Abstract
Increasing interest has been paid to applications of fluorescence measurements to analyze physiological mechanisms in living cells. However, few studies have taken advantage of DNA quantification by fluorometry for dynamic assessment of chromatin organization as well as cell motion during the cell cycle. This approach involves both optimal conditions for DNA staining and cell tracking methods. In this context, this report describes a stoichiometric method for nuclear DNA specific staining, using the bisbenzimidazole dye Hoechst 33342 associated with verapamil, a calcium membrane channel blocker. This method makes it possible to correlate variations of nuclear DNA content with cell motion in cells that are maintained alive. Motion measurement is the second goal of this paper and it explains the snake-spline method, and the associated cell following method.
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Berger, M.O. (1991). Les contours actifs: modélisation, comportement et convergence, INPL thesis.
Berthommier, F., O. François, T. Coll, T. Hervé, I. Marque, P. Cinquin and J. Demongeot (1971). Asymptotic behaviour of neural networks and image processing. In: A. Babloyantz, ed., Self-Organization, Emerging Properties and Learning, 219–230. New York, Plenum Press, NATO Series.
Bertran de Rio, L., A. Gomez and M. Jose Yacaman (1991). Image processing in TEM using the wavelet transform. Ultramicroscopy 38: 319–324.
Blankenship, T. (1987). Real-time enhancement of medical ultrasound images. In: L.W. Kessler, ed., Acoustical Imaging, 187–195. New York, Plenum.
Canny, J. (1986). A computational approach to edge detection. IEEE PAMI 8: 679–698.
Catte, F., P.L. Lions, J.M. Morel and T. Coll (1992). Image selective smoothing and edge detection by nonlinear diffusion. SIAM J. Numer. Anal. 29: 182–193.
Cohen, I. (1992). Modèles déformables 2D et 3D: application à la segmentation d'images médicales. Paris-Dauphine thesis.
Colomb, E., C. Dussert and P.M. Martin (1991). Nuclear texture parameters as discriminant factors in cell cycle and drug sensitivity studies. Cytometry 12: 15–25.
Cottet, G.H. (1991). Modèles de réaction-diffusion pour des réseaux de neurones stochastiques et déterministes. C. R. Acad. Sci Serie I Math. 312: 217–221.
Cottet, G.H. and L. Germain (1993). Image processing through reaction combined with non-linear diffuson. Mat. Comput. 61: 659–673.
Cottet, G.H. (in press). Neural networks: continuous approach and applications to image processing. J. Biol. Systems.
de Boor, C. (1978). A practical guide to splines. Berlin, Springer Verlag (ed.)
Duncan, J.H. and T.C. Chou (1992). On the detection of Motion and the Computation of Optical Flow. IEEE PAMI 14: 346–352.
Erba E., P. Ubezio, M. Broggini, M. Ponti and M. D'Incalci M. (1988). DNA damage, cytotoxic effect and cell-cycle perturbation of Hoechst 33342 on L1210 cells in vitro. Cytometry 9: 1–6.
Fay, D.A. and A.M. Waxman (1991). Real-time early vision neurocomputing. In: International Joint Conference on Neural Networks, IEEE 1: 621–626.
Fischer, J.R. and J. Dorband (1991). Applications of the MasPar MP-1 at NASA/Goddard. In: Compcon 91, IEEE: 278–282
Garcia-Segura, L.M., F. Soto, R. Planells-Cases, J. M. Gonzales-Ros and J.A. Ferragut (1992). Verapamil reverses the ultrastructural alterations in the plasma membrane induced by drug resistance. FEBS Lett. 314: 404–408.
Horn, B.K.P. and B.G. Schunk (1981). Determining optical flow. Artificial Intell. 17: 185–203.
Khellaf, A., A. Beghadi and H. Dupoisot (1991). Entropic contrast enhancement. IEE Trans. Med. Imaging 10: 589–592.
Kwon, T.M. and M.E. Zervakis (1992). Design of regularization filters with linear neural networks (image restoration). In: IEEE Trans Syst., Man Cybern. 1: 416–421.
Kobayashi, H., T. Matsumoto, T. Yagi, and T. Shimmi (1991). A layered architecture for regularization vision chips. In: IEEE International Joint Conference on Neural Networks, IEEE, 2: 1007–1020.
Kobayashi, H., T. Matsumoto, T. Yagi and T. Shimmi (1993). Image processing regularization filters on layered architecture. Neural Networks 6: 327–350.
Kobayashi, N., H. Saito and M. Nakajima (1994). Fast adaptive contrast enhancement method for the display of gray-tone images. Trans. IEICE 77: 502–509.
