Histology 101 Now In Progress, Among Others
The students working with me on the telescope shiner project are doing their first histological preparations this week, using scarlet shiners left over from the parasite project. The lab has a protocol for dehydrating and fixing the tissue so it can be mounted in paraffin, sliced and stained. The basic direction is: soak in Bouin's Fixative, two multiple-hour soakings in water, 70% ethanol, two round of 95% ethanol soakings, two rounds of 100% ethanol soakings, xylene, and tolulene/paraffin. It can be done in about 16 continuous hours of attention, or broken down into two days of paying attention to moving tissues from solution to solution. This is a technology that hasn't much changed since the 1930s. I think they're getting it, it's really nothing complicated but more similar to following a recipe to bake a cake.
I heard from Jo in the Alabama State Lands Division the other day. She called to say that they've granted me a permit to do scientific research at the Walls of Jericho, i.e. monthly collections of 30 telescope shiners. We're legal to go out this Saturday and seine telescopes. It will be something of a challenge, the temperature will be slightly above freezing but with sun if the forecast is to be believed. But we have at least 5 pairs of waders between 6 of us, so we'll be OK. We'll meet Nick at the vehicle entrance to the tract at 8:30 and he'll give me a key to all of the gates on the road into state property.
Yesterday I ran my first PCR in 2 years. Tomorrow we hope to run the results on a gel and see if they worked. I ran 2 DNA samples of Fundulus heteroclitus from Marion, Massachusetts, that I found in my freezer that I had neglected before, along with 3 samples from Nantucket and 1 sample from Virginia Beach. The last four had produced good amplification bands before but the sequences were screwed up, and I hope the Marion samples amplify. I extracted them in 1996 while I was still working out of UMass/Boston, and they've been frozen since. Spectrophotometric analysis at the time showed them to contain lots of DNA and relatively little protein (a PCR inhibitor) so in principle they should work well. I'll know by this time tomorrow...