xmipp3.protocols.protocol_particle_pick_pairs module
- class xmipp3.protocols.protocol_particle_pick_pairs.XmippProtParticlePickingPairs(**args)[source]
Bases:
ProtParticlePicking,XmippProtocolPicks particles in paired tilted micrographs. Using paired data improves particle localization and orientation determination, enhancing reconstruction accuracy.
AI Generated
## Overview
The Tilt Pairs Particle Picking protocol allows the user to pick corresponding particles in paired tilted and untilted micrographs.
Tilt-pair data contain two related images of the same field: one acquired without tilt, or with lower tilt, and another acquired at a known tilt angle. The same particles appear in both micrographs, but their positions are related by the geometry of the tilt. Correctly identifying corresponding particles in both views is essential for workflows such as random conical tilt, tilted-pair validation, and geometry-aware reconstruction strategies.
This protocol launches the Xmipp tilt-pair particle-picking graphical interface. The user can inspect the paired micrographs and define particle coordinates in a way that preserves the relationship between untilted and tilted views.
The main output is a CoordinatesTiltPair object, containing linked coordinate sets for the untilted and tilted micrographs, together with the tilt-angle information read from the paired micrograph metadata.
## Inputs and General Workflow
The input is a set of paired tilted micrographs, represented in Scipion as a MicrographsTiltPair object.
The protocol first converts the paired micrograph information into Xmipp metadata format. This metadata describes the relationship between the untilted and tilted micrograph sets and is used by the graphical picker.
The protocol then opens the tilt-pair picking interface. After the user has picked particles and saved the results, the protocol reads the coordinate files for both the untilted and tilted micrographs. It also reads the tilt-pair angle information and creates the final paired-coordinate output.
## Micrographs Tilt Pair
The Micrographs tilt pair parameter defines the paired micrograph dataset to be used for picking.
This input contains two linked micrograph sets:
the untilted micrographs;
the tilted micrographs.
Each untilted micrograph should correspond to a tilted micrograph showing the same field of view under the tilt geometry. The quality of the output depends on this pairing being correct.
If the micrograph pairs are mismatched, the picked coordinates will not represent the same physical particles in both views, and downstream tilted-pair analysis will be unreliable.
## Tilt-Pair Picking Interface
The protocol launches a graphical interface designed specifically for tilt-pair picking.
Unlike ordinary single-micrograph picking, the goal here is not only to select particle positions, but also to preserve their correspondence between the two views. The picker uses the paired micrograph metadata to help manage this relationship.
The user can inspect the untilted and tilted images, pick particles, and save coordinate information for both members of each pair.
This interactive step is important because tilted images may be harder to interpret than untilted images. Particles can appear distorted, shifted, or less contrasted because of the tilt geometry and increased effective ice thickness.
## Untilited and Tilted Coordinate Sets
After picking, the protocol creates two coordinate sets:
one coordinate set for the untilted micrographs;
one coordinate set for the tilted micrographs.
These coordinate sets are not independent outputs. They are combined into a single CoordinatesTiltPair object that stores their relationship.
This paired organization is essential. It tells downstream protocols which untilted coordinate corresponds to which tilted coordinate.
## Tilt Angles
The protocol also reads the angle information associated with the input micrograph pairs.
These angles describe the tilt geometry of the paired acquisition. They are stored together with the coordinate pairs and are needed by downstream tilted-pair processing protocols.
The angle information is not manually estimated by this protocol. It is read from the paired micrograph metadata generated from the input MicrographsTiltPair object.
Users should therefore ensure that the tilt-pair micrograph input contains correct tilt information.
## Output CoordinatesTiltPair
The main output is an outputCoordinatesTiltPair object.
This output contains:
the untilted coordinate set;
the tilted coordinate set;
the angle information linking the paired micrographs;
the box-size information when available.
This object is the standard Scipion representation of picked tilt-pair coordinates. It can be used by protocols that extract paired particles or perform tilted-pair analysis.
If the protocol is run several times and new coordinate outputs are registered, the outputs may receive numbered suffixes to avoid overwriting previous results.
## Particle Size and Box Size
The protocol can store a particle box size in the output coordinate sets when available.
The box size represents the approximate particle size used during picking and can guide later extraction of untilted and tilted particle pairs.
For tilted-pair data, the box size should be large enough to contain the particle in both views. Tilted projections may appear slightly elongated, distorted, or less well centered, so users should be careful not to choose a box that is too small.
## Interpretation of the Output
The output should be interpreted as a set of paired particle positions.
Each pair represents one physical particle observed in two acquisition geometries. The untilted coordinate indicates its position in the untilted micrograph, and the tilted coordinate indicates the corresponding position in the tilted micrograph.
This correspondence is more important than either coordinate set alone. Downstream analysis depends on the assumption that each pair refers to the same particle.
## Practical Recommendations
Use this protocol only with correctly paired tilted and untilted micrographs.
Before picking, inspect the micrograph pairs to verify that they correspond to the same field of view and that the tilt geometry is sensible.
Pick particles that can be reliably identified in both views. Avoid ambiguous particles, overlapping particles, strong contamination, carbon edges, and areas where the tilted view is too degraded to identify the same particle.
Choose a particle size or extraction box large enough for the tilted view, not only for the untilted view.
After picking, inspect the paired coordinates before extraction. Errors in pairing can seriously affect downstream tilted-pair workflows.
If several picking sessions are needed, keep track of the different output coordinate-pair objects generated by the protocol.
## Final Perspective
Tilt Pairs Particle Picking is the coordinate-generation step for tilted-pair cryo-EM workflows.
For biological users, its main role is to define which particles correspond between untilted and tilted micrographs. This correspondence is the foundation for subsequent extraction and analysis of particle pairs.
Good tilted-pair picking requires both accurate particle identification and careful preservation of the pairing relationship. When done correctly, it provides the geometrical information needed for downstream protocols that use tilt-pair data.