The translocation of long-chain molecules, such as DNA or peptides, through membranes is an integral process for the function of living cells. During the translocation process, a number of interactions of electrostatic or hydrophobic nature govern the translocation velocity. Most of these interactions remain largely unexplored on the single-molecule level due to a lack of suitable instrumentation. We have shown that a combination of optical tweezers, single solid-state nanopores, and electrophysiological ionic current detection can provide further insight into the behavior of polymers in confinement. Here, we describe the experimental procedures necessary for manipulation of single biopolymers in a single nanopore not only by electrical fields, but also through mechanical forces using optical tweezers.