Debbie Knight

A Day in the Life…April 14, 2011

In research log on April 14, 2011 at 5:34 pm

From time to time, I will give a glimpse into the “glamorous” life of a research associate and talk about what I’m doing in the lab on a particular day. These entries I will call “A Day in the Life…”

A  couple of weeks ago, we got a frozen vial of cells from the American Type Culture Collection (ATCC) – cells that were isolated from a colorectal cancer that we will use to model the lining of the intestines. Not a perfect model, the cells are, after all, cancer cells which means they’re different from the original cell type in several ways – they have mutations in their DNA, they may have more than the standard 46 chromosomes in the human cell, and they grow more quickly in culture than their original counterparts, but hopefully they will behave enough like the original cells that we can draw meaningful conclusions from our experiments.

So, in that frozen vial we got from the ATCC is perhaps a million cells – though it sounds like a lot, it’s not quite enough for our research needs. So, we have to place them in a small tissue culture flask, allow them to grow and crowd in, and then transfer them to more tissue culture flasks. We will repeat this a few times to further expand the culture.

Our cells look something like this when you look at them under the microscope. (This image taken from http://www.iclc.it/Listanuova.html)

Since this is a new cell line for us, I’ve been freezing a small portion of the culture at each transfer. We do this primarily in case our culture becomes contaminated with bacteria or fungus – not something we want to happen, but it can occur. If the culture does become contaminated, it’s nearly impossible to get rid of it so it’s best to discard the culture. By freezing some cells at each step, we ensure that we have a few cells saved for future experiments.

So, we store the frozen cells in a cryopreservation tank. Remember the scene from the movie “JurassicPark” when the guy who’s stealing the dinosaur embryos takes them out of a “steaming” container? Well, in the movies they probably used dry ice or a fog machine to achieve the “steaming” effect, but in the research lab, we use liquid nitrogen to keep the cells frozen. The “steam” actually looks similar to the “fog” produced by dry ice or the fog machine and a little like steam from boiling water.  The “steam” from liquid nitrogen is not warm at all, in fact it’s quite cold. Liquid nitrogen boils at minus 324 degrees Fahrenheit – brrr!

So, when you freeze the cells, you can’t just put the cells in the liquid nutrients (culture medium) you’ve been growing them in – if you did that, the ice crystals could pierce and shatter the fragile cells. To prevent the cells from rupturing during the freezing process, we add a commonly used chemical called dimethyl sulfoxide (DMSO) at a concentration of 10% just before we freeze them. There are more elaborate cryopreservatives that we could use, but DMSO is cheaper and works fine for our needs. We are able to thaw the cells on a later date, most of them will remain intact and grow.

So, now we have millions of cells stored in our lab making it possible to do many experiments in the future.

Cryopreserving cells is one of the things I did today in the lab – not the most exciting thing, but definitely an important task.

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  1. Hi Debbie, based on your experience, does the brand of cell culture plastic you use for culture effect how the cells behave or look?

  2. To be honest, I haven’t tried many of the tissue culture wares out there (the three labs I’ve worked in have pretty much “stuck” with Corning-Costar products), so I can’t really comment on cell behavior & plastics. I will say that they can display different adherence behavior on glass compared to plastic. Sorry I can’t directly address your question.

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