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The Exocytosis Detection recipe in Aivia detects exocytotic events in total internal reflection fluorescent (TIRF) microscopy images. The recipe provides an option for tracking vesicles associated with exocytosis. The recipe is tunable for the detection of small, point exocytosis events (e.g. kiss-and-run) as well as large, diffuse events (e.g. full fusion).

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Table of Contents

maxLevel	3
exclude	[4]

Parameters and Presets

Parameters

Recipe parameters for Exocytosis Detection and their descriptions are summarized in the table below.

Preset Group	Parameter Name	Min Value	Max Value	Description
Vesicle Detection	Vesicle Threshold (Vesicle Tracking only)	0	255	Adjusts the detection sensitivity for vesicle detection; a lower value will detect more vesicles
Vesicle Detection	Min Vesicle Size (Vesicle Tracking only)	0	100	Specifies the minimum size of a detected object to be included in the analysis results based on the area (in pixels) of the detected objects
Event Detection	Event Intensity Change	0	255 (8-bit) 65,535 (16-bit)	Adjusts the detection sensitivity for event detection based on the amount of intensity change (in number of gray levels) at the event from baseline; a lower value will detect more events
	Transience	0	100	Specifies the range of duration for the events by computing a normalized ratio for the difference between maximum and average intensities to the difference between the average and minimum intensities; a lower value will enable detection of events with longer durations
	Bright Detection Frame (Enable Bright Detection only)	0	255 (8-bit) 65,535 (16-bit)	Specifies the maximum number of frames for that an event with extended intensity increase in its surrounding regions may last; this parameter is used only when Enable Bright Detection is selected
	Duplicate Events Removal	Not applicable		Toggles the removal of detected events that occurs in a given region in successive time frames
Event Filter	Event Size Filter	0	1,000	Specifies the minimum size of a detected event to be included in the analysis results based on the area of the detected event
	Event Mean Intensity Filter	0	255 (8-bit) 65,535 (16-bit)	Specifies the minimum threshold for mean intensity of the detected event region to be included in the analysis results
	Event Total Intensity Filter (Enable Event Total Intensity Filter only)	0	1 x 10¹⁸	Specifies the minimum threshold for the total intensity of the detected event region to be included in the analysis results

Presets

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

Vesicle Detection

Parameter Name	Low	Medium	High
Vesicle Threshold	15	25	40
Min Vesicle Size	0.05	0.05	0.05

Event Detection

Parameter Name	Gross	Short	Long
Event Intensity Change	8 (8-bit) 1,966 (16-bit)	8 (8-bit) 1,966 (16-bit)	8 (8-bit) 1,966 (16-bit)
Transience	0 - 20	0 - 20	0 - 40
Bright Detection Frame	0	2	5
Duplicate Events Removal	Enable Duplicate Events Removal

Event Filter

Parameter Name	Small-Bright	Medium-Medium	Large-Dim
Event Size Filter	0.05	0.15	0.5
Event Mean Intensity Filter	19 (8-bit) 4,981 (16-bit)	12 (8-bit) 3,002 (16-bit)	5 (8-bit) 1,311 (16-bit)
Event Total Intensity Filter	15,000	45,000	50,000

Measurements

The Exocytosis Detection recipe generates intensity measurements for each detected event and, if Vesicle Tracking is enabled, for each detected particle plus track measurements for the particles. 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.

Measurement Type	Measurement Name	Events	Vesicle Tracks
Morphology	Area		x
Intensity	Mean	x	x
Intensity	Total	x	x
Track	Total Time		x
	First Frame		x
	Last Frame		x
	X		x
	Y		x
	Velocity Magnitude (instantaneous)		x
Miscellaneous	Additional measurements		Total Events Detected First Event Frame Has Relation (boolean) Events Detected on Track (boolean) Event on Frame (instaneous, boolean) Total Events in Image (Summary)

Tutorial

Before beginning the tutorial, please download the Exocytosis Detection 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, ExocytosisDemo.tif, into Aivia
In the Recipe Console, click on the Recipe selection dropdown menu and select the Exocytosis Detection recipe
Select the following preset for each preset group:
- Vesicle Detection: Medium
- Event Detection: Gross
- Event Filter: Large-Dim
Click on the caret Image Modified to the left of the Event Detection preset group to show the preset parameters
Modify the parameter value as follows, leaving the other parameters in tactintact:
- Event Intensity Change (counts): 1,000
Click the From beginningStart button or press the F4 key on your keyboard to begin applying the recipe to the image

The detected particle tracks and outline for the events will be overlaid on the image.

Results

Multimedia

name	ExocytosisDemo_output.mp4
autostart	true

Exocytosis Detection results

Image credits

Takashi Tsuboi, University of Tokyo, Tokyo, Japan

Versions Compared

Old Version 7

New Version 8

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