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In a typical single-molecule mechanical assay, the molecule of interest is attached to a sensitive probe such as two micron-sized beads held in optical tweezers.
By using conditional Piezo1 deletion from red blood cells, a novel mechanical assay combining fluorescent calcium-imaging with cell membrane stretch and the novel chemical Piezo1-specific agonist described in the accompanying manuscript they demonstrate that calcium-influx is completely dependent on Piezo1 and leading to cell dehydration via the ion channel KCAa3.1.
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Once we obtained microcores of different shapes and size manufactured from mortars of known dosage, the mechanical assays were performed for normalized specimens and microcores.
Meanwhile, coupling the typical fluorescent assays such as fluorescence resonance energy transfer with those mechanical assays also enables to visualize the intracellular in situ events under given mechanical stimuli.
Thus, pericarp tissue at MG, BR and BR+8 stages was subjected to two types of mechanical assays, i.e. compression and penetration.
MKM performed all cell culture, and biochemical and mechanical assays, and scanning electron imaging.
Our results thus establish DNA helix bundles as an attractive linker system for single-molecule mechanical assays.
1 Single-molecule mechanical assays have been successfully applied to analyze large conformational changes like the ones that occur in protein unfolding or in the motion of molecular motors.
The role of CXCR2 was explored using three cell-mechanical assays.
We, therefore, developed a mechanical stimulation assay combining the advantage of calcium imaging (no gigaohm seal necessary) and patch clamp (a pipette to capture the cell).
2) When explaining the novel mechanical stimulation assay the authors fail to mention that cells are loaded with fluorescent calcium indication (Fluo-4) to monitor calcium levels.
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