Automated cell cycle analysis with fluorescence microscopy and image analysis

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Abstract
Automatic cell cycle analysis (DNA histograms) is usually performed with flow cytometry. In cases when only few cells are available, the DNA content has to be measured with a fluorescence microscope combined with sensitive camera systems (SIT, MCP, cooled CCDs) and a frame grabber for image analysis. The fluorescent cells are observed on the monitor of the image analyser. The DNA content of specific cells of interest is calculated after an interactive selection via mouse click on the monitor screen. It is desirable to automate this time-consuming procedure by a computer "cell-finding' and "cell-measurement' system, which is comparable in speed and measurement accuracy with manual scoring. A software program, based on image analysis, was designed for the automated cell finding, the cell measurement, and finally the interactive DNA data evaluation. The system has already been used to determine cell cycle stages in which specific manipulations of a lymphocyte culture were required. However, the system is also able to determine, automatically, DNA distributions of different kinds of cell (different leucocytes) within one sample, as long as the automatic recognition software can distinguish between them by morphological differences.