Hi
Could you use some thing like saturated chloral hydrate in water (you'll
need a drug permit), or a lactic acid/glycerol solution both are water
miscible. I use these on plant material quite often. The major problem
is having to image in water, can you use an objective made for glycerin
immersion?
Russ
Michael Weber wrote:
> Dear all,
>
> first of all, thanks for the replies off- and online!
>
> I should have mentioned a bit more details in my initial post. The
> embryos get mounted in agarose and investigated with dipping
> objectives with water as medium, so the RI of the surrounding medium
> is around 1.33. I expect the RI of the embryo to be higher, so the
> limiting factor for penetration depth is diffraction between medium
> and embryo. But, there is nothing I can do about that, right? Clearing
> with i.e. BABB would not make any sense, if I put the samples back in
> water-like medium afterwards and do not use oil objectives anyway. So
> the imaging conditions are already as optimized as possible - is that
> a valid conclusion, or am I missing something?
>
> cheers,
> Michael
>
>
> Phil Hertzler schrieb:
>> Mike,
>>
>> Methyl salicylate (oil of wintergreen) also works well and smells
>> better than BABB. :-) Transition through 100% ethanol from aqueous
>> buffer. I've stored samples over 15 years in MS without loss of
>> fluorescence.
>>
>> Best regards,
>>
>> Phil
>>
>> At 01:21 PM 10/7/2009, you wrote:
>>> Mike
>>> This is a review that describes our procedure of clearing mammalian and
>>> insect tissue with BABB. Reprints are available on request
>>>
>>> Zucker, R.M.Technical note: Whole insects and Mammalian Embryo Imaging
>>> with Confocal Microscopy: Morphology and Apoptosis. Cytometry 2006 69A:
>>> 1143-1152
>>>
>>> Best wishes
>>> bob
>>>
>>> Robert M. Zucker, PhD
>>> U.S. Environmental Protection Agency
>>> Office of Research and Development
>>> National Health and Environmental Effects Research Laboratory.
>>> Toxicology Assessment Division
>>> Telephone: 919-541-1585 Fax: 919-541-4017
>>> e-mail: [log in to unmask]
>>>
>>> Mail address: USEPA,ORD,NHEERL,TAD
>>> Developmental Biology Branch ( MD 67)
>>> Research Triangle Park, North Carolina, 27711
>>>
>>> Shipping address:
>>> 2525 E.NC Highway 54
>>> Durham, NC, 27713
>>>
>>>
>>>
>>>
>>>
>>> From: Michael Weber
>>> <[log in to unmask]>
>>>
>>>
>>>
>>> To:
>>> [log in to unmask]
>>>
>>>
>>>
>>> Date: 10/07/2009 11:56
>>> AM
>>>
>>>
>>>
>>> Subject: optical clearing of
>>> tissue
>>>
>>>
>>>
>>> Sent by: Confocal Microscopy List
>>> <[log in to unmask]>
>>>
>>>
>>>
>>>
>>>
>>>
>>>
>>>
>>> Dear list,
>>>
>>> I am looking for advice on optical "clearing" of fixed tissue before
>>> staining it and using it for light microscopy. Actually "tissue" is not
>>> the precise term, since I would like to clear whole fly embryos. This
>>> process seems to be well established in histology, i.e. using Xylene. I
>>> also found a commercial product called "Histo-Clear" (National
>>> Diagnostics), which claims to preserve tissue structures rather well,
>>> while being less nasty compared to Xylene. Did you guys ever use
>>> something
>>> like that? Any input welcome.
>>>
>>> cheers,
>>> Michael
>>
>> ------------------------------------------------------------------------
>> Philip L. Hertzler
>> Professor
>> Central Michigan University
>> Dept. of Biology, Brooks Hall 217
>> 200 Library Dr.
>> Mount Pleasant, MI 48859
>>
>> Phone: (989) 774-2393
>> Fax: (989) 774-3462
>> Email: [log in to unmask] or
>> [log in to unmask]
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