Re: Need help ... special processing. Plants!
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|From:||"J. A. Kiernan" <firstname.lastname@example.org> (by way of Marvin Hanna)|
Another rather long one from me. For the short form
of my answer to Maria's questions, skip to the last
couple of paragraphs.
John Kiernan, London, Canada.
On Sun, 23 Jan 2000, Maria Saenz wrote:
> We are in the process of helping out a doctor's son doing research and have
> been asked to process , cut, stain, etc his specimens. The only thing is
> that I've never processed anything but human tissue. The specimens we will
> be processing is of the plant kingdom variety ...
In the Lower 6th form in Britain (that's usually 2nd last year of
high school, in which you turn 17), a good while back, in the
Biology Lab, we cut lots of plant material by hand, using a
cut-throat razor as the microtome blade, elder pith or carrot as
the embedding medium, and forefinger & thumb as the chuck.
Everything was kept swimming with water or (?)70% alcohol, the
liquid in which the pieces were preserved. Nobody in our class
(of about 10) collected a section of finger or thumb. More
than 10 years later I came to know a distinguished professor of
botany in Cambridge who was hand-cutting sections of large
tropical fruits; he later wrote a book about his findings.
Hand-cut sections float into the puddle on top of the blade,
and are collected into a watch-glass and then wet-mounted
under coverslips. With practice we could get them one cell thick.
The sections were stained by running a drop of iodine solution
under the coverslip: starch grains blue, other things yellow
to brown. We also stained with phloroglucinol-HCl, which made
the lignin of xylem vessels red.
The preparations were temporary, and we made drawings of them.
At the end of the Upper 6th, the practical part of the public exam
(then and now known as A-level) consisted largely of making, drawing
and labelling a plant anatomy preparation of this kind and
doing a dissection to show one of the systems (circulatory in my
year) of a rat.
Each boy in the Biology 6th at every British grammar school in
those days carried around a honed and stropped razor, a scalpel,
pointed scissors etc. No doubt it was the same at girls' schools
because they did the same exams. Things were much more egalitarian
then than now. I never heard of a kid going berserk with a razor,
but wonder if things have changed. About 5 years ago, in London,
Canada, my youngest son, then 15 or 16, was told he could not
bring an ordinary domestic table knife to school to cut chunks off
his lunch time apple.
Freehand sectioning is not dead, or wasn't in 1999. There is are
at least two excellent textbooks of botanical microtechnique
published much more recently than John Kiernan's schooldays, that
say so. May I suggest that you give this advice to the the doctor
whose son's homework you seem to be doing.
1. Buy the following two textbooks. They're both available from
www.amazon.com and will answer all the questions.
Berlyn,GP & Miksche,JP (1976) Botanical Microtechnique and
Cytochemistry. Ames, IA: University of Iowa Press.
Ruzin,SE (1999) Plant Microtechnique and Microscopy. New
York: Oxford University Press.
2, Pay you an appropriate hourly sum for the time you take
reading these books, practising the techniques and doing
most of his son's assignment.
During the last 3 years I have compared conventional animal
fixing & processing into paraffin with recommended plant
procedures, especially those in Berlyn & Miksche. I'm using bits
of various house plants and garden weeds. A tentative suggestion,
based on this limited experience (only 20 specimens and fewer
than 200 slides, stained by 4 methods), is that plant specimens
are very easily damaged by cutting and by sudden changes in
the ambient solvent. You cannot move a 2mm cube of leaf from
an aqueous to a 70% alcohol environment without wrecking the
architecture formed by the cell walls. Stems and roots are
slightly more forgiving. These remarks, based on crude tests
of the botanists' assertions, indicate that animals and plants
are different, and require different methods of fixation and
processing. Both the recommended textbooks explain this. They
are by plant histology experts.
John A. Kiernan,
Department of Anatomy & Cell Biology,
The University of Western Ontario,
LONDON, Canada N6A 5C1
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