The Dark Room

Some people at Toolik upon the eve of their departure get to throw up their hands in mock despair and say, “Its going to be so weird going back home where it gets dark! I’ve totally forgotten what that’s like!”. To my great dismay, I will not be able to say that and really mean it, because every day I spend a couple of hours in… the DARK ROOM.

The dark room is the only place in camp where no light shines 24/7. We run two analyses in there, OPA for ammonia, and chlorophyll. A fluorometer works by shining a light on an object and detecting how much of the light the object reemits via flurescence. To do this, it needs to be in the dark. On top of that, the reagent for OPA is light sensitive, and chlorophyll needs to be kept in the dark all the time, both of which reinforce the unfortunate necessity of the dark room being, well, dark. As the newest member of the Kling team, I got the exciting assignment of running OPA. As Sara so nicely first described the process to me so as not to discourage me, “Its a little long… but you get to spend a lot of time in the dark room by yourself which is kinda nice”. What she really meant is, its mind-numbing drudgery that takes forever, and you get to spend most of it sitting by yourself in the dark listening to other people in the lab having fun. I’m exaggerating, its really not that bad, and getting some time alone is nice… at least that’s what I tell myself when I’m in the dark squinting at the fluorometer.

So what do you need to run OPA? First off you need the following, sample, working reagent, buffer solution, and standards. The buffer solution is a weaker version of the working reagent, and standards are solutions of known ammonia concentration. We use these to make a standard curve which is essential to calculate the concentration of ammonia in the samples. The principle is as follows. There no way to directly test for ammonia, so what we do is add a reagent called OPA to the samples, let the mix react for 16 to 24 hours, and then read the fluorescence off the fluorometer. Except that in addition to the samples that have straight working reagent added to them, we have two more sets of samples that are spiked with two volumes of highly concentrated standard, and another row of tubes to which is not added working reagent but buffer, which is a weaker solution of the working reagent. We do this for calculation reasons involving matrices. Feel confused? Me too.

Though I haven’t tried to understand the math surrounding the production of the actual values of ammonia, what I do understand is that OPA quadruples the time I have to spend with the fluorometer, because if I’m running a medium run of 30 samples, I actually have 120 tubes to rinse, pipette, shoot up, and read; plus 12 more for the standards. Its an ordeal. After returning from the field on Mondays, Wednesday, and Fridays, I’ll start the long process of preparing my tubes for OPA. In the dark room I rinse them with DI water and then buffer (~ 45 minutes), then I pipette my samples and standards in the common area of the lab (~ 90 minutes) then back to the dark room to shoot up, i.e. add OPA or buffer to the tubes (~ 45 minutes). Then the next day back to the dark room to read the samples (~ 2 hours).

I have spent so much time with the fluorometer I feel as though it is another person, a very needy and fickle person. It was not enough that three labs were battling over the precious time slots to spend at its side, it just HAD to break down half way through the summer. We yelled at it, threatened to throw it on the ground, but it didn’t listen. I guess it finally got its attention fix after a week and a half of being fawned over, taken apart, and settings readjusted, before finally deciding to work again. Of course at that point, our backlogs had accumulated so quickly that it didn’t even have to try to give us good data, we took whatever shoddy numbers it threw at us. What an unappreciative piece of equipment.

Sometimes Jason goes all misty eyed and says, “Well you know back in 08 when I ran OPA, we had to pipette the samples in the dark room on top of everything else. That was the woooooorst.” Very soon, I’ll be able to join that exclusive club of ex-OPAers. Sometimes when I’m alone squinting at the ever fluctuating values of the fluorometer I dream about the last time I’ll exit the dark room, stumbling around for a few seconds while my eyes get used to the brightness, and look back upon OPA as Jason does; with a wistful fondness and a sense of accomplishment from having gone a whole summer without throwing the fluorometer on the ground. Even once.