Two-color fluorescence labeling in acrolein-fixed brain tissue
Keywords: 
sodium borohydride
endogenous peroxidase
inactivation
autofluorescence
ethanol
enhanced penetration
hydrogen peroxide
aldehyde fixation
Issue Date: 
16-Dec-2009
Publisher: 
Histochemical Society
ISSN: 
0022-1554
Citation: 
Luquin E, Perez-Lorenzo E, Aymerich MS, Mengual E. Two-color fluorescence labeling in acrolein-fixed brain tissue. J Histochem Cytochem 2010 Apr;58(4):359-368.
Abstract
SUMMARY Acrolein is a potent fixative that provides both excellent preservation of ultrastructural morphology and retention of antigenicity, thus it is frequently used for immunocytochemical detection of antigens at the electron microscopic level. However, acrolein is not commonly used for fluorescence microscopy because of concerns about possible autofluorescence and destruction of the luminosity of fluorescent dyes. Here we describe a simple protocol that allows fine visualization of two fluorescent markers in 40-mm sections from acroleinperfused rat brain. Autofluorescence was removed by pretreatment with 1% sodium borohydride for 30 min, and subsequent incubation in a 50% ethanol solution containing 0.3% hydrogen peroxide enhanced fluorescence labeling. Thus, fluorescence labeling can be used for high-quality detection of markers in tissue perfused with acrolein. Furthermore, adjacent acrolein-fixed sections from a single experiment can be processed to produce high-quality results for electron microscopy or fluorescence labeling.

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