Fluorophore´s role on the reliability of microorganism detection by Fluorescence In Situ Hybridisation (FISH).

Abstract

FISH has been applied in many ecological and phylogenetic studies becoming the method of choice for the direct detection and identification of microorganisms in their natural environments [1]. For reliable results, it is crucial to minimise or avoid background fluorescence and cellular autofluorescence but also to maximise the specific FISH signals obtained. Red-emitting dyes have been usually used for eliminating the problems of non-specific fluorescence interference. Therefore, in this work we evaluated the role of three of these fluorophores (Alexa Fluor 647 (AF647), ATTO 647N and Cy5) on the reliability of the RNAFISH results obtained both with a universal and a specie-specific probe.