Thank you for a very informative post. Would you please recommend a
non-aqueous fixative?
Cyndi
________________________________
What are the "cyclic dextrins" that you are asked to stain in "formalin
fixed tissue"? Are they cyclodextrins, dextrins or dextrans?
If the fixative was mostly water, dextrins (and dextrans if present)
have probably all been dissolved out. These oligo- and polysaccharides
are soluble in water. Unlike proteins, they do not react with
formaldehyde to form insoluble products. Read on.
Cyclodextrins are made by the action of a bacterial amylase on starch.
The long chain of 1-->4-linked glucose units is broken down to shorter
chains (6, 7 or 8 units), with the ends of each chain joined 1-->4 to
make a ring. There are three types of cyclodextrin: alpha (smallest,
with 6 glucose units), beta (7), and gamma (8 units). In three
dimensions the ring is a shell or cage, with hydrophilic -OH groups of
glucose units on the outside and a hydrophobic interior. A hydrophobic
organic molecule can be enclosed within a cyclodextrin molecule without
chemical change. The complex, known as a clathrate, is soluble in water;
this provides a way to make an aqueous solution of an otherwise
insoluble hydrophobic substance.
The ordinary non-cyclic dextrins (3 types) are produced by either dry
heating (toast) or acid hydrolysis of starch. Properties vary with
method of production. They are soluble in water and precipitated by
addition of alcohol. The information in this and the previous paragraph
is from the Merck Index (which gives some references) and from an old
biochemistry textbook for medical students (West & Todd, 1956).
I do not have any authoritative statement about the solubilities of the
three cyclodextrins in liquids other than water. They may, like simple
dextrins, be insoluble in alcohols. You can be sure they won't dissolve
in xylene or paraffin.
Dextrins must not be confused with dextrans, which are bacterial
polysaccharides composed of glucose units linked 1-->6 (in contrast to
the 1-->4 of plants and animals). Dextrans are the protein substitutes
in traditional artificial plasma expanders (Rheomacrodex when I was
young; is it still used?) and they can end up in the renal tubules.
According to Lillie's big book, which cites a paper by Bob Mowry (Am. J.
Path. 29:523, 1953), water must be avoided at all stages prior to
staining dextran. That means a non-aqueous fixative and floating out
paraffin sections on 95% alcohol, not water. The reagents for staining
(periodic acid-Schiff) should be made up in alcohol rather than water.
Mowry's and Lillie's simple rules probably apply also to the
histochemical detection of cyclo- and other dextrins: Non-aqueous
fixative --> wax --> flatten on warm alcohol --> dewax with xylene and
then --> alcoholic PAS method. Does this help? Please reply to all.
John Kiernan
Anatomy, UWO
London, Canada
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