High-Throughput Proteomics Sample Preparation Using a 96-Channel Pipettor and Magnetic Pin Device
Georgia Roumelioti, Alex Montoya, Gemma L. M. Fisher, Eneko Pascual Navarro, Angela Woods, Jane Bennett, Naveenan Navaratnam, Oliver Gonzalez-Carvajal, Jodie Birch, Elizabeth Pyman, Sijia Yu, Aleksandra Gruevska, Luc-Alban Vuillemenot, Oleh Lushchak, Zoe Hall, Alexis R. Barr
TL;DR
This paper introduces a cost-effective, semi-automated workflow for high-throughput phosphoproteomics using 96-channel devices, reducing workload and variability.
Contribution
A practical, semi-automated phosphoproteomics workflow using 96-channel devices with cost-efficient solid-phase extraction plates and simplified digestion.
Findings
A 96-well phosphoproteomics workflow was completed in 2 days with minimal operator workload.
Custom solid-phase extraction plates using Oasis HLB sorbent were characterized for loading capacity and lipid removal.
PAC digestion can be simplified by aspirating beads without requiring continuous shaking.
Abstract
High-throughput proteomics requires efficient and highly reproducible sample processing, yet workflowsparticularly for PTM profilingremain complex and costly to fully automate. Here, we present a practical intermediate solution using manually operated 96-channel devices: the Gilson Platemaster P220 pipettor and VP Scientific 96-well magnetic pin device. Using this setup, we achieved robust and reproducible phosphoproteomics in a 96-well format, completing protein aggregation capture (PAC/SP3) digestion, desalting, phosphopeptide enrichment, and a second desalting step within 2 days while minimizing operator workload and variability. Several innovations enable this workflow. First, we describe a cost-efficient method to generate 96-well solid-phase extraction plates by directly packing the Oasis HLB sorbent into tapered filter plates. We extensively characterize these plates in terms…
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Taxonomy
TopicsAdvanced Proteomics Techniques and Applications · Advanced Biosensing Techniques and Applications · Diatoms and Algae Research