Immunoprecipitation isolates a protein of interest and its interactors from a complex protein mixture. Combined with TMT labeling and LC-MS, it gives an accurate, quantitative measure of that protein and its network, compared across several conditions.
At a glance
- Quantification from as few as 5 million cells or 20 µg of protein
- Up to 18 samples quantified together with no missing values (TMT 18-plex)
- Suited to quantifying a protein of interest, mapping protein–protein interactions and active site mapping
How it works
Immunoprecipitated samples are alkylated and digested with trypsin to obtain peptides from the protein of interest and its partners. The peptides are labeled with TMT so the samples can be combined into a single multiplexed set. The mixture is fractionated and analyzed by LC-MS, and the results are scored against interaction databases to produce a map of interactors.
Example data: cell-surface microenvironments
Immunoprecipitating three different cell-surface antigens, CD47, CD45 and CD29, and quantifying the proteins enriched with each revealed a distinct cell-surface microenvironment around each antigen.
Spotlight: shared and distinct partners
CD87 sits in a microenvironment shared by CD45 and CD47, while CD29 differs between them. Protein-level and peptide-level measurements agree, with close agreement between replicate experiments (n=3).