Krishan, A. (1987). Effect of drug efflux blockers on vital staining of cellular DNA with Hoechst 33342. Cytometry 8: 642–645.
Lalande, M.E., V. Ling and R.G. Miller (1981). Hoechst 33342 dye uptake as a probe of membrane permeability changes in mammalian cells. Proc. Natl. Acad. Sci., USA 78: 363–367.
Latt, S.A. and R.G. Langlois (1990). Fluorescent probes of DNA microstructure and DNA synthesis. In: M.R. Melamed, T. Lindmo and M.L. Mandelsohn, eds., Flow Cytometry and Sorting, 249–290. New York, Wiley-Liss, Inc.
Laurent, P.J. (1972). Approximation et Optimisation. Paris, Hermann (ed.)
Le Negrate, A., A. Beghadi and H. Dupoisot (1992). An image enhancement technique and its evaluation through bimodality analysis. CGVIP: Graph. Models Image Process 54: 13–22.
Leitner, F. (1993). Segmentation dynamique d'images tridimensionnelles. INPG thesis.
Leitner, F., I. Marque, F. Berthommier, T. Coll, O. François, P. Cinquin and J. Demongeot (1991). Neural networks, differential systems and image processing. In: J.C. Simon, ed., From Pixels to Features II, 253–274. North Holland.
Martin, J.P. (1988). Fast enhancement technique for PC-based image processing systems. In: Electronic Imaging 88, Diagn. Imag. Mag. 1: 194–198.
Nakahira, H., M. Maruyama, H. Ueda and H. Yamada (1993). An image processing system using image signal multiprocessors (ISMPs). J. VLSI Signal Process. 5: 133–140.
Neycenssac, F. (1993). Contrast enhancement using the Laplacian of a Gaussian filter. CVGIP: Graph. Models Image Process. 55: 447–463.
Ning, F. and C. Ming Chieh (1993). An automatic cross-over point selection technique for image enhancement using fuzzy sets. Pattern Recognition Lett. 14: 397–406.
Paillasson, S., G. Brugal and X. Ronot (1994). Marquage du noyau de cellules vivantes par le Hoechst 33342: un nouveau concept pour l'analyse dynamique du contenu en ADN. Innov. Technol. Biol. Med. 15: 583–594.
Paillasson, S., F. Germain, P. Stelmaszyk and X. Ronot (1994). Dynamic analysis of cell motion: a new method based on dense vector field estimation. Anal. Cell. Pathol. 6: 201.
Paillasson, S., J.M. Millot, M. Manfait and X. Ronot (1995). DNA analysis in living cells: cytometric approaches. In: G. Morel, ed., Visualization of Nucleic Acids. New York, CRC Press.
Ritter, C. and A. Tanner-Martin (1992). Facilitating the Gibbs sampler: the Gibbs stopper and the Griddy-Gibbs sampler. J. Amer. Statist. Assoc. 87: 861–868.
Rougon, N. (1993). Eléments pour la reconnaissance de formes tridimensionnelles déformables. Application à l'imagerie biomédicale. ENST thesis.
Sablonnière, P. (1978). Spline and Bézier polygons associated with a polynomial spline curve. Computer Aided Design 10: 257–261.
Sablonnière, P. and A. le Méhauté (1987). Courbes et surfaces. Bézier/B-splines, A.T.P. du CNRS.
Santisteban-Otegui, M.P. Montmasson, F. Giroud, X. Ronot and G. Brugal (1992). Fluorescence image cytometry of nuclear DNA content versus chromatin pattern: a comparative study of ten fluorochromes. J. Histochem; Cytochem. 40: 1789–1797.
Shumaker, L.L. (1981). Spline Functions: Basic Theory. John Wiley & sons.
Taylor, D.L. and Y.L. Wang (1980). Fluorescent labelled molecules as probes of the structure and function of living cells. Nature 284: 405–410.
Terzopoulos, D., M. Kass and A. Witkin (1987). Snakes: active contour models. Proc of ICCV: 259–268.
Toet, A. (1992). Multiscale contrast enhancement with applications to image fusion. Opt. Eng. 31: 1026–1031.
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Leitner, F., Paillasson, S., Ronot, X. et al. Dynamic functional and structural analysis of living cells: New tools for vital staining of nuclear DNA and for characterisation of cell motion. Acta Biotheor 43, 299–317 (1995). https://doi.org/10.1007/BF00713555
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DOI: https://doi.org/10.1007/BF00713555