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Widefield deconvolution epifluorescence microscopy combined with fluorescence in situ hybridization reveals the spatial arrangement of bacteria in sponge tissue.
Citation key Manz2000
Author Manz, W. and G. Arp and G. Schumann-Kindel and U. Szewzyk and J. Reitner
Pages 125–134
Year 2000
DOI 10.1007/s002489900148
Journal Journal of Microbiological Methods
Volume 40
Number 2
Month Apr
Institution Technische Universität Berlin, Institut für Technischen Umweltschutz, Fachgebiet Okologie der Mikroorganismen, Sekretariat OE 5, Franklinstrasse 29, D-10587, Berlin, Germany. manz0654@mailszrz.tu-berlin.de
Abstract Widefield deconvolution epifluorescence microscopy (WDEM) combined with fluorescence in situ hybridization (FISH) was performed to identify and characterize single bacterial cells within sections of the mediterranean sponge Chondrosia reniformis. Sponges were embedded in paraffin wax or plastic prior to the preparation of thin sections, in situ hybridization and microscopy. Serial digital images generated by widefield epifluorescence microscopy were visualized using an exhaustive photon reassignment deconvolution algorithm and three-dimensional rendering software. Computer processing of series of images taken at different focal planes with the deconvolution technique provided deblurred three-dimensional images with high optical resolution on a submicron scale. Results from the deconvolution enhanced widefield microscopy were compared with conventional epifluorescent microscopical images. By the application of the deconvolution algorithm on digital image data obtained with widefield epifluorescence microscopy after FISH, the occurrence and spatial arrangement of Desulfovibrionaceae closely associated with micropores of Chondrosia reniformis could be visualized.
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