Abstract: Reliable autofocus is required to obtain accurate measurements of fluorescent stained cellular components from a system capable of scanning multiple microscope fields. Autofocus could be performed directly with fluorescence images, but due to photobleaching and destructive fluorescence by-products, it is best to minimize fluorescence exposure for photosensitive specimens and live cells. This exposure problem could be completely avoided by using phase-contrast microscopy, implemented through the same optics as fluorescence microscopy. Functions for both phase-contrast and fluorescence autofocus were evaluated using the present invention and the suitability of phase-contrast autofocus for fluorescence microscopy was determined. The present autofocus system for scanning microscopy can be performed at least as fast as 0.25 s/field without loss of precision.

Abstract: An operator independent image cytometer having a method for image segmentation. Image segmentation comprises the steps of filtering a digital image of a cellular specimen and thresholding the resultant image. In addition, the thresholding may include the sorting of features extracted from the filtered image. The present invention also includes a method for cytometer autofocus that combines the benefits of sharpening and contrast metrics. The present invention further includes an arc lamp stabilization and intensity control system. The image cytometer has broad applications in determining DNA content and other cellular measurements on as many as 10.sup.5 individual cells, including specimens of living cells. Image segmentation applications include PAP smear analysis and particle recognition.

Abstract: Reliable autofocus is required to obtain accurate measurements of fluorescent stained cellular components from a system capable of scanning multiple microscope fields. Autofocus could be performed directly with fluorescence images, but due to photobleaching and destructive fluorescence by-products, it is best to minimize fluorescence exposure for photosensitive specimens and live cells. This exposure problem could be completely avoided by using phase-contrast microscopy, implemented through the same optics as fluorescence microscopy. Functions for both phase-contrast and fluorescence autofocus were evaluated using the present invention and the suitability of phase-contrast autofocus for fluorescence microscopy was determined. The present autofocus system for scanning microscopy can be performed at least as fast as 0.25 s/field without loss of precision.

Abstract: In an image segmentation system that processes image objects by digital filtration, a digital filter is defined. The digital filter includes a neighborhood operator for processing intensity values of neighborhoods of pixels in a pixel array. A first pixel array is received defining a pixelated image including one or more objects and a background and a second pixel array is received that defines a reference image. The reference image includes at least one object included in the pixelated image in a background. In the reference image, pixels included in the at least one object are distinguished from pixels included in the background by a predetermined amount of contrast. Pixels of the first and second images are compared to determine a merit value; the merit value is used to compute neighborhood operator values; and, the neighborhood operator is applied to images in order to create or enhance contrast between objects and background in the images.