I have been doing immunofluorescence on paraffin-embedded tissue sections and I was wondering if we can still dehydrate the fluorescence labelled tissue sections and mount them exactly like we do for immunohischemistry. If this process is possible, what are the do's and dont's?
Thank You for your help!
As far as I know fluorescence would be destroyed by the organic solvents and mounting medium. You have to use special aequous mounting media, that support the fluorescence.
Although I have not personally tried it, but have been intending to do so. It is conventional knowledge that use of aqueous mounting medium is necessary for preventing photobleaching and preserving fluorescence. However, even at 4C in the dark the is gradual loss of fluorescence making most slides useless in about a weeks time. I have come across an article by Espada et al (Histochem Cell Biol. 2005 Mar;123(3):329-34) mentioning that they have used normal dehydration and mountings for FFPE sections and have stable long lasting fluorescence. Maybe you can try it. Do let us know how it turned out.
The one time I did try it - I got no fluorescence at all. I do not have access to the full text of that paper. plant get hold of it one day and try it our again.
@ Deep Kumar Raman : you should have access to the (whole) article at:
Article Non-aqueous permanent mounting for immunofluorescence microscopy
.Best regards and wishes, good luck, WM
@ Monyer Abd Al-Fatlawi :
for convenience (hoping it IS the article you meant indeed)::
Sanchez, NP, Pathak, MA, Sato, S, Fitzpatrick, TB, Sanchez, JL, Mihm, MC Jr. Melasma: a clinical, light microscopic, ultrastructural, and immunofluorescence study. J Am Acad Dermatol. 1981;4:698–710.
Abstract
Melasma is an acquired brown hypermelanosis of the face. Although it is thought that melasma is associated with multiple etiologic factors (pregnancy, gastric, racial, and endocrine), one of the primary causes of its exacerbation appears to be exposure to sunlight. Three patterns of melasma are recognized clinically: (1) a centrofacial pattern, (2) a malar pattern, and (3) a mandibular pattern. Examination of patients with Wood's light (320--400 nm) is useful in classifying the specific type of melasma in correlation with the localization of pigment granules (melanosomes) in the epidermis and dermis. Four types of melasma are described on the basis of Wood's light examination: (1) an epidermal type, (2) a dermal type, (3) a mixed type, and (4) a fourth type, described in patients of dark complexion, in which the lesions, for lack of contrast, are not discernible on Wood's light examination, perhaps due to the increased number of melanosomes in the normal skin of black individuals. Light, histochemical, and electron microscopic studies revealed an increase in number and activity of type-specific melanocytes which appeared to be engaged in increased formation, melanization, and transfer of pigment granules (melanosomes) to the epidermis as well as to the dermis. The melanocyte seems to undergo a functional alteration brought about by a combination of multiple factors, including persistent sun exposure, hormonal factors, and genetic predisposition.
PMID: 6787100 [Indexed for MEDLINE]
PS: URL to access online / purchase [ PPV: $35.95 USD|PDF Download and 24 Hours Online Access: http://www.jaad.org/action/ecommerce?backUri=/article/S0190-9622(81)70071-9/pdf&offerId=146166,10.1016/S0190-9622(81)70071-9&addToCart=true ] the pdf: http://www.jaad.org/article/S0190-9622(81)70071-9/pdf.
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