Single Particle Em
Single Particle Em - Related Videos
09:06
Cryo-EM and Single-Particle Analysis with Scipion
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Cited by 8 •
2021
Single-particle analysis in cryo-electron microscopy is one of the main techniques used to determine the structure of biological ensembles at high resolution. Scipion provides the tools to create the whole pipeline to process the information acquired by the microscope and achieve a 3D reconstruction of the biological specimen.
04:52
Single-Particle Cryo-EM Data Collection with Stage Tilt using Leginon
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Cited by 6 •
2022
The present protocol describes a generalized and easy-to-implement scheme for tilted single-particle data collection in cryo-EM experiments. Such a procedure is especially useful for obtaining a high-quality EM map for samples suffering from preferential orientation bias due to adherence to the air-water interface.
10:13
A Microfluidic-based Hydrodynamic Trap for Single Particles
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Cited by 14 •
2011
In this article, we present a microfluidic-based method for particle confinement based on hydrodynamic flow. We demonstrate stable particle trapping at a fluid stagnation point using a feedback control mechanism, thereby enabling confinement and micromanipulation of arbitrary particles in an integrated microdevice.
13:28
High-resolution Single Particle Analysis from Electron Cryo-microscopy Images Using SPHIRE
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Cited by 218 •
2017
This paper presents a protocol for processing cryo-EM images using the software suite SPHIRE. The present protocol can be applied for nearly all single particle EM projects that target near-atomic resolution.
13:43
A Robust Single-Particle Cryo-Electron Microscopy (cryo-EM) Processing Workflow with cryoSPARC, RELION, and Scipion
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Cited by 17 •
2022
This article describes how to effectively utilize three cryo-EM processing platforms, i.e., cryoSPARC v3, RELION-3, and Scipion 3, to create a single and robust workflow applicable to a variety of single-particle data sets for high-resolution structure determination.
FAQs
Computational alignment places particle images into a common orientation, while classification groups images that share similar structural features. Averaging the aligned images strengthens consistent signal and reduces the influence of image noise. This processing converts many two-dimensional views into a three-dimensional density map that can support interpretation of molecular architecture and structural differences.
Rapid immobilization preserves purified biological particles in a vitrified, glass-like form rather than allowing structural organization into crystals. The particles can then be imaged while retained in the frozen state, helping capture their molecular shapes for analysis from multiple orientations. This condition is central to examining individual proteins and complexes in a cryogenic workflow.
Particle images can be classified according to differences in their observed structural features. Distinct classes may correspond to alternative conformational states, allowing researchers to compare how the same complex adopts different shapes. These structural differences can help connect molecular architecture with biological function and provide context for mechanistic studies of proteins and larger molecular assemblies.
Single-particle EM analyzes separate biological particles rather than requiring them to form an ordered crystal. This distinction allows structural investigation of proteins and molecular complexes that are examined in an individual-particle workflow. By combining images from many particle orientations, the method can produce a three-dimensional map while retaining access to structural variation among particles.
A typical workflow begins with purified particles, followed by rapid immobilization in vitreous ice. The frozen sample is imaged from many orientations with an electron beam. Computational analysis then aligns and classifies the resulting particle images before averaging them to reconstruct a three-dimensional density map for biological interpretation.
The resulting three-dimensional density map can show overall protein architecture, ligand-binding sites, and distinct conformational states. These outputs give biologists structural evidence for examining how molecular shape relates to function. The information is useful when interpreting the organization of individual proteins or complexes and when investigating changes associated with different structural states.
The technique is useful for structural biology and mechanistic studies because it connects molecular shape with biological function. Its maps can also identify ligand-binding sites, making the method relevant to rational drug research. Researchers can use these structural insights to examine how biological particles are organized and to investigate molecular features related to potential ligand interactions.