Team:Heidelberg/Notebook/miMeasure/August

From 2010.igem.org

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===09/08/2010===
 
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seeding and transfecting cells for microscopy test measurements
 
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* add 0.6ul FuGENE reagent to 20ul of OptiMEM
 
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* mix and incubate 5' at RT
 
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* add 0.2ug DNA
 
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* mix and incubate 15' at RT
 
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* add DNA-FuGENE solution to 10 000 cells (400ul)
 
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* mix and incubate 10' 37degC 300rpm (shaker for 15ml falcon tubes is in the cell culture room at 3rd floor)
 
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* transfere cells to the plate
 
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* grow 48h
 
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===10/08/2010===
 
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coating plates with poly-L-lysine
 
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* add 15ul of poly-L-lysine solution to each well (make sure whole surface is covered with solution)
 
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* leave for 30' in the incubator
 
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* remove poly-L-lysine solution
 
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* wash once with PBS
 
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new 96-well plate was prepared (wells 2500 cells Huh7 are contaminated with HeLa) like previously
 
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===25/08/2010===
 
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seeding cells for pilot FACS and Tecan measurements (96-well plate)
 
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===26/08/2010===
 
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transfection - 96-well plate
 
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===27/08/2010===
 
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pilot FACS and Tecan measurements (96-well plate)
 
{{:Team:Heidelberg/Pagemiddle}}
{{:Team:Heidelberg/Pagemiddle}}

Revision as of 08:55, 26 October 2010

08/08/2010

seeding cells for test measurements - 96 well plate for plate-reader and FACS

test of media:

  • OptiMEM 0% FBS + Pen/Strep + L-Glu + other according to cell line
  • OptiMEM 2% FBS + Pen/Strep + L-Glu + other according to cell line
  • DMEM 10% FBS+ Pen/Strep + L-Glu + other according to cell line


  • cells did not adhere properly so we decided to coat 96-well plate with poly-L-lysine and repeat
  • significantly better growth in DMEM with 10% FBS, so we decided to grow cells in it and before measurement to change it to OptiMEM, to reduce high background fluorescence coming from FBS.


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