We investigated how bats use habitat structure along the vertical gradient of an old-growth Sequoia sempervirens (redwood) forest. Ground-based detection methods would underrepresent bats that use the canopy and above-canopy airspace in this forest as they reach far beyond practical netting and acoustic detection range. We equipped two tall trees with full spectrum automated bat detectors at treetop (108 m), lower crown (55 m), and ground level (5 m) from April 2008 to November 2009, excluding December and January. We sampled 1,365 detector nights, recorded 3,769 echolocation sequences (bat passes), and found 12 species, two of which, Lasiurus blossevillii and Tadarida brasiliensis, that had no prior documented presence in redwood forests. The maximum proportion of bat activity occurred at treetop, although ground level species diversity exceeded that of the lower crown and treetop, and species composition differed among locations. Non-Myotis species composed 95% of the calls at the treetop, 88% at lower crown, and 21% at ground level. Calls from Myotis species averaged 71% of all calls recorded at ground level compared to less than 4% at both lower crown and treetop. Activity declined markedly, but did continue, during the winter months we sampled. The combination of a temperate climate and observations of larger, migratory species during November, February, and March suggested the potential for resident populations or inland migrants overwintering in this forest. These findings emphasize the importance of sampling throughout the calendar year and including the full reach of the vertical habitat when quantifying bat activity in forests.