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The growing interest for high-throughput proteomics requires standardization and simplified workflows. This is enhanced by automation to remove human errors and decrease data variation. The Opentrons OT-2 (OT-2) liquid handling robot is an excellent example of a low-cost and open-source platform that easily enables the development and integration of end-to-end workflows. Other liquid handling robots are often expensive, and their proprietary design limits the integration of specific applications. As a step towards end-to-end workflows, we have developed a simple and automation friendly protocol for sample loading on Evotips, which makes use of a layered sandwich approach with defined airgaps between the layers (Figure 1). This is then pushed through the Evotip with the OT-2 pipette for 100 seconds leaving the Evotip ready for injection on the Evosep One….