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. 2021 Mar 11;12:644725. doi: 10.3389/fimmu.2021.644725

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Copyright © 2021 Metzemaekers, Abouelasrar Salama, Vandooren, Mortier, Janssens, Vandendriessche, Ganseman, Martens, Gouwy, Neerinckx, Verschueren, De Somer, Wouters, Struyf, Opdenakker, Van Damme and Proost.

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Workflow for immunosorbent tandem mass spectrometry proteoform analysis (ISTAMPA). (1) Synovial fluids are collected from the inflamed joints of patients with arthritis. (2) Total CXCL8 is extracted from patient samples by immunosorbent purification using antibody-coupled magnetic beads. (3) Isolated CXCL8 proteoforms are subjected to analysis by nano-UPLC-MS/MS. (4, 5) The detection and quantification of signature fragment ions, generated by top-down collision-induced dissociation (CID) of a pre-selected precursor ion, at the expected elution time point, strongly indicates the presence of a specific CXCL8 proteoform. In general, truncated CXCL8 proteoforms display an enhanced capacity to induce neutrophil migration and activation. This figure is created with BioRender software.