Phosphoproteomics

Posttranslational modifications of proteins are relevant for a large variety of cellular functions, with phosphorylation being one of the most frequent and relevant ones in many signal transduction pathways.
We constantly optimize protocols to specifically enrich and analyze phosphorylated peptides from tissue or cell lysates. We managed to significantly reduce the amount of sample material from several mg per sample down to 50 – 100 µg of sample input. Automation of protocols will further decrease necessary input to the low µg range.
Samples are measured in data independent acquisition (DIA) mode followed by label-free quantification and peptide centric statistical analysis. In every experiment, we deliver total proteome results in addition to the phosphorylation-enriched peptide fraction. Data are generated from the same sample to enable normalization and evaluation of changes on the phosphorylation level in relation to changes observed in the total proteome. Meaningful phosphoproteome profiling requires prior experimental knowledge about the dynamics (chronic or transient effects on phosphorylation) and the timing of the respective stimulation (such as time course experiments).