Electrokinetic trapping at the one nanometer limit

Abstract

Anti-Brownian electrokinetic traps have been used to trap and study the free-solution dynamics of large protein complexes and long chains of DNA. Small molecules in solution have thus far proved too mobile to trap by any means. Here we explore the ultimate limits on trapping single molecules. We developed a feedback-based anti-Brownian electrokinetic trap in which classical thermal noise is compensated to the maximal extent allowed by quantum measurement noise. We trapped single fluorophores with a molecular weight of <1 kDa and a hydrodynamic radius of 6.7 Å for longer than one second, in aqueous buffer at room temperature. This achievement represents an 800-fold decrease in the mass of objects trapped in solution, and opens the possibility to trap and manipulate any soluble molecule that can be fluorescently labeled. To illustrate the use of this trap, we studied the binding of unlabeled RecA to fluorescently labeled single-stranded DNA. Binding of RecA induced changes in the DNA diffusion coefficient, electrophoretic mobility, and brightness, all of which were measured simultaneously and on a molecule-by-molecule basis. This device greatly extends the size range of molecules that can be studied by room temperature feedback trapping, and opens the door to further studies of the binding of unmodified proteins to DNA in free solution.