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The Nuclei Tracking recipe in Aivia detects nuclei and tracks their motility over time in fluorescence time-lapse microscopy images. The recipe measures the morphology, intensity and motion attributes of the detected nuclei for comprehensive characterization of nuclei migrations. In addition, the recipe tracks nuclei divisions and generates lineages for all detected nuclei.

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Table of Contents

maxLevel	3
exclude	[4]

Parameters and Presets

Parameters

Recipe parameters for Nuclei Tracking and their descriptions are summarized in the table below.

Preset Group	Parameter Name	Min Value	Max Value	Description
Detection	Background Removal Factor (Remove Background only)	0	100	Adjusts the sensitivity of the background removal operation; a lower value will preserve larger objects and more background variations
	Contrast Threshold (Remove Background) Intensity Threshold (Skip Remove Background)	0	255 (8-bit) 65,535 (16-bit)	Adjusts the detection sensitivity on the segmentation image; when Remove Background is selected, the threshold will be applied to the background removed image; whereas when Skip Remove Background is selected, the threshold will be applied to the input image; a lower value will detect bigger and more objects
	Fill Holes Size	0	5,000	Adjusts the maximum size threshold for filling in gaps inside a detected object; a lower value will preserve more holes in the detection
	Smoothing Factor	0	100	Adjusts the amount of smoothing applied to the outline of the detected objects; a lower value will preserve more of the object's morphological features
Tracking	Object Size	0	50,000	Specifies the range of objects to be included in the analysis results based on the area of the detected objects
	Max Search Range	0	5,000	Specifies the maximum distance for track-point matchmaking between successive time frames; a higher value will expand the search distance for fast-moving cells
	Min Track Length	0	50,000	Specifies the minimum number of time frames before a detected object is considered a valid track; a lower value will generate more, and often shorter, tracks
	Separation Factor	0	100	Adjusts the sensitivity of the object separation operation, a lower value will preserve larger objects with multiple intensity peaks
	Matchmaking Option	Not applicable		Toggles the tracking algorithm used for track-point matchmaking between time points; the available options are Greedy Matching and Hungarian Matching
	Track Lineage Option	Not applicable		Specifies whether to track cell divisions for quantifying lineages

Presets

There are two preset groups in the recipe: Detection and Tracking; each group has three pre-configured parameter groupings to help you get started on the analysis. The default preset values are as follows:

Detection

Parameter Name	Low	Medium	High
Background Removal Factor	75	75	75
Contrast Threshold Intensity Threshold	23 (8-bit)	8 (8-bit)	1 (8-bit)
Contrast Threshold Intensity Threshold	5,898 (16-bit)	1,966 (16-bit)	262 (16-bit)
Fill Holes Size	50	50	50
Smoothing Factor	4	4	4

Partition

Parameter Name	Small	Medium	Large
Object Size	10 - 500	50 - 3,000	100 - 6,000
Separation Factor	75	75	75
Max Search Range	20	20	20
Min Track Length	4	4	4
Matchmaking Option	Use Greedy Algorithm
Track Lineage Option	Track Lineage

Measurements

The Nuclei Tracking recipe generates morphological and intensity measurements for each detected cell as well as a count of the total number of cells on the image. You can add additional measurements to the analysis results by using the Measurement Tool in Aivia. The measurements generated by the recipe are as follows.

Morphological	Intensity	Track	Lineage
Area Circularity	Mean Total Standard Deviation	Total Time First Frame Last Frame X Y Acceleration Magnitude (instantaneous) Velocity Magnitude (instantaneous)	Track Counts Minimum Generation Maximum Generation Mean Generation

Tutorial

Before beginning the tutorial, please download the Nuclei Tracking Demo image. For information on how to select presets or modify parameter values, please refer to the tutorial on how to use the Recipe Console.

Unzip the demo file and load the demo image, NucleiTrackDemo.tif, into Aivia
In the Recipe Console, click on the Recipe selection dropdown menu and select the Nuclei Tracking recipe
Select the Low presets for both the Detection and Tracking preset groups
Click on the caret to the left of the Tracking preset group to show the preset parameters
Modify the parameter value as follows, leaving the other parameters intact:
- Separation Factor: 92
Click the Start button or press the F4 key on your keyboard to begin applying the recipe to the image

The detected objects outline and tracks will be overlaid on the image.

Results

Multimedia

name	NucleiTrackDemo_output.mp4
autostart	true

Nuclei Tracking results

Image credits

Harvard Stem Cell Institute, Harvard University, Boston MA

Versions Compared

Old Version 11

New Version 12

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