Imaging technique implemented in CellTracks system.
BACKGROUND
We developed the CellTracks cell analysis system that, similar to flow cytometry, yields multiparameter information by which the cells can be differentiated. We describe the implementation of a laser scanning imaging method in the system. Image analysis of the cells improves the specificity of cell classification, especially in cases where the particular cells are found relatively infrequently and one has to discriminate between artifacts and real events.


METHODS
Fluorescent images of immunomagnetically labeled and aligned cells are obtained by passing the cells through a laser focus. The laser focus is smaller than the objects and subsequent frames captured by a regular surveillance CCD camera with a frame grabber board represent different parts of the cells. Complete images of the cells are constructed by shifting each image with respect to each other and adding individual pixel values.


RESULTS
The power of combining a fluorescent image with multiparametric data is demonstrated by imaging fluorescent and magnetically labeled beads and cells. The image gives additional information about the dye distribution across the objects. Changes in dye distribution as a function of time were observed in leukocytes labeled with the red fluorescent label, Oxazine750, which are imaged at different time intervals.


CONCLUSIONS
An imaging technique implemented in the CellTracks system provides high-resolution fluorescent images of events previously identified by the system. The images of the fluorescent cells enhance the ability to classify rare events.
Tibbe
Arjan G J
AG
Faculty of Applied Physics, University of Twente, Enschede, The Netherlands. A.G.J.Tibbe@tn.utwente.nl
de Grooth
Bart G
BG
Greve
Jan
J
Dolan
Gerald J
GJ
Terstappen
Leon W M M
LW
D001773:Blood Cells / Q000166:cytology*; D016250:Compact Disks / Q000639:trends; D005434:Flow Cytometry / Q000295:instrumentation / Q000379:methods*; D005456:Fluorescent Dyes; D006801:Humans; D007091:Image Processing, Computer-Assisted / Q000295:instrumentation / Q000379:methods*; D018189:Immunomagnetic Separation / Q000295:instrumentation / Q000379:methods*; D007834:Lasers; D007962:Leukocytes / Q000166:cytology; D008029:Lighting / Q000295:instrumentation / Q000379:methods; D008863:Microspheres; D010078:Oxazines; D013997:Time Factors
D005456:Fluorescent Dyes; D010078:Oxazines; C048170:oxazine 750