Heat Activation: What is it and should I be doing it? How is it done?
Picture this: It’s 1976. The US of A is celebrating her Bicentennial and has the spiffy quarters to show for it. Gas was $0.59 per gallon, Rocky was killing it at the box office, and you just got your hands on some righteous new music. You pop in your new 8-track and start grooving to the awesome music that defined your generation. Just as you start to realize what the lyrics to “Afternoon Delight” mean, your PI calls you on your Snoopy phone and tells you that you have to start your experiments all over again. “Hey student. You just got yourself 3 more years in grad school thanks to these people.” Fast forward to today, and here’s a link to that fateful paper that made your stomach ache: http://www.springerlink.com/content/9466611g3p5n7224/
I imagine it as a figurative punch to the gut: Your stuff is invalid. “Your FBS is loaded with viruses and we have no way of knowing if your results are because of contamination, or experimentation. Start over.” For the rest of your life, you get inexplicably irritable when you hear the Starland Vocal Band.
Ok, a bit dramatic. It does paint a nice picture though, doesn’t it? Back in the late 1960s and 1970s the realization that contaminants in FBS are a real problem prompted scientists to try and figure out ways to clear up the issue. It wasn’t uncommon to find published reports of FBS lots that were tested with lists for what came up positive. It was clear that something needed to be done. Hepa filters weren’t an option. However, heat inactivation of serum was, and was used. Essentially, one would heat the FBS in a water bath to a temperature that would denature the proteins and genetic material in the contaminants. The magic recipe: 56C for 30 minutes. The side effects: Denaturation of the very growth factors and other yummy bits that made FBS useful in your cell cultures.
Now that gas is over 4 DOLLARS A GALLON!…………I mean, now that we have had some technological advancements since the 70s, FBS has become a lot cleaner. As a matter of fact, I am willing to bet that 99.9% of all contamination in cell cultures, comes from us: The researchers growing the cells with their grubby little hands, under- or unwashed lab coats, non-sterile latex gloves, runny noses, and particle catching arm hairs (I have a great story about my wife’s PhD advisor shaving himself, but I’ll save that for another blog…once I am assured he doesn’t know who actually writes these) and immersion in the bacteria factories we call water baths. I know, we try, and we have our techniques and ethanol spray bottles but inevitably our stuff gets dirty. It’s a dirty fact of life and our methods are vastly out-numbered by the opportunities our tissue culture facilities provide, either in the room , or by those who enter them.
Aseptically harvested and triple filtered, FBS and other sera now pose little to no threat of contamination. Gemini sometimes filters 4 times, as an added protective measure. It is overkill? Probably. But why risk contamination? Time is just as valuable as your cells, right? This “new way” of acquiring clean serum has made it possible to have clean cultures without damaging the trophic agents in the serum. Pretty cool no? However, heat inactivated serum is still widely used in labs all over the world. Why? I don’t really know. What I do know is this: Heat inactivate your serum if
1-You are in an immunology lab and you are worried about complement, which will confound your experiments, as FBS is loaded with various complement.
2- You are studying a particular growth factor that you know is present in your serum and you want to inactivate it.
3-You have been doing it for so long that you risk changing the cell culture environment you’ve made and the change would affect your research.
With that last one, there is no quick way to make the switch but,
as Gemini’s super scientific blogger-type guy and unsolicited
advice-giver, I’d be happy to help if you’re interested in
learning a bit more about heat inactivation and switching to
non-heat inactivated serum. Just let the good folks at Gemini
know and they will shoot me an email and we’ll talk.
Sky rocket in flight………………………………….