Summary: Here I used protocol outlined in Grover et al. (PNAS 2016) to make glass supported lipid bilayer. I used the lipids that Joe gave me. I repeated two key observations reported in Grover et al., and one additional key observation per personal communication from two groups.
1. MTs do not tend to cross each other when gliding on bilayer. They tend to cross each other when gliding on glass,
2. The velocity of MT gliding on bilayer is slower than that on glass,
3. The velocity of MT gliding on bilayer does not increase appreciably over time.
What seems important for glass supported bilayer (from literature review):
1. The glass surface needs to be ultra clean. This paper gets cited a lot for bilayer preparation: "Single-molecule fluorescence studies of a PH domain: new insights into the membrane docking reaction."
2. I also see people including NaCl in their SUV solution (0.075- 1M quoted). Sandra Schmid's lab published a few papers/protocols on using NaCl (http://www.utsouthwestern.edu/labs/schmid/).
Experimental condition: Identical with the exception of bilayer as indicated. The two bilayer flowcells were prepared separately as they would be if prepared on different days.
Video display frame rate: Identical.
MTs: I have not cleaned up the MT in these experiments, the bright bundles can in principle be removed before experiment.
Lipids: 0.2 mM SUVs in Grover buffer, diluted from 2 mM in ddH2O.