Related Experiment Videos
High-resolution cytometry of FISH dots in interphase cell nuclei
M Kozubek1, S Kozubek, E Lukásová
1Faculty of Informatics, Masaryk University, Brno, Czech Republic. kozubek@fi.muni.cz
Cytometry
|July 15, 1999
Summary
We developed a high-resolution cytometer (HRCM) that analyzes cells with high accuracy, bridging the gap between low-resolution flow cytometry and high-resolution confocal microscopy for cell research.
Area of Science:
- Cell biology
- Microscopy
- Cytometry
Background:
- Flow cytometry (FCM) and laser scanning cytometry (LSCM) offer low-resolution analysis of many cells.
- Confocal microscopy provides high-resolution analysis of fewer cells.
- A need exists for a system combining high throughput with high resolution.
Purpose of the Study:
- To present a novel high-resolution cytometer (HRCM) as a compromise between FCM/LSCM and confocal microscopy.
- To enable automated, high-resolution analysis of FISH-stained interphase nuclei.
Main Methods:
- Developed a fully automated, high-resolution cytometer (HRCM).
- Utilized a motorized epi-fluorescence microscope and CCD camera with computer control for acquisition and analysis.
- Employed sequential dye acquisition with specific filters and off-line computation of nuclear and dot attributes.
Main Results:
- HRCM analyzes multi-color and repeatedly hybridized preparations.
- Achieved acquisition and analysis speeds of approximately 50 nuclei/min (2D) and 1 nucleus/min (3D).
- Demonstrated lateral and axial measurement precision of approximately 100 nm.
Conclusions:
- HRCM provides quantities comparable to FCM/LSCM with accuracy similar to confocal microscopy.
- Suitable for routine diagnostics, therapy follow-up, chromatin structure studies, and cell research.
- Enables high-accuracy cell analysis bridging existing technological gaps.