Haynes Lab:Notebook/Engineering PC-TFs/2013/10/22: Difference between revisions
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# '''Pre-warm all liquid reagents to 37°C in the bead bath'''. | # '''Pre-warm all liquid reagents to 37°C in the bead bath'''. | ||
# After reagents are warmed, spray bottles down with 70% ethanol and prepare the biosafety hood as for routine work. Do all work in the biosafety hood. | # After reagents are warmed, spray bottles down with 70% ethanol and prepare the biosafety hood as for routine work. Do all work in the biosafety hood. | ||
# Retrieve the cell culture from the incubator. Stand the flask up | # Retrieve the cell culture from the incubator. Stand the flask up. Open the cap and aspirate old culture media from the bottom of the flask. | ||
# Add '''5 mL 1x PBS''' to the flask. Lay the flask flat (see image above) and wash by gently tilting the flask back and forth. | # Add '''5 mL 1x PBS''' to the flask. Lay the flask flat (see image above) and wash by gently tilting the flask back and forth. | ||
# Stand the flask up. Open the cap and aspirate off the PBS. | # Stand the flask up. Open the cap and aspirate off the PBS. | ||
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# Place the new flask into the 37°C incubator. The cells should adhere to the growth surface in a couple of hours. | # Place the new flask into the 37°C incubator. The cells should adhere to the growth surface in a couple of hours. | ||
# Discard the left-over cells as liquid waste by aspirating them into the vacuum trap. Discard all plastics, etc. that have come into contact with growth medium and cells as biohazard waste. | # Discard the left-over cells as liquid waste by aspirating them into the vacuum trap. Discard all plastics, etc. that have come into contact with growth medium and cells as biohazard waste. | ||
*Seed cultures in 6-well plates at ~2.5x105 so that cells will be ~5.0x105 at the time of transfection. | *Seed cultures in 6-well plates at ~2.5x105 so that cells will be ~5.0x105 at the time of transfection. |
Revision as of 09:30, 25 October 2013
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October 22, 2013Materials
Procedure
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