I'm still testing treatments to get cells to stick to coverslips (with more success):
I used more poly-L-lysine this time, but the important thing I realized was that I need to tap the slides briskly to loosen the cells that are just sitting on the surface but not attached,a nd to rinse a lot of medium through the chamber. This let me see that there were usually substantially more B. subtilis cells attached to the coverslip than to the glass slide.
So then I tested whether my competent H. influenzae cells would also bind to the coverslips. They did, better than the B. subtilis cells. I could see that many of the cells were only attached at one end, with the other end moving in the medium. But what was surprising was how vigorously the free parts of the cells were moving around. Not just bouncing back and forth (can Brownian motion be that vigorous?), but sometimes spinning around almost like cells tethered by their flagella (see this movie, which I think is Salmonella but might be E. coli).
But H. influenzae definitely doesn't have flagella. So I'm plating the cells I have onto LB and BHI plates with and without hemin and NAD, to check that some flagellated imposter hasn't snuck in to my culture. (It would have to make colonies that look just like H. influenzae colonies, and be sensitive to novobiocin...)
If at first you don't succeed, put it all back they way it was and keep lumbering along
11 hours ago in The Phytophactor