This review highlights the current trends and the most recent advances in the field of preparation and application of organic polymer‐based monolithic materials and covers literature published in 2020. A short historical background is provided and protocols to anchor monoliths covalently to the wall of the column/separation device are discussed. Furthermore, advances in tuning the macroporous structure and establishing its link to separation performance are conferred. Finally, method development and key applications using novel monolithic columns are discussed.

T lymphocytes are key components in adaptive immunity and their activation naturally involves mitochondrial-derived oxygen species (mtROS). In particular, H2O2 has been implicated as an important signaling molecule regulating major T cell functions. H2O2 targets the oxidation status of functional cysteine residues but knowledge if and where this happens in T cell signaling networks is widely missing. This study aimed to identify mtROS-sensitive processes in activated primary human CD4 + T cells. By using a thiolspecific redox proteomic approach we examined the oxidation state of 4784 cysteinecontaining peptides of ex vivo stimulated T cells from healthy individuals. Upon activation, a shift in oxidation was observed at catalytic cysteine residues of peroxiredoxins (PRDX5 & PRDX6), and T cells were found to maintain their global thiol-redox homeostasis. In parallel, a distinct set of 88 cysteine residues were found to be differentially oxidized upon T cell activation suggesting novel functional thiol switches. In mitochondria, cysteine oxidations selectively modified regulators of respiration (NDUFA2, NDUFA8, and UQCRH) confirming electron leakage from electron transport complexes I and III. The majority of oxidations occurred outside mitochondria and enriched sensitive thiols at regulators of cytoskeleton dynamics (e.g. CYFIP2 and ARPC1B) and known immune functions including the non-receptor tyrosine phosphatase PTPN7. Conversely, cysteine reduction occurred predominantly at transcriptional regulators and sites that coordinate zinc-binding in zinc-finger motifs. Indeed, fluorescence microscopy revealed a colocalization of zinc-rich microenvironments and mitochondria in T cells suggesting mtROS-dependent zincrelease of identified transcriptional regulators including ZFP36, RPL37A and CRIP2. In conclusion, this study complements knowledge on the mtROS signaling network and suggests zinc-dependent thiol switches as a mechanism of how mtROS affects transcription and translation in T cells.

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This study reports on the potential of using ion-exchange suppressor technology in liquid chromatography—electrospray ionization mass spectrometry workflows. The aim was to use high salt concentrations to improve separation performance, while overcoming the resulting significant ion suppression during electrospray ionization. As a case study, we apply suppressor technology to the hydrophilic interaction liquid chromatography separation and detection of taurine and glycochenodeoxycholate sulfate, endogenous biomarkers for organic anion transporter protein inhibition. The desired chromatographic selectivity was achieved applying 100 mM ion-pairing agent, while competing ions negatively affecting MS sensitivity were actively removed post-column from the solvent via a charged partially permeable membrane and replaced with protons, resulting in an up to 10-fold increase in detection sensitivity.

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