I’ve not written in a bit about the trials and tribulations of getting an electrophys rig up and running. As an update, I believe that I’ve located the major problem in the system. The ringers solution being used was made by a previous grad student, and had been sitting in the fridge for some time with parafilm over it. Well, it turns out the parafilm wasn’t sealing things in so well, and the solution was supersaturated to the point of precipitating solid. I didn’t realize this was way off until looking up ringers solutions and finding that not not that much in the way of salts go into the stock, and therefore precipitation shouldn’t be happening. This was making the electrode pulling almost impossible (I’m guessing the ones I pulled before will be rediculously high impedance with the new solution), and led me to try innumerable solutions. While some might consider all that time wasted, I really don’t look at it that way.
In graduate school, I believe, one needs to develop a bit of an immunity towards repeated obstacles. Keeping focused on finding solutions, and realizing that much time will be spent getting past obstacles is the only way to survive. If you don’t expect all that, or come to expect all that, I’d imagine that things could be quite miserable. I think that’s also probably why graduate school isn’t for everyone. If one is already frustrated after undergrad, you’re probably not going to pick up steam spending another several years working on things. If, however, you’re not, and you keep in mind Rule #1 (most of your time will be spent trying to fix things), it can be pretty awesome. You basically get to pick out your own project and work on what you like without worrying about getting product out the door by next quarter, regardless of flaws still present. I’m not saying here that one’s research ends up being perfection in the end, I would say instead that things don’t generally get swept under the carpet. They definitely do when immediate profit is the goal.
What I thought initially would be hard, proved to be exceptionally easy then exceptionally impossible. I’d heard from others long before I’d gotten into the business of pulling my own electrodes that this would be a road fraught with dismay and frustration, or at least fraught with cursing and many discarded electrodes that did not work out. When I started in on this, I got our lab’s shared laser puller up and running. This part wasn’t hard, and merely required reading the directions. Then after less than half a day of pulling electrodes, I was getting ones that were around 30-40 Megohm. What, I found soon after, however, was that that was 30-40 Megohm with a tiny bit of air still holding out near the tip, and that filling the very tip of the electrode by capillary action is necessary to get a proper electrode. As an aside on this matter, if you have crazy noise issues that creep into your signal and you’re sure it isn’t clogged: you may have a bubble in there somewhere. It seems however, that getting this capillary action going becomes exponentially harder and slower as the tip diameter closes down. Currently, this is showing itself in the sense that I can only get 4-10 Megohm micropipettes that actually sip from the KCL I’m placing them in.
I’m writing this, and posting it, both for my own reference, and for the fact that I couldn’t really easily find anything on methodology for pulling electrodes. Perhaps I should start a wiki for such things? It would not have to be limited in scope only to that particular topic, but rather be a site covering lab technique and the various ins and outs. Personally, this rather appeals to me for the reason that a lot of the manual lab work requires what seems to be almost a black art, and either someone in your lab can show you this art, to some extent, or you just have to spend enough time banging into things that you figure it out on your own. I will have to think about this…
So, I’ve recently been training up on doing electrophysiology in the lab, and I thought I’d blog about the experiences a bit (mainly technical details) as I go along since I’ve encountered a number of issues along the way, and some I’ve resolved so far, whereas others still need some work to get going.
This is basically both for personal reference, as well as for anyone who happens to be interested in these details. One of my standard solutions when looking for information about things that don’t seem to be working correctly is through googling, and I’ve really not been able to find any good information on all this stuff.