Slowly, but surely I am getting into the habit of working. 7:30 wake up, leave 8:30, work at 9:00, lunch at 1:00, and end workday at 6:00-at this point I am starving so dinner immediately follows. These past two weeks I have been gaining different skills in the lab, most notably learning how to do tissue culture and how to use a wide field live imaging microscope and a confocal microscope.
I was intimidated by tissue culture at first. When Amy first showed me how to do it I feverously took notes. The next day I typed these notes and studied them intensely because I needed to subculture the next day. Kuayping (who is now watching over me because Amy went to the US for two weeks) showed me again and then let me try.
The steps to tissue culture of fairly simple; remove the old medium, wash with buffer, add trypsin (an enzyme) to dislodge the cells, remove the trypsin because you don’t want to have too much enzyme, incubate the cells so that the remaining trypsin can do its job, add a buffer that inactivates the trypsin because you don’t want to digest your cells, suck up and squirt out the cells to put them all into solution, and finally add new growth media and call it a day. Overall, pretty simple, the most complicated procedural task is remembering which button on the electronic pipette sucks up liquid and which squirts it out.
But, the procedure for tissue culture isn’t what is tricky-being sterile is the though part.
Your tissue culture to do list:
Spray down the hood with ethanol-check. Spray every bottle that enters the hood with ethanol-no problem. Wash your gloves with ethanol before you put them in the hood-really cell culture is an alcoholic’s dream (just kidding, never ever think to drink the ethanol).
Why all the ethanol? You need to have a completely sterile environment, 70% ethanol will kill most biological organisms that threaten your sterile hood atmosphere.
Alright-I’ve been working spray bottles as water guns since the second grade; so far everything I’ve explained about sterility shouldn’t be a problem. But now we in da hood (science gangster reference, not typo).
When working in the hood you need to be much more conscious about everything you do. This means when I reach over my sterile pipette, with my sterile glove, for my sterile media bottle I have risked contamination of my sample because my unsterile arm may have dropped dust on my once sterile pipette. Even when being rigidly careful, Kuayping still pointed out how I broke the sterility code at least three times-which means throwing out a perfectly clean pipette tip (I felt bad about that).
During my first tissue culture venture, I literally felt like I had never used my hands or arms before. To set the dramatic scene:
The setting: Biopolis, Singapore on another 85 degree day, 100% humidity. On the fifth floor of IMB which is kept at about 10 degrees (I wish I brought more long sleeved clothing), in the tissue culture room whose doors open only if you wave your hand in front of a scanner to the left of the door (it is really cool, I couldn’t figure out where else to put that). Devinn, the narrator of this telling tale, is diligently working on cell culture. The hood is gently humming in the background as it sucks up air to keep the environment clean. She is working with a flask which has the general shape of a metal whisky container that you see men have in the movies-except lie the tin bottle flat on the table.
My motivation: I need to make sure I dispense the media everywhere in the flask except the bottle neck of the flask because it is too close to the air and that compromises sterility.
The Conflict: I can’t figure out how to rotate a foot long pipette so that I cover the area around the mouth of the flask without touching it. I awkwardly struggle and fail multiple times.
The Climax: Kuayping either out of pity, boredom, or frustration reaches in to demonstrate this task.
The solution: he uses his wrist!!
Anticlimatic? Maybe. My point is that my arms felt so foreign to me that this simple motion was revolutionary.
On my second time sub culturing, Kuayping didn’t watch over me- I was surprised. It was like the mother bird pushing the hatchling out of the nest- thankfully I flew instead of splatting on the ground (This reminds me of a really funny website with analogies-I suggest you check it out http://writingenglish.wordpress.com/2006/09/12/the-25-funniest-analogies-collected-by-high-school-english-teachers/).
Every attempt after the first has gone fairly smoothly-now that I know the procedure by heart I can focus more on my movements to make sure I keep everything sterile.
I will dedicate another blog to my microscopy work once I have some beautiful fluorescent photos.
Till then, keep it real USA