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RESEARCH DIVISION Laboratory Manual

 


 

Media and Solutions Required for Routine ES Cell Culture (Version 2)

(We have used both 1 & 2)

Media Used

To prepare 100 ml medium :

DMEM (GIBCO cat# 11965-062) 80 ml
15% FCS 15 ml
Pen/Strep 1 ml
L-Glutamine 1 ml
MEM non essential amino acids (GIBCO cat. # 11140-050) 1ml
Lif (105 U /ml) 1 ml
BME (0.1 M) 0.2 ml

LIF

comes in 1 ml ampules as LIF ESGRO AMRAD at 107U/ml dilute in 100 ml DMEM + 10% FCS and store in 5 ml aliquots at -20.

BME

Add 0.1ml of BME (14.4 M) to 14.3 ml PBS and filter through a 0.2 micron acrodisc. Store at -20 for up to 1 month.

PMEF Media

To prepare 100 ml of medium

DMEM (GIBCO cat# 11965-062) 88 ml
10% FCS 10 ml
Pen/Strep 1 ml
L-Glutamine 1 ml
BME (0.1 M) 0.2 ml
Disclaimer: The following set of protocols were contributed by various members of our lab (past and present): Christine Andrews, Fiona Christensen, Neil Della, Ross Dickins, Debbie Donald, Andrew Holloway, Gary Hime, Colin House, Yinling Hu, Rachael Parkinson, Nadia Traficante, Hannah Robertson, Ping Fu and Dennis Wang. Special thanks to Vicki Hammond, Frank Kontgen and Maria Murphy, who contributed many of the ES cell protocols. Sections dealing with Photomicroscopy, Polyclonal and Monoclonal Antibody Production were provided by members of Gerry Rubin's Laboratory (Berkeley). Any comments in the methods (technical errors etc.) E-mail: d.bowtell@pmci.unimelb.edu.au
David Bowtell PMCI October 1998