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Subcellular Fractionation

Subcellular Fractionation

The homogenate obtained after cell lysis contains various membrane-bound organelles that can be further separated into pure fractions by subcellular fractionation. These isolates are used to study specific cellular components, analyze localized protein activity, and are even employed in diagnostics. Fractionation is typically achieved using centrifugation methods, the most common being density-gradient and differential centrifugation.
Differential Centrifugation
Differential centrifugation is...

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Related Experiment Video

Updated: Dec 7, 2025

Substructure Analyzer: A User-Friendly Workflow for Rapid Exploration and Accurate Analysis of Cellular Bodies in Fluorescence Microscopy Images

Substructure Analyzer: A User-Friendly Workflow for Rapid Exploration and Accurate Analysis of Cellular Bodies in Fluorescence Microscopy Images

Published on: July 15, 2020

Quantitative analysis of subcellular distributions with an open-source, object-based tool.

Pearl V Ryder¹, Dorothy A Lerit¹

¹Department of Cell Biology, Emory University School of Medicine, Atlanta, GA 30322, USA dlerit@emory.edu ryderp@gmail.com.

|September 25, 2020

Summary

This summary is machine-generated.

A new computational tool, SubcellularDistribution pipeline, automates 3D microscopy image analysis for subcellular localization. It accurately detects RNA distribution relative to centrosomes in Drosophila embryos.

Keywords:

Centrosome Colocalization Fluorescence microscopy Image analysis RNA localization

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Area of Science:

Cell Biology
Computational Biology
Genetics

Background:

Subcellular localization of cellular components is crucial for cell function.
Colocalization analysis of microscopy images is vital for understanding spatial relationships and biological mechanisms.
Automated tools are needed to streamline complex image analysis.

Purpose of the Study:

To develop an automated and customizable computational tool, the SubcellularDistribution pipeline, for object-based image analysis of 3D fluorescence microscopy data.
To assess the utility of the SubcellularDistribution pipeline by examining mRNA distribution relative to centrosomes in Drosophila embryos.
To provide an open-source software solution for efficient and accurate subcellular localization studies.

Main Methods:

Generation of the SubcellularDistribution pipeline, an automated computational tool for image analysis.
Application of the pipeline to analyze three-dimensional (3D) fluorescence microscopy images of syncytial Drosophila embryos.
Comparison of the pipeline's RNA distribution detection with commercially available software.

Main Results:

The SubcellularDistribution pipeline successfully automated the analysis of subcellular localization from 3D microscopy images.
The software accurately detected RNA distributions relative to centrosomes in Drosophila embryos, comparable to commercial software.
The pipeline guides users through image segmentation, data processing, and visualization for publication-ready data.

Conclusions:

The SubcellularDistribution pipeline is an effective, open-source tool for analyzing subcellular object localization in 3D microscopy.
This tool facilitates the study of spatial relationships, such as RNA enrichment at centrosomes, in biological systems.
The software enhances the efficiency and accessibility of complex image analysis for biological research.