Tracking fast cell migration in phase contrast 2D 10x movies

[Michael Shannon]

Hi all

Hope everyone is good.

I had a chat with Alexandre Dufour after a talk he gave at Imperial college the other day, and he advised me to post some data on here to see whether anyone can help with my problem of tracking phase contrast cells.

The reason we started talking was that I have tried a few different programs with limited success in this endeavour, but I have never tried 'contour mapping'.

It would be really useful if I was able to automatically track these cells (and filter out round/non moving ones) without resorting to cell/nuclear dyes, as these primaries are quite sensitive to such treatment.

Below is a link to a dropbox folder containing two files. The imaging isn't perfect.

Number 1 has pixel size 0.29 um/px and FPS 0.31

Number 2 has pixel size 0.29 um/px and FPS 0.02

https://www.dropbox.com/s/2ewi72uwlvkxvhu/for%20ICY%20contour%20mapping.zip?dl=0

If anyone can track these cells, I will buy them a pint!

Mike

[Irina Catrina]

Hi Mike,

I looked at your files and for files similar to 1.tiff if I may suggest that you use flat-field correction (ImageJ link:http://imagej.net/Image_Intensity_Processing#Flat-field_correction).  Also, I found that for me the spot detector worked better for 1.tif after I inverted the image, but for 2.tiff both the original or the inverted gave similar tracking results.  Tracking for 2.tif worked much better than for 1.tif, and one of the reasons is the higher sampling rate for 2.tiff. 

For spot detection I used the 25 pixel scale with sensitivity 70 for 1.tiff (bright spots over dark background), and 80 for 2.tiff (dark spots over bright background for the inverted stack or bright spots over dark background from original stack).

After that the round/non-moving cells were filtered out using the motion profiler.  I attached three snapshotps with the tracks.  Are these results similar to your limited results?

Irina

[Michael Shannon]

Hi Irina

Thanks for this. 

Yeah so the tracks look pretty good, enthused that 'spot detection' seems to have worked...

Do you have the speed data for these? Does it give an overlay of the tracks on top of the movie also?

Mike

[Irina Catrina]

Hi Mike,

Yes, you can get the speed information in the track manager using the
Motion Profiler. But you may want to carefully optimize both the spot
detection and tracking parameters.

I attached the speed info for all these (preliminary) tracks as an xlsx
file. Also, find below a dropbox link for an AVI movie showing the
tracks with a total displacement greater than 50 pixels, which are
represented as tails of 10 consecutive frames at 10 frames/sec.

https://www.dropbox.com/s/7jjhtdcvsejiq8o/Rendering%20of%20%272_inverted.tif%27%20view%20%28RGB%20rendering%292.avi?dl=0

Hope this helps,
Irina

On 6/6/2016 5:59 AM, Michael Shannon wrote:
> Hi Irina
>
> Thanks for this.
>
> Yeah so the tracks look pretty good, enthused that 'spot detection'
> seems to have worked...
>
> Do you have the speed data for these? Does it give an overlay of the
> tracks on top of the movie also?
>
> Mike
>
> On Sunday, June 5, 2016 at 6:38:59 PM UTC+1, Irina Catrina wrote:
>
> Hi Mike,
>
> I looked at your files and for files similar to 1.tiff if I may
> suggest that you use flat-field correction (ImageJ
> link:http://imagej.net/Image_Intensity_Processing#Flat-field_correction

> <http://imagej.net/Image_Intensity_Processing#Flat-field_correction>).

> <https://we.tl/5setztagwd>

>
> If anyone can track these cells, I will buy them a pint!
>
> Mike
>
>

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