Validation

A synthetic video of five cells with changing shape was used in the evaluation of tracking software. Each cell was tracked using respective tracking software, and their coordinates in pixels were obtained.

The following steps were taken to analyse tracking data:

1. Convert synthetic trajectory coordinate points into pixels.
2. Calculate the displacement in each frame for both synthetic and tracking data
3. Calculate the distance travelled in each frame for both synthetic and tracking data
4. Obtain the errors in coordinates, dislacement and distance travelled by the cell

The errors associated with manual tracking and Volocity software are shown below. All errors are given in terms of pixels.

Manual Tracking

Error in X Coordinate	Error in Y Coordinate
Mean = 1.7354, Standard Deviation = 1.7187	Mean = 1.2524, Standard Deviation = 1.0568
Error in X Displacement	Error in Y Displacement	Error in Distance
Mean = -0.0022, Standard Deviation = 1.0002	Mean = -0.0018, Standard Deviation = 0.6795	Mean = -0.1506, Standard Deviation = 0.9196

Volocity

Error in X Coordinate	Error in Y Coordinate
Mean = -1.7149, Standard Deviation = 0.1376	Mean = -0.4907, Standard Deviation = 0.1614
Error in X Displacement	Error in Y Displacement	Error in Distance
Mean = 8.2732e-005, Standard Deviation = 0.1111	Mean = 6.1416e-004, Standard Deviation = 0.0712	Mean = 1.4262e-004, Standard Deviation = 0.0947

Evaluation

Volocity

Pros

• Lower error in terms of tracking centroid than other tracking software.
• Less time consuming.

Cons

• Failure to track cells continuously over a long number of frames > 70
• Baised to cells which run, rather than tumble

Manual Tracking

Pros

• Able to discern cells visually
• Able to track over larger number of frames

Cons

• Time consuming
• Higher error in terms of tracking centroid as compared to tracking software.

Conclusion

• Manual tracking will be used to track B.Subtilis.