SlipOChip - single-cell respiration under tuneable environments.

In disciplines like toxicology and pharmacology, oxygen (O) respiration is a universal metric for evaluating the effects of chemicals across various model systems, including mammalian and microalgal cells. However, for these cells the common practice is to segregate populations into control and exposure groups, which assumes direct equivalence in their responses and does not take into account heterogeneity among individual cells. This lack of resolution impedes our ability to precisely investigate differences among experimental groups with small or limited sample sizes. To overcome this barrier, we introduce SlipOChip, an innovative glass microfluidic platform for precisely quantifying single-cell O respiration in the coordinated absence and presence of chemical solutes. SlipOChip comprises a wet-etched fused silica channel plate on the top and a dry-etched borosilicate microwell plate at the bottom. The microwells are coated with Pt(II) -tetra(pentafluorophenyl)porphine (PtTFPP), an O sensing optode material and an O-independent reference dye. A custom 3D-printed holder facilitates the controlled horizontal movement ('slipping') of the channel plate over the microwell plate, thereby establishing or disrupting the fluid path over microwells. Collectively, these design elements enable the immobilization of single-cells in microwells, their exposure to controlled fluid flows, the coordinated opening and closing of microwells and repeated measurements of single-cell O respiration. Uniquely, by sequentially executing opening and closing it becomes possible to measure single-cell respiration prior to and after exposure to chemical solutes. In a proof-of-concept application, we utilized SlipOChip to measure the impact of increasing exposures of the marine bacterial signal 2-heptyl-4-quinolone (HHQ) on the dark respiration of the diatom at single-cell resolution. Results revealed a concentration-dependent decrease in per-cell O dark respiration, with a maximum reduction of 40.2% observed at HHQ concentrations exceeding 35.5 μM, and a half-maximal effective concentration () of 5.8 μM, consistent with that obtained conventional bulk respiration methods. The ability of SlipOChip to sequentially assess the effects of chemical substances on single-cell O metabolism is advantageous for research where sample volumes are limited, such as clinical biopsies, studies involving rare microbial isolates, and toxicological studies aiming to address exposure effects while accounting for cell-to-cell variability.