With further development, this instrument could enable biodosimetry on a large scale.

A developmental instrument for assessment of radiation-induced damage in human lymphocytes includes an automated fluorescence microscope equipped with a one or more charge coupled device (CCD) video camera(s) and circuitry to digitize the video output. The microscope is also equipped with a three-axis translation stage that includes a rotation stage, and a rotary tray that holds as many as thirty specimen slides. The figure depicts one version of the instrument. Once the slides have been prepared and loaded into the tray, the instrument can operate unattended. A computer controls the operation of the stage, tray, and microscope, and processes the digital fluorescence image data to recognize and count chromosomes that have been broken, presumably by radiation.

The Original Version of the Automated Fluorescence Microscope included four CCD cameras and dichroic beam splitters for acquiring images in four fluorescence wavelength bands. The instrument was then modified to incorporate a single, more sensitive video camera and a filter wheel for taking a sequential exposures of each microscope field in the fluorescence wavelength bands of interest.

Under control by the computer, the microscope is automatically focused on the cells and each slide is scanned while the computer analyzes the DAPI-fluorescence images to find the metaphases. Each metaphase field is recentered in the field of view and refocused. Then a four-color image (more precisely, a set of images of the same view in the fluorescent colors of the four dyes) is acquired. By use of pattern-recognition software developed specifically for this instrument, the images in the various colors are processed to recognize the metaphases and count the chromosome fragments of each color within the metaphases. The intermediate results are then further processed to estimate the proportion of cells that have suffered genetic damage.

The prototype instrument scans at an average areal rate of 4.7 mm2/h in unattended operation, finding about 14 metaphases per hour. The false-alarm rate is typically less than 3 percent, and the metaphase-miss rate has been estimated to be less than 5 percent. The counts of chromosomes and fragments thereof are 50 to 70 percent accurate.

This work was done by Kenneth R. Castleman and Mark Schulze of Perceptive Scientific Instruments, Inc., for Johnson Space Center. For further information, contact the Johnson Technology Transfer Office at (281) 483-3809. MSC-23072