IGEM:IMPERIAL/2008/Prototype/Drylab/Motility data collection

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(Manual Tracking Methodology)
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*We intend to track for each cell:
*We intend to track for each cell:
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**The centroid  
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**the centroid  
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**The part of the cell in the direction of movement
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**the part of the cell in the direction of movement
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*We attempt to reduce the bias in our manual tracking
*We attempt to reduce the bias in our manual tracking
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**By disabling the view path option  
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**by disabling the view path option  
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**By displacing the cursor away from the tracked cell before adding a new point
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**by displacing the cursor away from the tracked cell before adding a new point
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==Data Clustering==
==Data Clustering==
{{Imperial/EndPage}}
{{Imperial/EndPage}}

Revision as of 11:01, 4 September 2008

Home The Project B.subtilis Chassis Wet Lab Dry Lab Notebook


Contents

Our Approach

Material

We will be using the Zeiss Axiovert 200 inverted microscope with a fully motorised stage, controlled by Improvision Volocity acquisition software. This system offers a full incubation chamber with temperature and CO2 control, a large range of filter sets from UV to far-red and a highly sensitive 1300x1000 pixel camera for fast low-light imaging.

Video images are captured into memory by the system at a basal video frame rate of 16.3Hz. This can be further increased by performing binning.


Microscope Video Characteristics

From our experience on the microscope, we observe the following:

Bacteria are rod shaped. As the bacteria transverse across the plate, it may change shape and orientation. Due to tumbling in the z-direction, bacteria may go in and out of focus. This leads to change in light intensity. Background noise is also present. We also observe a few static noisy spots over all images ( might be due to dust particles ???)

  • Video 1

Our first video.The cells are out of focus.

Notice (see below) the presence of some noisy circular patterns. These patterns are found at fixed positions over all frames.

Image:Noise1.tif

Intensity properties for the background, cells and noisy circular patterns can be found below. As well as shape characteristics for the cells.

Background Cells Circular Patterns

  • Video 2

This video was recorded with higher contrast and with a better focus than for Video1. The video was shot at 12 fps for a duration of 1 min. The cells and their wall can clearly be seen over all frames. Note that the inside of the wall is darker than the cell membrane.

Notice (see below) the presence of some noisy circular patterns. These patterns are found at fixed positions over all frames.

Image:Noise2r.tiff

Intensity properties for the background, cells and noisy circular patterns can be found below. As well as shape characteristics for the cells.

Background Cells Circular Patterns

Manual Tracking Methodology

  • We intend to track for each cell:
    • The centroid
    • The part of the cell in the direction of movement
  • We attempt to reduce the bias in our manual tracking
    • By disabling the view path option
    • By displacing the cursor away from the tracked cell before adding a new point

Data Clustering


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