ZEISS Knowledge Base

Introduction

This guide explains you how to detect big structures manual using the pipeline with the Intensity Threshold Segmenter operation. The purpose of the pipeline is to detect objects having a random shape and not well-defined borders. It can be applied to any cellular compartments or biological structures.

Activating the Sample Pipeline

In the Shortcut Toolbar, click Analysis Panel .
In the Sample Pipelines list, double-click the Detect Big Structures Manual Pipeline.
If you have activated a pipeline, it will be replaced by the new one.

You can open the appropriate How to guide. By hovering over a pipeline, this button is displayed. When clicking on it, the option Open How to appears.

The following operations are part of the pipeline.

This operation allows to select the region of interest (ROI). ROI defines the dataset subarea that will be processed and analyzed by the pipeline.

Parameter		Description
ROI		Sets the processing and analysis target space.
–	Current View	The selected Z plane and the viewer area are processed.
–	Current Plane	The selected Z plane is processed (XY).
–	Current Time Point	The selected time point is processed (XYZ).
–	Current Image Set	The complete dataset (XYZ and time) is processed.
–	Custom	Allows to mix the previous methods. Note: Use the Custom option during the pipeline setting and testing. Set a sub volume (XY, Planes, Time Points, channels) of your dataset on which perform the trial. This will speed up the setting process.
Channels		Sets the processing and analysis target channels. Selecting a single channel, all the operations in the pipeline will be forced to use it.
Scaling		Sets the scaling of the dataset, which reduces it size. The measurements will not be modified by the scaling factor
Crop input data		Sets the limitation of the data used for the calculation to only the cropped selection defined above. If this option is not selected, the entire image set is used for the calculation.

Additional Parameters

Additional settings for Input ROI expand, when you click in the operation.

Parameter	Description
Bounds	Sets the analysis area edges. The whole XY bounds, the viewing area or a custom space can be applied.
Planes	Sets the analysis planes range. A single plane, a range of planes or the whole stack can be selected.
Time Points	Sets the analysis time points range. A single time pint, a range of time points or the whole movie can be selected.

Intensity Threshold Segmenter

Allows the segments detection using one of the available automatic threshold methods.

Intensity Threshold Segmenter parameters

By default, the Method: Range is selected. Depending on the selection, different parameters are available.

Parameter		Description
Channel		Sets the processing and analysis target channel.
Method		Sets the threshold algorithm. By default, Auto is selected.
–	Simple	This method uses a single threshold and segments everything below or above it.
–	Range	This method uses two Thresholds to define the range of intensities to segment. Everything outside of this range is neglected.
–	Percentile	This method uses a single threshold and segments all intensities below or above it. The threshold is given as a percentile and is based on the data.
–	Percentile range	This method uses two thresholds to define the range of intensities to segment. Everything outside of this range is neglected. The thresholds are given as percentiles and are based on the data.
–	Auto	This method uses a single threshold and segments all intensities below or above it. The threshold is automatically calculated based on the data.
–	Adaptive mean	This method uses local information around each pixel to set a threshold in relation to the local background. It is based on Niblack's thresholding method.
–	Mean	This method uses local information around each pixel to set a threshold in relation to the local background.
Threshold		Sets the threshold values accordingly to the image dynamic range.

Store Objects

Store the detected segments (tag) in the active dataset.

Executing the pipeline

You can execute the pipeline step by step or in a single run. To do this, use the executing buttons in the Pipeline toolbar.

Executing step by step

You can execute the pipeline step by step (back and forth). This method allows to run and undo a single operation. You can either use the executing buttons in the Pipeline toolbar or in the Operation toolbar to go through the operation list.

To run the single operation, click .
To undo the single operation, click .
Note: Undo the last operation executed if you need to change the operation settings.

Executing in a single run

As alternative to executing step by step, execute the pipeline in a single run.

To run the whole pipeline, click .
To stop the pipeline execution, click .

Modifying the current pipeline

You can modify the pipeline to adapt to another datasets. Therefore, all the pipeline parameters should be set according to the new dataset features.

Adjusting the operations

The parameters of each operation are described here: Pipeline operations layout

You have opened the pipeline.

Go to the operation you want to change.
Set the appropriate parameters as desired.
Execute the pipeline (see Executing the pipeline).

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Detecting big structures manually

Introduction

Workflow