I'm kind of afraid Sarah will sneak into my room at night and shave my eyebrows off if I don't post...so here it goes.
I (along with most of the other contributors to this wonderful blog) have recently successfully completed the Hughes Program. After working in our labs every day, all day for ten weeks we're finally done with our projects. Or at least that was the idea. I am nowhere near done with my project.
Swastik, my lab's awesome grad student (I, too, am not a part of the MacQueen lab), told me the other day that when Thomas Edison finally succeeded in making the light bulb after his 30th attempt (or something like that), someone asked him if he felt like he was wasting his time for all those years he spent unsuccessfully making the light bulb. Apparently, the wise Mr. Edison replied that he did not, in fact, feel as though he had wasted his time because he now knew of 29 ways not to make a light bulb.
Well, I've spent the last 10 weeks growing insect cells, and I've learned at least 5 ways not to grow insect cells. I'm using insect cells as a way to express a eukaryotic protein and make a lot of it in hopes of one day crystallizing it and finding its structure. The cells I'm using are ovarian cells from this caterpillar, called the cabbage loper (Trichoplusia ni):
Because they're just the ovarian cells, they don't have any kind of immune system, so they have to be cultured with sterile technique. This means I have to work with them inside a sterile hood, spray everything that goes into the hood (including my hands and arms) with ethanol and generally make sure I don't contaminate them. If they die, then I have to start over from a frozen stock.
In order to make them express my protein, I infect them with a virus called baculovirus. In the wild, I'm told this virus causes the caterpillars to burst and drip down onto the caterpillars on tree branches below them, infecting these caterpillars as well. In the lab, we use a virus that has the instructions for making our protein in it, so before it kills the cells, it causes them to make a bunch of protein.
Anyway, I'm the first student in my lab (the Olson lab) to use this expression system, as we, like the MacQueen lab, just started last fall. Therefore, it's been kind of an adventure for me to figure out what to do and not to do, and while I did manage to express enough protein to squeeze out some data this summer, I also discovered at least five ways NOT to grow insect cells. Because I did not have a chance to share this accomplishment on my Hughes poster, I will take the opportunity to do so here.
Five Ways Not to Grow Insect Cells by Falcon Tube:
Don't grow insect cells by...
1) ...Letting them get too hot. This is tricky if you're working somewhere where they keep turing off the AC when it gets above 95 outside (as they do at Wesleyan).
2) ...Being too obsessed and look at them every day twice a day, as you'll shake them around too much and they'll die, or at least be unhappy and grow slowly.
3) ...Growing them without antibiotics. They can get infected with bacteria and die. (Some people do actually grow them without antibiotics, but it didn't work for me).
4) ...Letting them get infected with fungus. They will die. (There's nothing you can really do about this except being extra sterile).
5) ...Letting all of your cells get infected with your virus. All of them will make your protein...but then they'll die. This makes it a) impossible to have negative controls b) really hard to get anything done.
So now I know all of these things can happen to insect cells. And I know (at least a little bit better) how to stop them from happening. I'm proud to say I've had happy, living insect cells for about two or three weeks now, and I hope to keep them that way for a productive semester.
Photo credit: http://blog.ecosmart.com/index.php/2009/07/31/cabbage-looper-control/
