Multiplexed single-molecule measurements with magnetic tweezers.

We present a method for performing multiple single-molecule manipulation experiments in parallel with magnetic tweezers. We use a microscope with a low magnification, and thus a wide field of view, to visualize multiple DNA-tethered paramagnetic beads and apply an optimized image analysis routine to track the three-dimensional position of each bead simultaneously in real time. Force is applied to each bead using an externally applied magnetic field. Since variations in the field parameters are negligible across the field of view, nearly identical manipulation of all visible beads is possible. However, we find that the error in the position measurement is inversely proportional to the microscope's magnification. To mitigate the increased error caused by demagnification, we have developed a strategy based on tracking multiple fixed beads. Our system is capable of simultaneously manipulating and tracking up to 34 DNA-tethered beads at 60 Hz with approximately 1.5 nm resolution and with approximately 10% variation in applied force.