Scanning fluorescent microscopy is an alternative for quantitative fluorescent cell analysis

Cytometry. Part a : the Journal of the International Society for Analytical Cytology
Viktor VargaBela Molnar

Abstract

Fluorescent measurements on cells are performed today with FCM and laser scanning cytometry. The scientific community dealing with quantitative cell analysis would benefit from the development of a new digital multichannel and virtual microscopy based scanning fluorescent microscopy technology and from its evaluation on routine standardized fluorescent beads and clinical specimens. We applied a commercial motorized fluorescent microscope system. The scanning was done at 20 x (0.5 NA) magnification, on three channels (Rhodamine, FITC, Hoechst). The SFM (scanning fluorescent microscopy) software included the following features: scanning area, exposure time, and channel definition, autofocused scanning, densitometric and morphometric cellular feature determination, gating on scatterplots and frequency histograms, and preparation of galleries of the gated cells. For the calibration and standardization Immuno-Brite beads were used. With application of shading compensation, the CV of fluorescence of the beads decreased from 24.3% to 3.9%. Standard JPEG image compression until 1:150 resulted in no significant change. The change of focus influenced the CV significantly only after +/-5 microm error. SFM is a valuable method for the eval...Continue Reading

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Citations

Sep 13, 2011·Apoptosis : an International Journal on Programmed Cell Death·Leo Li-Ying ChanBo Lin
Jul 27, 2007·Lab on a Chip·Shannon E WeigumJohn T McDevitt
Feb 26, 2005·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Dominik LenzUlrich Sack
Jun 2, 2004·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Andreas O H GerstnerAttila Tárnok
Sep 8, 2004·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·József BocsiAttila Tárnok
Sep 25, 2008·Cytometry. Part B, Clinical Cytometry·Arkadiusz PierzchalskiAttila Tárnok
Feb 16, 2006·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·József BocsiAttila Tárnok
Jan 30, 2008·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Levente FicsorBela Molnar
Mar 17, 2006·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Attila TárnokFrank Emmrich
Jun 30, 2006·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Attila Tárnok
May 4, 2011·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Leo L ChanBo Lin
Feb 22, 2011·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Tycho M ScholtensLeon W M M Terstappen
Sep 12, 2009·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Viktor Sebestyén VargaBéla Molnár
Mar 31, 2016·Journal of Microscopy·Peet KaskThora Pommerencke
Mar 17, 2015·Nature Methods·Kevin SmithPeter Horvath
Jun 9, 2017·Nature Communications·Tingying PengNassir Navab
May 9, 2006·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Howard M Shapiro, Nancy G Perlmutter
Sep 16, 2006·Cytometry. Part B, Clinical Cytometry·Levente FicsorBela Molnar
Apr 15, 2005·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Anja MittagAttila Tárnok

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