Updated project metadata.
Isobaric labeling has the promise of combining high sample multiplexing with precise quantification. However, normalization issues and the missing value problem of complete n-plexes hamper quantification across more than one n-plex. Here we applied two novel algorithms implemented in MaxQuant that substantially improve the data analysis with multiple n-plexes. First, isobaric matching between runs (IMBR) makes use of the three-dimensional MS1 features to transfer identifications from identified to unidentified MS/MS spectra between LC-MS runs in order to utilize reporter ion intensities in unidentified spectra for quantification. On typical datasets, we observe a significant gain in quantifiable n-plexes MS/MS spectra that can be used for quantification. Second, we introduce a novel PSM-level normalization, applicable to data with and without common reference channel. It is a weighted median-based method, in which the weights reflect the number of ions that were used for fragmentation. This dataset is TMT 8-plex samples without any referene channels.