Etchevers:Notebook/Genomics of hNCC/2009/03/27: Difference between revisions

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Note that [http://www.openwetware.org/wiki/Phototoxicity some others have experienced CMFDA toxicity] although that was on exposure to light and creating free radicals (vitamin E seemed to protect against it).
Note that [http://www.openwetware.org/wiki/Phototoxicity some others have experienced CMFDA toxicity] although that was on exposure to light and creating free radicals (vitamin E seemed to protect against it).
CMFDA manual [http://probes.invitrogen.com/media/pis/mp02925.pdf here].


Review of previous trial:
Review of previous trial:

Revision as of 23:07, 26 March 2009

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Animal injections

Fed cells yesterday, cleaned up debris. Lots of cells in R1113, but they look dominated by large, fiber-containing cells with serrated edges. The rabbit corneal endothelial cells, in passing, are not proliferating - they have spread, but are too far apart on the plastic surface.

Note that some others have experienced CMFDA toxicity although that was on exposure to light and creating free radicals (vitamin E seemed to protect against it).

CMFDA manual here.

Review of previous trial:

  • Warm CMFDA to room temperature, dilute in 10.76 μL DMSO, which is 10 mM.
  • Dilute in Stemline medium *with no serum* to 10.76 mL. Rinse one flask with same medium, replace with CMFDA, incubate for 30 minutes.
  • Aspirate and replace with Rich medium. I think you have to let the cells sit for 45 minutes.
  • Trypsinize and resuspend in 2 mL tube with 0.2 mL Rich medium for 50 μL injections
  • FIND YOUR HAMILTON SYRINGE!

Some use 10 μM final concentration; the 25 μM last time was too much and I was thinking of using 5 μM (2000x dilution). But 10 is easier (1/1000).

  • Heather 02:05, 27 March 2009 (EDT):


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