Strategies for Overcoming the Single-Molecule Concentration Barrier

Fluorescence-based single-molecule approaches have helped revolutionize our understanding of chemical and biological mechanisms. Unfortunately, these methods are only suitable at low concentrations fluorescent molecules so that single species interest can be successfully resolved beyond background signal. The application techniques has therefore been limited to high-affinity interactions despite most processes occurring much higher reactant concentrations. Fortunately, recent methodological advances demonstrated this concentration barrier indeed broken, with reaching as high 1 mM. goal Review is discuss the challenges in performing fluorescence high-concentration, offer applications both biology chemistry, highlight major milestones shatter barrier. We also hope inspire widespread use we begin exploring new physical phenomena lying