Sensitive detection of hybridocytochemical results by means of reflection-contrast microscopy

Abstract

A new sensitive method for visualization of nonautoradiographic hybridization results in microscopic preparations is described. The method is based on the reflection of the incident light by diaminobenzidine precipitates deposited at the site of hybridization during an indirect hybridocytochemical procedure. The reflected light is detected by means of reflection-contrast microscopy. The applicability of the procedure is demonstrated with nucleic acid probes modified with 2-acetylaminofluorene groups. These in turn are localized in situ by an indirect immunoperoxidase reaction. Besides its sensitivity, this simple visualization technique possesses the additional advantages, over absorption and fluorescence microscopy, that it provides a total DNA counterstain and a chromosomal banding pattern.