Targeted proteomics analysis of plasma proteins using recombinant protein standards for addition only workflows

Targeted proteomics is an attractive approach for the analysis of blood proteins. Here, we describe a novel analytical platform based on isotope-labeled recombinant protein standards stored in a chaotropic agent and subsequently dried down to allow storage at ambient temperature. This enables a straightforward protocol suitable for robotic workstations. Plasma samples to be analyzed are simply added to the dried pellet followed by enzymatic treatment and mass spectrometry analysis. Here, we show that this approach can be used to precisely (coefficient of variation <10%) determine the absolute concentrations in human plasma of hundred clinically relevant protein targets, spanning four orders of magnitude, using simultaneous analysis of 292 peptides. The use of this next-generation analytical platform for high-throughput clinical proteome profiling is discussed. METHOD SUMMARY Here, we describe a novel approach suitable for routine multiplex measurements of plasma proteins using a mass spectrometry-based protocol, enabled by the use of a chaotropic agent in combination with stable isotope-labeled recombinant protein standards. The method combines established technologies, such as stable isotope methods, generation of recombinant protein standards and mass spectrometry analysis using data independent acquisition with novel use of a chaotropic agent in combination with a vacuum-dried format.

Automated summary

This novel approach utilizes isotope-labeled recombinant protein standards stored in a chaotropic agent, allowing storage at ambient temperature, for high-throughput clinical proteome profiling.