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Antibody-Based Proteomic Analysis of Apoptosis Signaling

Protocol
Part of the Methods in Pharmacology and Toxicology book series (MIPT)

Abstract

Reagents that assess activation of apoptosis and associated signaling pathways are critical for greater understanding of the molecular basis of programmed cell death. The advent of proteomic technologies to probe these events allows monitoring of hundreds to thousands of proteins, as well as sites of posttranslational modification involved in apoptosis at one time. This view of apoptosis at a network level is a powerful tool in studying known apoptotic pathways, as well as elucidating novel signaling events that affect or are affected by apoptotic signaling. The following is a detailed method for successful proteomic profiling of apoptosis using antibody-based enrichment methods along with a liquid chromatography–tandem mass spectrometry analytical platform.

Key words

Apoptosis Proteomics Posttranslational modification Antibody LC-MS/MS Phosphorylation Caspase 

Abbreviations

IAP

Immunoaffinity purification

LC

Liquid chromatography

MS/MS

Tandem mass spectrometry

MeCN

Acetonitrile

MS

Mass spectrometry

PTM

Posttranslational modification

TFA

Trifluoroacetic acid

Supplementary material

313506_1_En_9_MOESM1_ESM.xlsx (610 kb)
Supplemental Table S1 (XLSX 609 kb)

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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  1. 1.Cell Signaling Technology, Inc.DanversUSA

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