Spoke too soon

Yup, I spoke too soon. Got too excited. I must always remember...PCR-SSCP is cursed in the hands of researchers in lab number 7309. Cursed. The image looked great. The overstaining problem appeared to have been solved. I changed the resolution setting, and clicked go for the high resolution image of my gel to be scanned. This is what I got:



Waaay too dark! Not enough contrast to run and decent image analysis and develop electroferrograms. And this happened repeatedly for the next 4 hours. Garbage. As a last ditch effort, I turned the scanner off and repositioned the gel in a different corner of the scanner:

Much lighter background....but my bands were lighter too...disappearing. It had been 5 hours since I unhooked the voltage on the gel, and now the DNA was starting to diffuse through the gel. These faint fuzzy bands are worthless to analyze. I tried restaining to see if I could get the bands any darker. No good. The stain wasn't faded, the bands were just diffusing and fuzzing out. Boo. PCR-SSCP strike 3. This is the closest I've come to getting actual results. The bands I see confirm my hypothesis, so I know I'll be happy with the result and that it fits in nicely with the rest of my data. But this is the one last crowning experiment that I'm trying to finish so that I can include it in a paper. And I really want to get the paper submitted before I start taking my prequalifying exams this fall. To top it off, I'm just about out of the DNA I was using in these experiments. That means I have to go back to square one in order to repeat the SSCP. It'll take at least a week. Grr. That's the last time I celebrate until the data is actually in hand.