Experiment 3: Cell Motility Part 1:  First Trial of Cell Movement analysis

1.Take a 60 mm Petri dish and add about 5x 105 cells in 3.5 ml of HL5 (a bit under 1x105 cells/ml).

2.Allow the cells to settle for at least an hour

3.Using Micro-Manager- take a time lapse movie of the cells for about 30 minutes using the 10x objective and phase contrast.

4.You can try the other objectives as well so you get a sense of how the field of view varies so you can pick an appropriate objective for your experiment.

5.When done watch the playback so you can see the frame to frame movement and overall movement.  This will help you to figure out how frequently to take images for future experiments.

6.Watch the movie to get a descriptive view of what happens. Do all the cells move? Do they all move at the same rate? What can you say about the mechanism of movement from watching?

7.Use mTrackJ to analyze the movement

Goal- You will be repeating this experiment next time with wild-type and myosin mutants, so this is a trial to work out the software and parameters you will use for that experiment.  You need to figure out what density of cells to plate, how frequently to take images, what magnification to use, how long to image each cell type, etc.