notions of the actions
  of various thiazine dyes present in the traditional staining

 John A. Kiernan,
 Department of Anatomy & Cell Biology,
 The University of Western Ontario,
 LONDON,  Canada  N6A 5C1

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