The development and use of chemical probes for redox biology has been hampered by several systematic problems. Only very few redox-active trigger chemotypes have been explored; in our field, this lack of diversity is exacerbated by the overwhelming use of linear disulfides in probes (Fig. ). The rational mechanism-based design of new redox-active motifs that are chemocompatible with the major biological redox systems, is urgently needed to drive innovation and research across the field. A second problem is the lack of clarity about what molecular information the known redox chemotypes do, or do not, provide. We believe that high-quality chemotypes should exhibit logical structure-activity relationships: for redox just as much as is accepted for ligand binding. A third problem is that redox selectivity testing is rarely as thorough as it could be. Selectivity is often stated despite it being obscure which cellular reductants have been controlled, and which have not; justifications of how much of a probe is claimed to be cellularly processed by the claimed species would be less ambiguous. The combination of these problems has resulted in a large number of redox probe reports claiming selectivity for various species, with even the same redox-active motifs being claimed as selective for a range of different redox species (Fig. ), occasionally without or against supporting data or rationale. We also intended this probe research to test how such systematic problems may be overcome with chemistry. The two cyclic selenenylsulfide cores are new chemical species that open up broad possibilities for probe adaptations, which are easily accessible through their operationally simple, divergent, gram-scale syntheses. The exploration of the solubiliser sidechains, and the comparisons to related probe structures, confirms the rule of structure-activity relationships from cell-free to cellular applications. The battery of cell-free and cellular tests we have employed have stringently assayed all key aspects of redox probe performance, in particular to validate RX1's TrxR1selectivity. The comprehensive cell-free characterisations, using key redox-active species and proteins in their full interacting systems, tested several key features that cellular assays later confirmed by orthogonal measurements. RX1's cell-free resistance to physiological levels of monothiols over hours was indeed shown to depend on mechanism, since its thermodynamically identical but mechanistically differentiated G1 analogue is GSH-labile. The cell-free selenolthiol-selectivity of RX1 was shown by its nonresponse to the TrxR(U498C) mutant, which until now was assumed to be similarly-reactive to most small molecule redox substrates including all that contain selenium . The cellular counterpoint of these assays has been the validation of TrxR1 as the only relevant cellular target of RX1, by three independent biological and chemical methods (knockout/knock-in, chemical inhibition, and selenium starvation). Comparison to the performance of mixed-mode G1 (processed by both TrxR and thiols, as revealed by all three methods) further substantiated RX1's cellular selectivity, and provides a comprehensive and quantitative basis for reporting RX1 as a TrxR1-selective probe.