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Western Blotting

Western Blotting

Western blotting is an analytical technique for protein identification. It has various applications in immunology and medicine, including detecting diseases like bovine spongiform encephalopathy, mad cow disease, and human and feline immunodeficiency virus from biological samples.
The technique begins with separating proteins from the sample using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), followed by protein transfer, immunoblotting, and finally, protein detection.

Caspases

Caspases

Caspase, a family of cysteine proteases, serve as effectors in apoptosis. The ced3 gene in C.elegans was first identified to be involved in apoptosis. This gene encodes the ced-3 caspase that is similar to the interleukin-1-beta converting enzyme or ICE in mammals. In addition to apoptosis, caspases also function in the inflammatory response. Inflammatory caspases are essential in activating pro-inflammatory cytokines that recruit immune cells and block the replication of pathogens inside...

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Related Experiment Video

Updated: Apr 27, 2026

Evaluation of Caspase Activation to Assess Innate Immune Cell Death

Evaluation of Caspase Activation to Assess Innate Immune Cell Death

Published on: January 20, 2023

Detection of caspase activity using antibody-based techniques.

Gavin P McStay¹, Douglas R Green²

¹Department of Life Sciences, New York Institute of Technology, Old Westbury, New York 11568;

Cold Spring Harbor Protocols

|July 3, 2014

Summary

Antibodies targeting caspase cleavage fragments can identify activated executioner caspases in mammalian cells. This western blot protocol aids in determining caspase activation following apoptotic stimuli.

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Last Updated: Apr 27, 2026

Evaluation of Caspase Activation to Assess Innate Immune Cell Death

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Published on: January 20, 2023

Measuring Caspase Activity Using a Fluorometric Assay or Flow Cytometry

Measuring Caspase Activity Using a Fluorometric Assay or Flow Cytometry

Published on: March 24, 2023

Detecting Anastasis In Vivo by CaspaseTracker Biosensor

Detecting Anastasis In Vivo by CaspaseTracker Biosensor

Published on: February 1, 2018

Area of Science:

Molecular Biology
Cell Biology
Immunology

Background:

Caspases are key proteases in apoptosis, with specific antibodies available for mammalian models.
Antibodies can target either caspase pro-forms or their cleavage fragments.
Detecting activated executioner caspases is crucial for understanding apoptosis.

Purpose of the Study:

To present a protocol for identifying activated executioner caspases using fragment-specific antibodies.
To enable the determination of caspase activation extent and identity following apoptotic stimuli.

Main Methods:

Treatment of cultured cells with a proapoptotic stimulus.
Preparation of protein lysates from treated cells.
Western blot analysis using fragment-specific antibodies to detect cleaved executioner caspases.

Main Results:

Fragment-specific antibodies effectively identify activated executioner caspases.
Western blot analysis quantifies the extent of caspase activation.
This method aids in pinpointing which specific caspases are activated.

Conclusions:

Antibodies recognizing caspase cleavage fragments are valuable tools for apoptosis research.
The described western blot protocol provides a reliable method for assessing executioner caspase activation.
This approach facilitates the study of apoptotic pathways in mammalian systems.