In-gel stable isotope labeling for relative quantification using mass spectrometry.

In-gel stable isotope labeling for relative quantification using mass spectrometry. Research Abstract Details 

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In-gel stable isotope labeling for relative quantification using mass spectrometry. Abstract Text:

John M Asara,Xiang Zhang,Bin Zheng,Lisa A Maroney,Heather R Christofk,Ning Wu,Lewis C Cantley,John M Asara,Xiang Zhang,Bin Zheng,Lisa A Maroney,Heather R Christofk,Ning Wu,Lewis C Cantley,

Although differences in protein staining intensity can often be visualized by difference gel electrophoresis, abundant proteins can obscure less abundant proteins, and quantification of post-translational modifications is difficult. We present a protocol for quantifying changes in the abundance of a specific protein or changes in specific modifications of a protein using in-gel stable isotope labeling. In this protocol protein extracts from any source treated under two experimental conditions are resolved in two separate lanes by gel electrophoresis. Parallel gel regions of interest are reacted separately with either light or heavy isotope-labeled reagents, and the gel slices are then combined and digested with proteases. The resulting peptides are then analyzed by liquid chromatography/mass spectrometry (LC/MS) to determine relative abundance of light- and heavy-isotope lysine-containing peptide pairs and analyzed by LC/MS/MS for identification of sequence and modifications. This protocol should take approximately 24-26 h to complete, including the incubation time for proteolytic digestion. Additional time will be needed for data analysis and interpretation.

In-gel stable isotope labeling for relative quantification using mass spectrometry. Publishing Authors By Initials

JM Asara,X Zhang,B Zheng,LA Maroney,HR Christofk,N Wu,LC Cantley,JM Asara,X Zhang,B Zheng,LA Maroney,HR Christofk,N Wu,LC Cantley,

For similar genetic processes: gene expression regulation: protein modification, translational: protein processing, post-translational research abstracts see: genetic processes: gene expression regulation: protein modification, translational: protein processing, post-translational research

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In-gel stable isotope labeling for relative quantification using mass spectrometry. Journal Published:

PUBLICATION TYPE: Research Support, Non-U.S. Gov

Journal: Nature protocols

VOLUME: 1

Page Numbers: 46-51

Journal Abbreviation:

ISSN: 1750-2799

DAY: 3

MONTH: 12

YEAR: 2006

In-gel stable isotope labeling for relative quantification using mass spectrometry. Information

Number of References:

LANGUAGE: eng

NlmUniqueID: 101284307

In-gel stable isotope labeling for relative quantification using mass spectrometry. Keywords Mesh Terms:

KEYWORDS: Protein Processing, Post-Translational

MESH TERMS: analysis

Chemical & Substance for Abstract: In-gel stable isotope labeling for relative quantification using mass spectrometry. Information

Substance Name: Peptides

Registry Number: 0

Grant and Affiliation Information for In-gel stable isotope labeling for relative quantification using mass spectrometry.

AFFILIATION: Division of Signal Transduction, Beth Israel Deaconess Medical Center, Boston, Massachusetts 02115, USA. jasara@bidmc.harvard.edu

Country: England

AGENCY: United States NIGMS

GRANT: GM56203

ACRONYM: GM

MEDLINETA: Nat Protoc

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