Growth and Maintenance of Chick Embryo Fibroblasts (CEF)

Raquel Hernandez1, Dennis T. Brown1

1 North Carolina State University, Raleigh, North Carolina
Publication Name:  Current Protocols in Microbiology
Unit Number:  Appendix 4I
DOI:  10.1002/9780471729259.mca04is17
Online Posting Date:  May, 2010
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Abstract

Primary cultures of chick embryo fibroblasts (CEF) are widely used for the cultivation of viruses. Protocols for the growth and maintenance of CEF cells in the laboratory are provided. Curr. Protoc. Microbiol. 17:A.4I.1‐A.4I.8. © 2010 by John Wiley & Sons, Inc.

Keywords: chick embryo fibroblasts; primary cell lines; cell culture

     
 
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Table of Contents

  • Introduction
  • Basic Protocol 1: Preparation of Primary Chick Embryo Fibroblast (CEF) Culture
  • Basic Protocol 2: Freezing and Thawing of Chick Embryo Fibroblast (CEF) Cultures
  • Reagents and Solutions
  • Commentary
  • Literature Cited
     
 
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Materials

Basic Protocol 1: Preparation of Primary Chick Embryo Fibroblast (CEF) Culture

  Materials
  • Fertile chicken eggs, 10 days old (Spafas; http://www.criver.com/RM/spf/egg_chickens.html)
  • 70% (v/v) ethanol
  • 1× phosphate‐buffered saline (PBS) with calcium and magnesium (see recipe)
  • 0.25% trypsin (see recipe)
  • Versene solution (see recipe)
  • 1× complete EMEM (see recipe)
  • 1× PBS‐D (see recipe)
  • Strong light source (75 to 100 W)
  • Small sterile beaker
  • Sharp scissors and forceps, sterile
  • Petri dishes
  • 50‐ml conical tubes
  • 37°C water bath
  • 5‐ or 10‐ml sterile pipets
  • 25‐cm2 or 75‐cm2 tissue culture flasks
  • Additional reagents and equipment for counting cells (Strober, )

Basic Protocol 2: Freezing and Thawing of Chick Embryo Fibroblast (CEF) Cultures

  Materials
  • CEF in culture (see protocol 1)
  • 0.25% trypsin (see recipe)
  • Versene solution (see recipe)
  • PBS‐D (see recipe)
  • 1× complete EMEM medium (see recipe), 37°C
  • Freezing medium (see recipe) made with EMEM
  • 2‐ml cryotubes
  • Nalgene Cryo 1°C freezing container
  • Liquid nitrogen freezer
  • 37°C water bath
  • 25‐cm2 flasks
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Figures

Videos

Literature Cited

Literature Cited
   Strober, W. 1997. Monitoring cell growth. Curr. Protoc. Immunol. 21:A.3A.1‐A.3A.2.
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