Filter-Dense Multicolor Microscopy

PloS One
Siavash KijaniPer Fogelstrand

Abstract

Immunofluorescence microscopy is a unique method to reveal the spatial location of proteins in tissues and cells. By combining antibodies that are labeled with different fluorochromes, the location of several proteins can simultaneously be visualized in one sample. However, because of the risk of bleed-through signals between fluorochromes, standard multicolor microscopy is restricted to a maximum of four fluorescence channels, including one for nuclei staining. This is not always enough to address common scientific questions. In particular, the use of a rapidly increasing number of marker proteins to classify functionally distinct cell populations and diseased tissues emphasizes the need for more complex multistainings. Hence, multicolor microscopy should ideally offer more channels to meet the current needs in biomedical science. Here we present an enhanced multi-fluorescence setup, which we call Filter-Dense Multicolor Microscopy (FDMM). FDMM is based on condensed filter sets that are more specific for each fluorochrome and allow a more economic use of the light spectrum. FDMM allows at least six independent fluorescence channels and can be applied to any standard fluorescence microscope without changing any operative proced...Continue Reading

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Citations

Jul 27, 2017·Mucosal Immunology·N Y Lycke, M Bemark
Dec 3, 2017·Science Immunology·Jayendra Kumar KrishnaswamyStephanie C Eisenbarth
Apr 19, 2015·Genome Biology·Felix SommerFredrik Bäckhed
Aug 25, 2017·Nature Protocols·Alexandre BaccoucheAnthony J Genot
Jun 6, 2019·Methods and Protocols·Sebastian C Bhakdi, Ponpan Thaicharoen
Aug 18, 2021·The American Journal of Pathology·Ankur PanditaMax Levin

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Methods Mentioned

BETA
flow cytometry
fluorescence microscopy
antibody array

Software Mentioned

Axiovision
FDMM
Semrock SearchLight

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