centrifuge at 4°C - (Dec/12/2007 )
I centrifuge at RT, unless I want to stop an activationn then I centrifuge at 4C and I lyse the cells after.
I keep media at 4c, not to prevent from contamination, because I assume there is no bacteria in my medium, unless they will start to grow anyway when I will incubate the medium at 37C with the cells.
I warm the medium when I use cells that could detach from the plastic when you put cold medium, unless I use cold medium, or slghtly warmed at RT
I assume following this logic that you use PBS, Culture media and Trypsin at +4 degrees????????
Doing routine passaging of cells, they should be at room temperature.
Use +4 degrees when preparing cell pellets for enzyme preparations etc.
i don't see how you make that assumption based on what i wrote regarding preservation of a sample.
i don't do cell culture so i am not familiar with the conditions required to passage the cells but the question asked was why 4C is used during centrifugation of the cells.
if lower temperature is not required when the cells are being harvested to be regrown then the assumption is that joohn is going to process the collected cells, in which case you would want to preserve the cell components.
if there is something wrong with my understanding, please don't hesitate to correct it.
I always centrifuge my cells at RT .
Dear Lauralee and Minnie mouse,
Keep up the good work you seem to be the only one doing TC properly.
As always kindest regards.
Actually we spin not at 4 but at 10 C. We were told that it doesn't make a huge difference for the cells, but it makes a big difference for the centrifuge because there always working to get it at such a low temp. So they should live longer, anything for our fuges! . Since we've changed we haven't noticed anything different in our cell culture exp. So I guess it works.
Generally you work at 4 C while carrying on experiments in which there are possibility of enzyme activity that is undesirable. For example in RNA isolation all centrifugation steps are done at 4 C, to minimize possible activity of endogenous RNase activity in order to obtain intact RNA.
If you spin down cells for 5 minutes in a 4°C centrifuge, how much colder will your cells have gotten, assuming they started at anywhere between 37°C (temp from your incubator) and RT (because you removed cells from incubator some minutes ago, so they will have lost a bit of the temp and maybe you have added some room temperture solution)? Not too much.
Centrifuging stuff at 4°C only makes sense when you start with cooled material, or if you will centrifuge for a long time at high speed to prevent sample from heating (e.g. precipitation of a maxiprep or so according to qiagen's protocol).
BTW: I'm with Lauralee and Minnie (and Rhombus) on the storing and pre-heating cell culture material, yay!
I just wondered why sometimes (or even always) it is recommanded to centrifuge cells at 4°C? Has anyone an idea? May be to shut down the cell metabolism?
in most cases, I think, it is not necessary to spin down cells at 4°C; however, multi-used centrigues are often pre-cooled at 4°C, so you may have no alternative...
At 4 degree celcius most of the biological substances are stable. However, centrifugation at room temperature is also use in some cases.