Proteomics has revolutionized the field of biomarker discovery, offering unprecedented insights into cellular processes and disease mechanisms. However, as mass spectrometry (MS) technology advances, the bottleneck has shifted to the pre-analytical sample preparation phase. Evosep’s innovative solution addresses this challenge head-on with a fully automated, scalable workflow leveraging the Evotip platform. In collaboration with the Olsen group at University of Copenhagen, this has been evaluated and the findings detailed in the recent publication in Molecular & Cellular Proteomics.


Figure: Workflow of complete automated sample preparation for scalable clinical proteomics with Evotip.


Revolutionizing Sample Preparation

Traditional sample preparation in proteomics involves multiple manual steps that are time-consuming, prone to variability, and costly. The new workflow developed by Evosep integrates sample digestion, cleanup, and loading onto Evotips in a fully automated manner using the Opentrons OT-2 robot. This approach not only streamlines the process but also significantly enhances throughput and reproducibility.

The key to this efficiency lies in the Evotip, that facilitates the direct loading of peptide samples onto the MS. By incorporating this with the OT-2 robot, up to 192 samples can be processed in just 6 hours, a dramatic improvement over traditional methods. This advancement is particularly beneficial for large-scale clinical studies, where hundreds of samples need to be analyzed quickly and consistently.


High Sensitivity and Reproducibility

The automated workflow has been rigorously tested with HeLa cell samples, yielding impressive results. With a protein input of just 1 microgram, the workflow identified approximately 8,000 protein groups and 130,000 peptide precursors using the 100 SPD method in combination with the Orbitral Astral mass spectrometer. This level of sensitivity and reproducibility is crucial for detecting subtle changes in protein expression, which can be indicative of disease states or therapeutic responses.

Furthermore, the workflow demonstrated remarkable stability, with minimal sample degradation even after several days of storage. This flexibility allows for the seamless integration of sample preparation with subsequent MS analysis, reducing downtime and maximizing the use of instrumentation.


Expanding Capabilities with Phosphoproteomics

In addition to proteome analysis, the workflow can be extended to phosphoproteomics, a technique essential for studying cellular signaling pathways. By integrating an automated phosphopeptide enrichment step using magnetic Ti-IMAC beads, the system can process both proteomes and phosphoproteomes in parallel. This dual capability opens new avenues for investigating post-translational modifications, providing deeper insights into cellular dynamics and disease mechanisms.


Clinical Application in Cancer Research

The practical utility of this workflow is exemplified in its application to a clinical cohort of metastatic melanoma patients undergoing immune therapy. By analyzing plasma samples before and after treatment, the study identified several potential biomarkers associated with therapeutic response. These findings underscore the potential of Evosep’s automated workflow to facilitate large-scale biomarker discovery and personalized medicine. For instance, the study revealed significant changes in specific proteins, such as increases in liver and acute phase proteins and decreases in apolipoproteins in non-responding patients. These biomarkers can provide valuable information for predicting patient outcomes and tailoring treatment strategies.



Evosep’s automated workflow, based on the Evotip platform, represents a significant leap forward in clinical proteomics. By combining high-throughput capability, sensitivity, and reproducibility, it addresses the critical bottleneck in sample preparation. This scalable solution not only enhances the efficiency of biomarker discovery but also paves the way for more robust and cost-effective clinical proteomics studies.

As proteomics continues to evolve, innovations like these will be instrumental in translating complex biological data into actionable insights, ultimately improving patient care and advancing our understanding of disease. Evosep’s commitment to developing cutting-edge solutions ensures that researchers can keep pace with the ever-increasing demands of clinical proteomics.


The Evosep One instrument is for Research Use Only.

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