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  • Tandem Affinity Purification and Mass Spectrometry (TAP-MS) for the Analysis of Protein Complexes.

Tandem Affinity Purification and Mass Spectrometry (TAP-MS) for the Analysis of Protein Complexes.

Current protocols in protein science (2019-02-02)
Guillaume Adelmant, Brijesh K Garg, Maria Tavares, Joseph D Card, Jarrod A Marto
ABSTRACT

Affinity purification followed by mass spectrometry has become the technique of choice to identify binding partners in biochemical complexes isolated from a physiologic cellular context. In this report we detail our protocol for tandem affinity purification (TAP) primarily based on the use of the FLAG and HA peptide epitopes, with a particular emphasis on factors affecting yield and specificity, as well as steps to implement an automated version of the TAP procedure. © 2019 by John Wiley & Sons, Inc.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
DL-Dithiothreitol, ≥99.0% (RT)
Sigma-Aldrich
Phosphatase Inhibitor Cocktail 3, DMSO solution
Millipore
Anti-FLAG® M2 Magnetic Beads, affinity isolated antibody
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Acetic acid, glacial, ≥99.99% trace metals basis
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ANTI-FLAG® M2-Peroxidase (HRP) antibody, Mouse monoclonal, clone M2, purified immunoglobulin, buffered aqueous glycerol solution
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ANTI-FLAG® M2 Affinity Gel, purified immunoglobulin, buffered aqueous glycerol solution
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Anti-IL-2 Rα Antibody, clone 7G7/B6, clone 7G7/B6, Upstate®, from mouse