The ability of block copolymers (BCP) to self-assemble into highly ordered, periodic nanostructures makes them a promising candidate for nanoscale templating applications. In recent years, the refinement of BCP processing methods has enabled the fabrication of essentially defect-free patterns on the wafer scale. However, the dynamics of BCP assembly are still poorly understood, and the most important processing steps are also the least experimentally accessible. In particular, annealing procedures, which enhance BCP mobility for structural reorganization, involve high temperatures or harsh solvent environments that hinder many analytical methods. This thesis presents research on diverse BCP systems involving several different stages of the processing pipeline. Altogether, the differing topics presented here are unified in exploring the preparation and application of BCP patterns under controlled environmental conditions in situ with atomic force microscopy (AFM). On a flat surface, BCP films natively form disordered fingerprint patterns that lack long-range order. Several substrate modification strategies — collectively referred to as directed self-assembly (DSA) — may be used to control the orientation and alignment of a BCP overlayer for the preparation of controlled pattern morphologies. As part of the DSA process, thermal annealing of BCP above its glass transition temperature increases chain mobility and allows for pattern reorganization. In situ AFM enables the direct examination of the mechanisms and dynamics of defect healing during thermal annealing. The advent of high-speed AFM imaging has dramatically enhanced the imaging time resolution, allowing fine-grain analysis of structural intermediates in real time and space. In addition to mapping the BCP morphology, high-speed imaging enables the direct investigation of the interfacial dynamics and the influence of single-chain dynamics. Alongside thermal annealing, solvent treatment also facilitates polymer mobility and pattern rearrangements. Selective solvents, which have differing affinities for each of the component BCP blocks, partition into the polymer matrix and can produce diverse polymer morphologies with unique functional utility. In one example, solvent-vapor annealing selectively swells one block to produce a highly corrugated surface reconstruction. The resulting nanogrooves may then be used to template the end-to-end alignment of Au nanorods for the preparation of a molecular sensing surface. In a second experiment, BCP nanoislands act as a novel platform for the in situ measurement of nanoscale solvent-swelling behavior. By using the solvent as a carrier for fluorescent probes, this process offers a new, kinetically-controlled method for the localized functionalization of polymer surfaces. Collectively, these experiments demonstrate the utility of in situ AFM imaging for studying otherwise inaccessible BCP dynamics, while highlighting the method’s flexibility under a wide range of operational environments.