Automated image analyzing system for the quantitative study of living cells in culture

A fully automated image analyzing system was developed for the quantitative study of cells in culture. It was able to count cells, to classify cells according to their morphological characteristics and to follow cell culture development. A specific procedure was designed to process Hoffman modulation contrast images. It detects local gray level differences while using conditional dilation techniques. We were able to successfully detect aggregated unstained cells, presently a technical limit in image segmentation. Living cells can be studied in a noninvasive and nondestructive way with this system. An improved automatic focusing algorithm was developed which ensured an accurate prediction of the optimal focus position. A strictly defined sampling procedure was applied to estimate unbiasedly cell density and obtain precisely cell contours. The evaluation of the system was carried out on Chinese hamster ovary (CHO‐NTR) cell cultures treated with a newly developed neurotensin agonist JMV449. Chinese hamster ovary cell division was found to be retarded 20 hours after the JMV449 treatment, while the morphology of CHO‐NTR cells has already undergone significant changes 12 hours after the treatment. This image analyzing system provides the possibility to follow cell culture development (e.g., cell density evolution, cell morphological changes) under various experimental conditions. © 1994 Wiley‐Liss, Inc.

[1]  P. Kitabgi,et al.  Reduced peptide bond pseudopeptide analogues of neurotensin: binding and biological activities, and in vitro metabolic stability. , 1991, European journal of pharmacology.

[2]  D. Guidolin,et al.  Morphometric evaluation of populations of neuronal profiles (cell bodies, dendrites, and nerve terminals) in the central nervous system , 1992, Microscopy research and technique.

[3]  Jean Serra,et al.  Image Analysis and Mathematical Morphology , 1983 .

[4]  L. C. Kenny,et al.  Automated focusing of an optical microscope , 1983 .

[5]  T. W. Anderson.  An Introduction to Multivariate Statistical Analysis , 1959 .

[6]  R Di Paola,et al.  The use of an image analyser in human tumour clonogenic assays. , 1986, Cytometry.

[7]  H J Gundersen,et al.  The efficiency of systematic sampling in stereology and its prediction * , 1987, Journal of microscopy.

[8]  G Burnstock,et al.  Quantitative method for the study of the morphological changes induced by vasoactive agents in single aortic myocytes grown in primary cultures. , 1987, Laboratory investigation; a journal of technical methods and pathology.

[9]  Masayuki Masu,et al.  Structure and functional expression of the cloned rat neurotensin receptor , 1990, Neuron.

[10]  J. Chassery,et al.  An iterative region-growing process for cell image segmentation based on local color similarity and global shape criteria. , 1986, Analytical and quantitative cytology and histology.

[11]  P Bodin,et al.  Study of living single cells in culture: automated recognition of cell behavior. , 1988, Laboratory investigation; a journal of technical methods and pathology.

[12]  Robert Hoffman,et al.  The modulation contrast microscope: principles and performance , 1977 .

[13]  G Brugal,et al.  A quantitative analysis of the human bone marrow erythroblastic cell lineage using the SAMBA 200 cell image processor. I. The normal maturation sequence. , 1987, Analytical and quantitative cytology and histology.

[14]  Jean-Pierre Royet,et al.  Stereology: A method for analyzing images , 1991, Progress in Neurobiology.

[15]  K Zilles,et al.  Image analysis of Nissl‐stained neuronal perikarya in the primary visual cortex of the rat: Automatic detection and segmentation of neuronal profiles with nuclei and nucleoli * , 1990, Journal of microscopy.

[16]  H. Gundersen,et al.  Notes on the estimation of the numerical density of arbitrary profiles: the edge effect , 1977 .

[17]  L. Yan,et al.  An automated method for autoradiographic analysis of cultured Schwann cells , 1987, Journal of Neuroscience Methods.

[18]  J. D. Clements,et al.  Automated image analysis for counting unstained cultured neurones , 1991, Journal of Neuroscience Methods.

[19]  Azriel Rosenfeld,et al.  Digital Picture Processing , 1976 .