SEM with coverslipped sections

From:Gayle Callis

I asked Phil if a coverslip over section would negate SEM.  Here is his

Thanks to Phil for reply, he asked it be forwarded to Histonet. 

Gayle Callis 

At 04:12 PM 8/15/02 -0500, you wrote:
>Yes, the coverslip would negate the SEM. I was assuming that these 
>sections were not coverslipped. I should've checked that.
>If the coverslips can be removed without damaging the sections, then 
>SEM is still possible, after drying the sections. Otherwise the SEM 
>would just be looking at the top of the coverslip.
>I would've copied this to the list, but that wouldn't be proper 
>without asking -- if you don't mind, please do send this to the list.
>>What about SEM on coverslipped sections, I was trying to interpret what she
>>meant, and if there is a coverslip, doesn't that negate the SEM?
>>At 03:08 PM 8/15/02 -0500, you wrote:
>>>The staining won't affect the SEM imaging. Any embedding medium must
>>>be removed and the samples dried. If there is no embedding medium,
>>>and the samples are dry, then they're ready for coating. This could
>>>be the hang-up. If the samples must be left unaltered, then they
>>>cannot be coated. This means a field-emission SEM operating at low
>>>voltage (<5kV, likely around 1 - 2 kV) may be necessary. Although I
>>>have photographed fresh plant leaves uncoated at 10kV. It's best to
>>>coat (gold-coat or gold-palladium in a sputter coater) however. This
>>>coating can only be removed with effort and the use of methods that
>>>might at best destain the samples. Also, depending on the thickness
>>>needed, it might hide the staining, and it certainly will affect the
>>>colors of the stains. The coating is gray to gold, depending on
>>>thickness and surface texture. It is also somewhat transparent to
>>>opaque (if thick enough).
>>>The size of the samples and whatever they're mounted on could also be
>>>a problem, but this depends on the stage.
>>>If there is a chance of ruining things, then perhaps it's best to
>>>wait for your SEM person to return. Keep the samples dry and
>>>dust-free, and ignore them until then.
>>>>It's a dog femur samples implanted with IM rods coated with Titanium foam.
>>>>The slides were grind and polished up to 60 microns and stained with
>>>>Stevenel's Blue/ Van gieson Picrofuchsin. We originally measure the
>>>>using Bioquant Image analysis and would like to compare the porosity using
>>>>SEM, but we have stained all the slides. Yes!! we do have an expert on
>>>>but he is on vacation...want to be safe... I can't go back and repeat all
>>>>-----Original Message-----
>>>>From: Philip Oshel []
>>>>Sent: Thursday, August 15, 2002 3:15 PM
>>>>To: Dimaano, Nena
>>>>Subject: Re: SEM inquiry
>>>>Any longer reply will require more information about the slides,
>>>>preparation, what prep equipment you have access to, what you're
>>>>after, what can be done to the slides, and so on.
>>>>But you have access to lots of SEMs in your area, both commercial and
>>>>university, and personnel to run them and prepare samples.
>>>>>To all SEM experts:
>>>>>Can you do SEM on a stained slides?
>>>>>Nena Dimaano,MT/HT(ASCP)
>>>>>Advanced Technology- Preclinical and Biological Affairs
>>>>>Stryker Howmedica Osteonics
>>>>>300 Commerce Court
>>>>>Mahwah, NJ 07430
>>>>>tel: 201-831-5338
>>>>>fax: 201-831-6224
>>>>Philip Oshel
>>>>Supervisor, BBPIC microscopy facility
>>>>Department of Animal Sciences
>>>>University of Wisconsin
>>>>1675 Observatory Drive
>>>>Madison,  WI  53706 - 1284
>>>>voice: (608) 263-4162
>>>>fax: (608) 262-5157 (dept. fax)
>>Gayle Callis
>>Research Histopathology Supervisor
>>Veterinary Molecular Biology - Marsh Lab
>>Montana State University - Bozeman
>>19th and Lincoln St

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