QUBIC: a method for mapping protein–protein interactions by combination of BAC TransgeneOmics and quantitative MS. (A and B) Two optimized AP-MS approaches of QUBIC are shown using either SILAC (A) or label-free (B) protein quantitation. (A) In SILAC experiments, the WT cell line without a BAC transgene is cultured in a medium containing the C¹²N¹⁴ form of lysine, and the tagged cell line is cultured in a medium containing the C¹³N¹⁵ form of lysine. Separate pull-downs using magnetic beads coupled to anti-GFP antibody are performed, and elutes merged directly after elution by in-column digestion. Peptides are identified by high resolution LC-MS/MS and quantified by directly comparing relative intensities of the light and heavy forms of each peptide present in the mass spectrum. Specific interaction partners show high H/L ratios, whereas background binders have a ratio of 1. (B) In label-free experiments, tagged and control cells are cultured in normal media, and separate pull-downs are performed. Eluates are not mixed but analyzed separately by LC-MS/MS. Proteins are quantified with the label-free algorithm in MaxQuant software.