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Histol Histopathol. 2010 Aug;25(8):1017-24. doi: 10.14670/HH-25.1017.

Sudan Black B treatment reduces autofluorescence and improves resolution of in situ hybridization specific fluorescent signals of brain sections.

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1
Department of Pathology, Ribeirão Preto School of Medicine - University of São Paulo (USP), Ribeirão Preto - SP, Brazil. vivianecassia@usp.br

Abstract

Interference by autofluorescence is one of the major concerns of immunofluorescence analysis of in situ hybridization-based diagnostic assays. We present a useful technique that reduces autofluorescent background without affecting the tissue integrity or direct immunofluorescence signals in brain sections. Using six different protocols, such as ammonia/ethanol, Sudan Black B (SBB) in 70% ethanol, photobleaching with UV light and different combinations of them in both formalin-fixed paraffin-embedded and frozen human brain tissue sections, we have found that tissue treatment of SBB in a concentration of 0.1% in 70% ethanol is the best approach to reduce/eliminate tissue autofluorescence and background, while preserving the specific fluorescence hybridization signals. This strategy is a feasible, non-time consuming method that provides a reasonable compromise between total reduction of the tissue autofluorescence and maintenance of specific fluorescent labels.

PMID:
20552552
DOI:
10.14670/HH-25.1017
[Indexed for MEDLINE]
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