Wash. Rock. Aspirate. Repeat.
Six weeks of diligent work had culminated with six delicately miniscule mouse embryos, so transparent that my eyes often lost track of them as they sloshed around in solution. My mentor and I had spent the past month running PCRs, conjuring up gels, and transforming plasmids into bacteria, all in preparation for this final step: the in situ hybridization. The purpose of this complicated sounding process was to insert RNA probes into the embryos, which would allow us to observe the expression pattern of the gene Fgf12 in embryonic heart development (specifically in the atrium). The next four days would be filled with myriads of strange solutions comprised of mysterious, unpronounceable substances. These deceptively clear liquids were capable of performing painstaking chemical processes at the molecular level, something that my mind could hardly fathom. Each step involved washing the embryos with different solutions (which could contain anything from methanol to antibodies), then rocking them for a certain amount of time, and lastly aspirating out the no longer needed liquid.
Wash. Rock. Aspirate. Repeat--with a different solution.
This procedure eventually became ingrained in my mind as it became my daily routine, but as it turns out, it was nowhere close to being uneventful. One solution that I’ll probably forever have nightmares about is the harmless PBT buffer, which when combined with proteinase K, was capable of removing protein from the embryos. This would allow our RNA probes to more easily penetrate the relatively thick outer layers of the embryo. However, what I didn’t learn until it was almost too late was that the solution would gradually cause the embryos to disintegrate if they were left in solution for too long. Fortunately, the embryos managed to miraculously survive the harrowing ordeal, and as I frantically scrambled to aspirate the solution before it could do any more damage, I carved myself a mental note to never let a disaster like that happen again. From then on, my eyes meticulously scanned the lab protocol at every step, double-checking to make sure the solution I would be using wouldn’t dissolve my embryos into oblivion.
Wash. Rock. Aspirate. Repeat.
On day 4, we finally reached the last step, which was adding the BCIP/NBT solution, which would stain the sections of the embryo where the Fgf12 gene was expressed. Since the solution was light sensitive, we had to place the vial in a box, encase it in aluminum, and hide it in a drawer. Now, all there was left was an agonizing thirty minute wait. After half an hour of anxiety inducing anticipation, my fingers slid open the drawer, peeled off the aluminum foil and popped open the box. Sure enough, the barely detectable embryos contained a speck of purple in the center. Closer examination with a microscope confirmed that our experiment had been a success: the atriums of the embryos were stained a deep violet, while the ventricles retained the transparency of the surrounding tissue. At that point, my excitement was almost palpable, but since we were technically still in a lab, I had to content myself with high-fiving my mentor.
As the summer draws to a close, I realize that I eventually have to part ways with the lab. On the flip side however, I’m incredibly grateful to have had the opportunity to delve into such ground-breaking research. Regardless of where my future takes me, being able to explore the vast fields of science and medicine this summer has lead me to the conclusion that, just like how those embryo hearts will remain permanently stained, my fascination with the science of discovery will never fade.