Motility Assays

Motility Assays used to study motor use two different geometries, called the bead assay and the gliding assay. In the bead assay the microtubules/microfilaments are fixed to the surface and the respective motors move on them. A bead is bound to one or more motors to allow visualization or the exertion of force in an optical trap. In a gliding assay, the tails of the motors move the microfilaments/microtubules across the surface.

The experimental set up for the gliding assay involves four basic steps:

1)adsorbing a protein monolayer onto the surface by flushing in a solution containing casein or albumin to reduce denaturation

2)adsorbing the molecular motors to the penetrated surface by exchanging solutions

3)adsorbing the microtubules/microfilaments to the motors from a third solution

4)imaging motility with optical microscopy

In the bead assay, motors and filaments/ microtubules are interchanged in steps (2) and (3)

Flow cells

A simple flow cell of a classic motility assay allows easy exchange of solutions, illumination with light and a fluorescence microscope to be used which will aid in visualization.

The most common way  to construct a flow cell is to place two strips of double-stick tape/ spacers on a glass slide ~7-10 mm apart and cover with a 18x18 or 22x22 mm coverslip. This results in a ~12-15 µl flow cell. Solutions are pipeted on one side and sucked out the other side by capillary action using Whatman #1 filter paper or a Kimwipe.

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