Skip to content

Changes

Page history
Short description of tracking pipeline authored by Antonio Politi's avatar Antonio Politi
Show whitespace changes
Inline Side-by-side
AdaptiveFeedback.md
View page @ 4bff5441
...@@ -103,7 +103,25 @@ Segmentation of the DNA channel (red) returned 17 ROIs. <br/>Only for ROI 10 and ...@@ -103,7 +103,25 @@ Segmentation of the DNA channel (red) returned 17 ROIs. <br/>Only for ROI 10 and
Segmenation of the DNA channel (red) returned 2 ROIs. <br/> For ROI 1 and 2 the intensity of the fluorescent protein (green) is within the specified range. | The cell closest to the center of the image is picked (green outline).<br/> The red outline shows a cell that has not been picked because it is too far away from the center of the image. <br/> Coordinates for the two FCS points are passed to MyPiC. Segmenation of the DNA channel (red) returned 2 ROIs. <br/> For ROI 1 and 2 the intensity of the fluorescent protein (green) is within the specified range. | The cell closest to the center of the image is picked (green outline).<br/> The red outline shows a cell that has not been picked because it is too far away from the center of the image. <br/> Coordinates for the two FCS points are passed to MyPiC.
## [<img src="./images/up.PNG">](#back) <a name=windowsregistry></a> Windows registry commands ## Object based tracking
MyPiC and the FiJi macro can be used to track objects based on the segmentation result. The tracking is simple nearest neighbour where the coordinates of the segmented object closest to the center of the image are passed to the microscope.
We define two images:
* The tracking image, the image used to compute XYZ coordinates, e.g. a low zoom and low resolution image.
* The acquisition image, e.g. a high resolution image.
Depending on the sample a single image can be used for the tracking and acquisition. Typically for a fast moving or large object one would use a low zoom tracking image and high zoom acquisition image.
To enable object based tracking the processing settings in **MyPiC** must be `Online image analysis` and `Track XY` and/or `TrackZ` for the tracking image. Within [**Automated FCS**](https://git.embl.de/politi/adaptive_feedback_mic_fiji) use the command `focus` and `Pick particle in = center`. This sends the coordinates computed in **Automated FCS** to **MyPiC**.
> **Airy detector**: It is optimal to use the airy detector for the tracking as well as the acquistion image. This avoids repeated changes of the settings and minimizes hardware wear out. Maintain very similar imaging settings between tracking and acquisition image (e.g. just change the zoom, pixel dwell, laser lines). In particular avoid using different pinhole sizes. Analyze the channel that corresponds to the sum projection of the 32 airy detectors (raw image not the airy processed image). For a single channel image this corresponds to channel number 2. If not sure which channel to use open the Airy raw image in ImageJ and find the sum projected channel. ** Save the images in czi format**.
# [<img src="./images/up.PNG">](#back) <a name=windowsregistry></a> Windows registry commands
MyPiC communicates with external programs by reading and writing in the Windows Registry: MyPiC communicates with external programs by reading and writing in the Windows Registry:
......