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Identification of Protein Interaction Partners in Mammalian Cells Using SILAC-immunoprecipitation Quantitative Proteomics
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Quantitative Comparison of Proteomes Using SILAC.

Jingjing Deng1, Hediye Erdjument-Bromage1, Thomas A Neubert1

  • 1Kimmel Center for Biology and Medicine at the Skirball Institute and Department of Cell Biology, New York University School of Medicine, New York, New York.

Current Protocols in Protein Science
|September 22, 2018
PubMed
Summary
This summary is machine-generated.

Stable isotope labeling by amino acids in cell culture (SILAC) is a popular quantitative proteomic method. Protocols are described for basic, triplex, and newly synthesized protein measurements using this metabolic labeling strategy.

Keywords:
BONLACSILACprotein synthesis and proteomics

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Area of Science:

  • Proteomics
  • Cell Biology
  • Biochemistry

Background:

  • Stable isotope labeling by amino acids in cell culture (SILAC) is a widely adopted quantitative proteomic technique.
  • Introduced in 2002, SILAC relies on metabolic incorporation of isotope-labeled amino acids into proteins during translation.
  • This method allows for precise quantification by mixing differentially labeled samples early in the workflow.

Purpose of the Study:

  • To provide detailed protocols for implementing SILAC.
  • To describe variations of SILAC for different experimental needs, including non-dividing cells.
  • To outline methods for quantifying newly synthesized proteins using SILAC.

Main Methods:

  • Metabolic labeling of cells with stable isotope-enriched amino acids.
  • Differential labeling of samples followed by immediate mixing post-lysis.
  • Application of protocols for basic duplex SILAC, triplex SILAC, and newly synthesized protein quantification.

Main Results:

  • Established protocols for SILAC enable accurate quantitative proteomic analysis.
  • The described methods are applicable to various cell types, including non-dividing cells like neurons.
  • SILAC effectively facilitates the measurement of newly synthesized proteins.

Conclusions:

  • SILAC is a robust and versatile method for quantitative proteomics.
  • The provided protocols offer practical guidance for researchers utilizing SILAC.
  • This technique minimizes errors and enhances the reliability of proteomic data.