Tomogram segmentation¶

Closed surfaces¶

nextPYP can segment closed surfaces (such as virions or spherical vescicles) using an energy-based algorithm described in Bartesaghi et al. (2005). Segmentation proceeds in two steps:

Find centers¶

The first step is to locate a marker inside each virion or vescicle, which can be done using any of the particle picking methods implemented in nextPYP:

• Click on Tomograms (output of the Pre-processing block) and select Particle Picking

• Select the desired particle picking algorithm and corresponding parameters, see Particle picking. The “virion” picking method is especially designed to find the center of spherical virions and estimate their radius (which will be useful later)

• Click Save, Run, and Start Run for 1 block

• Navigate to the Particles tab to inspect the results of the detection

Segment surfaces¶

Once the location of each virion or vescicle center has been determined, the segmentation can be calculated:

• Click on Particles (output of the Particle-Picking block) and select Segmentation (closed surfaces)

• Set the desired Segmentation radius tolerance

• Click Save, Run, and Start Run for 1 block

• Navigate to the Segmentation (closed surfaces) block and select the Segmentation tab to inspect the results

nextPYP calculates implicit representations of surfaces and uses a single threshold to determine the location of the membranes

• (optional) In cases where the default value for the threshold gives innacurate results, users can select a different threshold by selecting a virion from the table to show its 3D slices and the segmentation thresholds (8 different thresholds are shown as yellow contours in columns 1-8). The highlighted column number represents the current threshold selection (default is 1, click on a different column to select a better threshold). If none of the columns look reasonable (or if you want to ignore the current virion), select the last column labeled as “-”. Repeat this process for all virions in the tilt-series and all tilt-series in the dataset

Open surfaces¶

nextPYP can also segment open surfaces using the package MemBrain-Seg:

• Click on Tomograms (output of the Pre-processing block) and select Segmentation (open surfaces)

• Select the desired parameters for MemBrain-Seg, including the location of a pre-trained model (*.ckpt) downloadable from their Github repository

• Click Save, Run, and Start Run for 1 block

• Navigate to the Segmentation (open surfaces) block to inspect the results of the segmentation