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|From:||Lynn Gardner <firstname.lastname@example.org> (by way of histonet)|
This is a very interesting protocol! It was nice for me to see that there
are others who do work routinely with eyes as I think we are a rare
commodity. We have a protocol that is a little different and also works
very well and we are able to produce slides without wrinkles and that stain
beautifully with routine, special, immuno and insitu staining. If anyone is
interested I would be happy to share the information. We also have a
different way of grossing the eyes. Good Luck!
Lynn Gardner, HT(ASCP)
Supervisory Research Assistant III
The University of Iowa
Iowa City, IA 52242-1182
At 07:36 AM 2/12/99 -0600, you wrote:
>Here I am, behind this bush over here...
>I've posted Annie (in Africa)'s message below, as published in Microscopy
>Today, May '98.
>Thanks for the article Annie! You are getting MT are you not?
>Hints for fixing vertebrate eyes:
> In my experience (up to 6 eyes per week for about 10 years), the
>globes MUST be properly fixed. I learned this the hard way, when I was a
>Medical Technologist, in charge of the Neuropathology Laboratory at the
>University of Cape Town Medical School, Anatomical Pathology Department.
> We would 'lop' off the 'top' and the 'bottom' - like a breakfast
>soft boiled egg - after 24 hours in 10% buffered formalin. The cuts are
>made in line with the cornea after orienting the specimen properly. The
>inferior and superior pieces were processed routinely overnight if properly
>fixed - with excellent results.
> The roughly 8mm thick mid-portion should be handled according to
>the contents of the globe - sometimes another 24 to 48 hours fixation is
>necessary for tumor or blood filled globes - particularly if the tumor is
> Thereafter a long dehydration program is required:
> 3 changes of absolute alcohol, total time about 8 or 10 hours
> follow with a manual inspection to check for proper dehydration
> 6 to 8 hours total in 3 changes of 100 % xylene (or other clearing
> ca . 12 hours total in 3 wax changes
> embed routinely in paraffin wax.
> When sectioning use lots of ice and section at 4 to 6 micrometers.
>I used to float out on 10% alcohol (or weaker) before the warm water bath.
>My results were excellent, with very few eyes giving me problems. I
>have used the double embedding method as well (to teach students) and have
>found it to be a poor substitute for the careful, slow processing of eyes
>using the classic pathway.
> I have to admit that many of my colleges did not enjoy handling the
>eyes, claiming that they could not get the results which I got. I had many
>years of practice and an excellent teacher. In addition, my Pathologist had
>"an understanding" of the lengthy schedule and he in turn educated the
>ophthalmic surgeons and the oncologists to be patient if they wanted good
>Anne S. van Binsbergen, South African Institute for Medical Research,
>Chris Hani Baragwanath Hospital in Soweto, South Africa
>>Philip Oshel - somewhere out there - has a protocol for whole eye
>>processing which I e-mailed him about a year or so ago, for ? pulication in
>>Philip, my system 'crashed' and I was out of action for ages - lost all my
>>e-mail info (sadly) - please would you re-post that protocol - if you have
>>I found that pig eyes were a fairly good substitute for human eyes.
>>The secret to proper processing of multi-layered tissues is 'slowly and
>>Annie (in Africa)
>>Anne S. van Binsbergen
>>T.I.C. Anatomical Pathology
>>Chris Hani Baragwanath Hospital
>>PO Box 1038
>>tel: 011 4898711
>>cell: 083 4403343
>****be famous! send in a tech tip or question***
>Technical Editor, Microscopy Today
>PO Box 620068
>Middleton, WI 53562
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