Team:Warwick/Notebook/cellmaintenance
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- | < | + | <body> |
+ | <div id = "pageWrapper"> | ||
+ | <!--CONTENT START--> | ||
+ | |||
+ | <img class = "headerImage" style = "width: 30%;" src="https://static.igem.org/mediawiki/2014/f/ff/RepliconLogoON.png"> | ||
+ | <img class = "headerImage" style = "width: 12%;" src="https://static.igem.org/mediawiki/2014/f/f6/Warwick_logo.png"> | ||
+ | <div id = "mainMenuBar"> | ||
+ | <li> <a href = "/Team:Warwick/Project"> PROJECT </a> </li> | ||
+ | <li> <a href = "/Team:Warwick/Team"> TEAM </a> </li> | ||
+ | <li> <a href = "/Team:Warwick/Parts"> PARTS </a> </li> | ||
+ | <li> <a href = "/Team:Warwick/Modelling"> MODELLING </a> </li> | ||
+ | <li> <a href = "/Team:Warwick/Notebook"> NOTEBOOK </a> </li> | ||
+ | <li> <a href = "/Team:Warwick/Human"> HUMAN PRACTICES </a> </li> | ||
+ | <li> <a href = "/Team:Warwick/Measurements"> MEASUREMENTS </a> </li> | ||
+ | <li> <a href = "/Team:Warwick/Interlab"> INTERLAB </a> </li> | ||
+ | <li> <a href = "/Team:Warwick/Attributions"> SPONSORS </a> </li> | ||
+ | </div> | ||
- | < | + | <div id = "pageContentWrapper"> |
- | </ | + | <div id = "pageContent"> |
+ | <!-- THIS IS WHERE YOUR MAIN BODY GOES --> | ||
+ | <h1> Human Cell Line Maintenance </h1> <br> <br> | ||
+ | <p> | ||
+ | Materials: 25ml DMEM media, 10ml PBS, 2ml Trypsin, (5ml Pen/strep, 50ml FBS) 1 tissue culture plate, 1 15ml falcon tube. | ||
+ | </p> | ||
- | < | + | <p> |
- | + | If using fresh DMEM, defrost pen/strep and 50ml FBS in 37⁰C waterbath (1 hour before starting). Then add 50ml FBS and 5ml pen/strep to DMEM media – label with name and date. | |
+ | Warm DMEM, PBS and Trypsin in 37⁰C waterbath at least 30min in advance of starting. | ||
+ | </p> | ||
+ | <ol> | ||
- | < | + | <li>Check cell confluency with microscope</li> |
+ | <li>Aspirate off old media (with vacuum pump</li> | ||
+ | <li>Wash cells with 10ml warm PBS and aspirate PBS off</li> | ||
+ | <li>Add 2ml Trypsin, move to incubator for 3-5min until cells begin to come off plate</li> | ||
+ | <li>Check cells have detached from plate (microscope)</li> | ||
+ | <li>Add 5ml DMEM media, wash plate thoroughly to re-suspend all cells, move to 15ml tube</li> | ||
+ | <li>Spin 4min at 1000rpm and room temp</li> | ||
+ | <li>Aspirate off media (do not disturb pellet)</li> | ||
+ | <li>Re-suspend cells in 1ml DMEM media, add a further 9ml DMEM. Mix thoroughly.</li> | ||
+ | <li>Plate 1ml culture and add 9ml DMEM (for a 1 in 10 dilution).</li> | ||
+ | <li>Swirl plate side to side and up and down (not in a circular motion)</li> | ||
+ | <li>Grow 2 days at 37⁰C and 5% CO2 </li> | ||
+ | </ol> | ||
+ | <p> Return DMEM and PBS to fridge, and pen/strep and trypsin to freezer. </p> | ||
+ | </div> | ||
+ | </div> | ||
+ | <!--CONTENT END--> | ||
+ | </div> | ||
+ | </body> | ||
- | </ | + | </html> |
Latest revision as of 10:21, 27 August 2014
Human Cell Line Maintenance
Materials: 25ml DMEM media, 10ml PBS, 2ml Trypsin, (5ml Pen/strep, 50ml FBS) 1 tissue culture plate, 1 15ml falcon tube.
If using fresh DMEM, defrost pen/strep and 50ml FBS in 37⁰C waterbath (1 hour before starting). Then add 50ml FBS and 5ml pen/strep to DMEM media – label with name and date. Warm DMEM, PBS and Trypsin in 37⁰C waterbath at least 30min in advance of starting.
- Check cell confluency with microscope
- Aspirate off old media (with vacuum pump
- Wash cells with 10ml warm PBS and aspirate PBS off
- Add 2ml Trypsin, move to incubator for 3-5min until cells begin to come off plate
- Check cells have detached from plate (microscope)
- Add 5ml DMEM media, wash plate thoroughly to re-suspend all cells, move to 15ml tube
- Spin 4min at 1000rpm and room temp
- Aspirate off media (do not disturb pellet)
- Re-suspend cells in 1ml DMEM media, add a further 9ml DMEM. Mix thoroughly.
- Plate 1ml culture and add 9ml DMEM (for a 1 in 10 dilution).
- Swirl plate side to side and up and down (not in a circular motion)
- Grow 2 days at 37⁰C and 5% CO2
Return DMEM and PBS to fridge, and pen/strep and trypsin to freezer.