Freaking micropipettes! Like fairies' wands! We'd get a lot more science done if we could make ourselves Lilliputian at will. And I should be good at this sort of thing! I made quite a name for myself as a youngster by making beautiful tiny Christmas ornaments that all the adults exclaimed at! I swear that science is like magic sometimes!
The resident electrophysiologists in the lab have been nothing but generous in their attempts to teach me how to patch cells. I greatly appreciate this, because I know that I'm exasperating. Especially today. I think I broke six of the ten glass micropipettes that I pulled today. I hope it wasn't any more than six, and if it turns out to have been more, I am not going to tell you. Of the four that I miraculously refrained from breaking, two of them turned out to have tips of insufficiently large diameter. Guess I got too heavy-handed with the pedal switch that controls the tiny heating filament over which one shapes the micropipette's tip for patching. That's one of those things that one doesn't necessarily discover until one realizes how shitty the seal resistance is. For those of you who are not electrophysiologists or their interested friends: This is far into the arduous cell-patching process. You really don't want to wait until that stage to discover that your tips are inadequate.
I mounted a micropipette incorrectly, which caused it to dive into the cell dish's bath upon a computerized command that should have suspended it neatly above the dish's center instead. It was almost funny. Meanwhile, the bath level kept mysteriously sinking, which never happens when the senior grad students patch. All unaware, I gazed through the microscope lens and slowly lowered the micropipette toward my chosen cell, thinking, "That's odd, the pipette shadow keeps looking like it should when the pipette is still suspended above the water--" only to see the crystalline tip crush open the cell and shatter into a dozen miniscule fragments as it struck bottom. Awesome.
Too fast or too slow, and heartily endeavoring to concentrate, I kept at it until the sun had long gone and the grad student who was training me discovered that I'd risen at five and begun work at 7:30 am and said kindly that no one should patch cells when she's tired. And I am tired. And I would like to continue to get to the lab by 7:30 am, and I'm old, so I will go to bed now.
11 comments:
I think this may be more common than you think. One of the boys went through don't-ask-how-many scanning electron microscope probes before getting it right.
Turns out that his mistake was following the documentation. Seems that "everyone" knew that there was an error there but hadn't annotated it, just passed it along over coffee until nobody remembered that "the way to do it" wasn't in the written instructions.
Don't listen to D.C. Yes, steep learning curves are common in science. However, you aren't doing science. You are performing the dark art of electrophysiology. There isn't one particular rookie mistake you are making that will magically make everything all right.
If you choose electrophysiology, you will sit at the rig for 8-12 hours a day. On a good day, you will get data from three cells. Once a week or so it might be informative data. You will pull 30 or 40 tips every day and go through them all. You will give up coffee, because it makes your hands too shakey. You will give up everything in your life except sleeping and patching (you can eat at the rig). And you will learn emotional control to remain calm at all times. It could make you stronger- you will need to be able to spend 10 hours at the rig and fail to get any good data and go back and do it the next day, without ever having your confidence shaken.
Becca, I don't think we're disagreeing: there's no easy way to do a hard thing, and learning a hard thing takes longer.
Anything that you do down at the cellular or atomic level is going to be hard. For crying out loud, some of this stuff is literally running up against Heisenberg's limit.
I collaborated with a lab that does patching and electrophys during my masters. I will never go near a rig for the reasons that becca states. Electrophy is the DARK SCIENCE. Ununderstanding electrophysiology and being able to explain reversal potentials etc is the first step. That was its own special hell learning for me. You will master this, but it will take time and patience. Trust me when I say your senior lab mates have been there and understand. No one does this on their first 20 attempts.
My point is that, while *understanding* the data follows learning curves like other things (in other words, it's very hard at first but one day you look back and are all 'wow, I KNOW stuff!'), *generating* electrophysiology data will always be hard. It was hard for the postdoc who had been doing it for a decade who was calm and bright and talented and who did it every freakin day. I didn't realize that (he never *told* me it was hard; and I didn't realize how many times he didn't get good data), and when I never had a breakthrough in generating the data, I got very depressed. I want Juniper to know that it's *fine* to give up on electrophysiology (ultimately, I landed in a lab I was much happier in) but that if she wants to stick with it, she *cannot get down on herself because it is hard, and it will still be hard even when she is 'supposed' to know how to do it*. Probably somewhat true in science generally, but electrophysiology really is in a class of it's own (ok, so it's in a class with microinjection of embryos for creating knockout mice, if I have been correctly informed. But still, those two things are a quantum leap in physical difficulty away from every day molecular lab techniques).
Becca: Emphatic agreement. Some things are hard. Period. At best, after much study and practice, they're only skullbustingly hard; to begin with, they bust your skull and then do unspeakable things with the contents.
Assuming that you've actually been granted access to the tribal knowledge that makes it possible to do it at all (not a guarantee by any means.) Otherwise it's much, much worse.
The cryostat used to be my worst enemy.
This comment thread made me laugh so hard.
D.C., I only ever used a scanning electron microscope to analyze archaeological specimens wrapped in glorified aluminum foil. And that was several years ago. So I have no idea what probes you are talking about. Clearly, I was not paying enough attention; it is not ingrained in my memory.
In all seriousness, I appreciate your encouragement. But I am pretty sure that I blow at patching cells, and that I am doing more than one thing wrong.
Becca . . . wow. Are you exaggerating? That's not a sarcastic question. The resident electrophysiologists in our lab told me that electrophysiology was a little difficult. They said nothing about 30-40 tips a day and veritable data droughts. Plus, an incredibly talented postdoc in our lab, who had never attempted electrophysiology before, successfully patched 12 cells on his second day of training. So I suspect that I just suck at this.
I am learning electrophysiology as part of my second rotation. I will keep practicing until I can patch a cell without assistance, and I will increase my efforts. I am nothing if not determined. However, I highly doubt that I will choose electrophysiology as a focus. It is becoming crystal clear to me that I am much better at other things. Besides, my hands are already shaky. From the fucking Wellbutrin. Which is one of two reasons why I'm already abstaining from caffeine as well as alcohol.
ScientistMother: I do feel a little like Harry Potter whenever I try to patch cells.
Dr. Isis: You know what's hilarious? Cryostat is actually something I rock at!
D.C.-I love that phrase. Unspeakable things, indeed!
Juniper- As I'm sure you've realized by now, the rig is everything. I've only seen electrophysiology in *one* lab. With two different rigs. Good equipment *helps*; it's always possible that the difficulties I and others had were partially related to the challenges of the equipment.
That said, what killed me in the e-phys lab was listening to what my PI said about how things should be easy, and not asking specifically how long things took the other benchscientists. Ask the specific questions- how long a particular figure in a paper took to generate, and what the longest part of the paper was, and that kind of thing. Past challenges aren't always a guide to future challenges, but it'll give you a good idea how much of a happy-face people are putting on (of course, happy-faces can be very useful for coping with things that are intrinsically challenging).
But my best guest? Your talented postdoc will be a legend... because patching cells is a very rare talent.
I tell you how hard it was for me so you *don't* come to the erroneous conclusion you suck. It's possible to suck at it, but it's NOT possible that you can know if you suck because you are not that postdoc.
This is a pretty hilarious thread you have going here. Sounds like that postdoc in your lab is either: 1) exaggerating his abilities or 2) a total freak of nature. I'd go with 1.
If you want to come cryostat all of my shit, you would be certainly welcome.
Post a Comment