A rapid, semiautomated system to quantitate and analyze leukocyte shape and locomotion was developed. Video images of moving leukocytes were obtained using a Vidicon camera mounted on a Nikon phase microscope. The video signal was either inputted directly, or indirectly via a video cassette recorder, to a Datacube video analog-digital, digital-analog converter. A Digital Equipment Corporation LSI 11/23 computer using the RT-II/ TSX-Plus operating system and computer programs written in FORTRAN and MARCO assembly language permitted image segmentation, image display, and calculation of position, speed, direction of movement and orientation of each leukocyte at 10 s intervals. These data were stored on a winchester disk for subsequent evaluation of the leukocyte orientation, speed and direction of movement using statistical and graphical methods. The reproducibility of measurements made with the video system was tested by comparison with manual measurements; a correlation coefficient of 0.998 was obtained for the two methods. Rates of chemokinesis were then determined for unstimulated and chemokinetically stimulated polymorphonuclear leukocytes (PMNs) and found to average 12.8 gm/min and 18.1 Ixm/ min, respectively. The high speed, ease of data analysis, and potential for multiparameter evaluation makes this system useful for directly evaluating leukocyte locomotion. chambers) do not provide precise data, measure a biased sample of the fastest cells and only measure distance moved in one direction rather than total chemotaxis and chemokinesis (Wilkinson et al. 1982).
Direct visual methods using time lapse microcinematography (Gruler and Bultmann 1984;Maher et al. 1984) or video techniques (Cheung and Miller 1984;Cheung et al. 1982;Wilkinson 1982) permit continuous observation of leukocyte locomotion thereby allowing precise and unbiased characterization of chemotaxis and chemokinesis. Unfortunately, these highly accurate means of characterizing leukocyte locomotion are limited by the need to use tedious manual measurement and analysis. For example, if the positions of 50 leukocytes are measured every 10 s for 30 min, a total of 9000 measurements must be made by hand using a ruler and recording the data on paper for subsequent analysis.
This paper describes a computer-assisted system for the quantitation of leukocyte locomotion that is much faster, less tedious and more objective than manual methods. This system automatically tracks up to 50 leukocytes at 10-s intervals, calculates the orientation, speed and direction of each leukocyte and records the data on a computer disk. Data obtained in this manner permit an in depth and unbiased analysis of leukocyte locomotion.