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Phase-Contrast Microscopy of Primary Chicken Embryo Fibroblast Cells

  • Authors: Kristina M. Obom 1, Patrick J. Cummings 2
  • VIEW AFFILIATIONS HIDE AFFILIATIONS
    Affiliations: 1: Johns Hopkins University, Baltimore and Rockville, Maryland, 21218; 2: Johns Hopkins University, Baltimore and Rockville, MD, 21218
  • Citation: Kristina M. Obom, Patrick J. Cummings. 2007. Phase-contrast microscopy of primary chicken embryo fibroblast cells.
  • Publication Date : September 2007
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Introduction



The figure shows a subconfluent (covering ~70% of plastic surface) monolayer (single layer of growing cells) of primary chicken embryo fibroblasts (a cell type that gives rise to connective tissue) under high magnification using phase-contrast microscopy (100X objective). Note the long spindle-shaped morphology of adherent fibroblasts and rounded refractile (dead) cells in the labeled view. 

Methods

Tissue from a 10-day-old embryonated chicken egg was dissected using aseptic technique, followed by tissue mincing, trypsinizing (addition of trypsin, a proteolytic enzyme that facilitates the breakdown of tissue into single cells), and seeding into T25 tissue culture flasks. The cells were grown at 37oC under 5% CO2 in Dulbecco's minimal essential medium containing 5% fetal calf serum (2).  

Discussion

Primary cells are harvested directly from an organism and can be grown for several weeks in vitro in specialized cell culture medium before the cells undergo senescence. Primary cells are especially sensitive to chemicals, toxins, and viruses including influenza virus (primary chick embryo kidney cells) (4) and Eastern equine encephalitis virus (primary chick embryo fibroblast cells) (6) and are often used for various research and industrial applications. Avian strains of influenza A virus replicate in chick embryo fibroblasts which is important for the study and isolation of these viruses. Investigators have used these cells to assay the infectivity of a strain of H5N2 in Texas (3) and test the antiviral properties of several antiviral compounds (1, 5).   

References

1. Conti, G., and P. Portincasa. 2002. Chromomycin A3 inhibits influenza A virus multiplication in chick embryo fibroblast cells.  New Microbiologica Official J. of the Italian Society for Medical, Odontoiatric, and Clinical Microbiology SIMMOC 25(4):385–398.  

2. Freshley, R. I. 2005. Culture of animal cells: a manual of basic technique, 5th ed.,  p. 177, 185, and 189.  John Wiley & Sons, Inc., New York, NY.  

3. Lee, C. -W., D. E. Swayne, J. A. Linares, D. A. Senne, D. L. Suarez. 2005. H5N2 avian influenza outbreak in Texas in 2004: the first highly pathogenic strain in the United States in 20 years? J. Virol. 79:11412–11421.

4. Ona, M., S. Melon, P. Iglesia, F. Hidalgo,  A. F. Verdugo. 1995. Isolation of influenza virus in human lung embryonated fibroblast cells (MRC-5) from clinical samples. Clin. Microbiol. 33:1948–1949.

5. Serkedjieva, J. 1995. Inhibition of influenza virus protein synthesis by a plant preparation from Geranium sanguineum L. Acta Virol. 39:5–10.

6. World Organization for Animal Health. 2004. Manual of diagnostic tests and vaccines for terrestrial animals.  Chap 2.5.3 Equine Encephalitis.  http://www.oie.int/eng/normes/mmanual/A_00081.htm.

 

 

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