User:Andy Maloney/Notebook/Lab Notebook of Andy Maloney/2009/10/23/Motility

Motility update

So I was able to get motility working again. Here's a quick run down of what's been going on.

Things stopped working again.
I got a new batch of kinesin and gave my old batch to Brigette so she can check to see if it is working or not with her bulk assay stuff.
I have determined that the old antifade that Brigette made back in the summer is completely useless.
I have determined that the new antifade Brigette made does work.
There seems to be a lot of attaching/detaching going on with the microtubules and the kinesin. Plus, there are a lot of squiggles in the microtubules which lends one to believe that there is a lot less active kinesin in the vials we have.
I have not ruled out if it is the ATP we are using as a cause for some distress. I should point out that the ATP is in PEM and not the usual Tris-HCl. I didn't see any reason why we wanted to put it in Tris-HCl other than it not hydrolyzing as quickly as it would in a buffer at a pH of 6.8. Who knows. It could be an issue. I will test this though.
Andy Maloney 19:29, 23 October 2009 (EDT): So I just noticed that after some time sitting on the microscope, things are moving better and more uniformly. I believe I heard Koch talk about this. Well, it turns out that the ATP is good because motility is rocking right now. I think I'm going to make sure that I don't take data for a specific amount of time. In other words, do what Koch suggested and not start until after 15 minutes has gone by after sealing the flow cell.
- Ant did bring up a good point and that is, why has no one studied kinesin accelerating?

^{SJK 23:25, 23 October 2009 (EDT)}