Six-colour fluorescent imaging of lymphoid tissue based on colour addition theory

B. Ma; S. Winkelbach; W. Lindenmaier; K.E.J. Dittmar

doi:10.1016/j.acthis.2006.02.002

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Six-colour fluorescent imaging of lymphoid tissue based on colour addition theory

Journal article

Peer reviewed

Six-colour fluorescent imaging of lymphoid tissue based on colour addition theory

B. Ma, S. Winkelbach, W. Lindenmaier and K.E.J. Dittmar

Acta Histochemica, Vol.108(4), pp.243-257

2006

DOI: https://doi.org/10.1016/j.acthis.2006.02.002

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Abstract

Multi-colour imaging of immunofluorescently labelled tissue using confocal microscopy was accomplished by using colour addition theory. This new technique includes several improvements for immunolabelling: (1) the co-localization of two or more markers on one cell for the identification of specific cell populations; (2) the co-localization of two fluorescent dyes from secondary reagents for the identification of the cells; (3) a multi-step staining protocol with two primary antibodies originating from the same host species or with two or three biotin-conjugated primary antibodies. After image acquisition, colour segmentation/unmixing are applied to the single multi-colour image to generate multi-pseudo-channels for individual or co-localized fluorescent dyes. With this new technique, we have been able to visualize six cell populations simultaneously in the mouse lymph node and intestine. The efficiency of this method has also been demonstrated in the three-dimensional reconstruction of thick sections from mouse ileum. Our method is simple, efficient, and may be indispensable in experimental cell and tissue studies requiring multiple immunolabelling.

Details

Title: Six-colour fluorescent imaging of lymphoid tissue based on colour addition theory
Authors/Creators: B. Ma (Author/Creator)
S. Winkelbach (Author/Creator)
W. Lindenmaier (Author/Creator)
K.E.J. Dittmar (Author/Creator)
Publication Details: Acta Histochemica, Vol.108(4), pp.243-257
Publisher: Elsevier GmbH
Identifiers: 991005543456707891
Copyright: © 2006 Elsevier GmbH.
Murdoch Affiliation: Murdoch University
Language: English
Resource Type: Journal article

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Collaboration types: Domestic collaboration
Citation topics: 2 Chemistry; 2.259 Optical Chemistry; 2.259.815 Fluorescence Microscopy
Web Of Science research areas: Cell Biology
ESI research areas: Biology & Biochemistry