The gas-phase acidities of the two aromatic sites in benzocyclobutene were measured in a Fourier transform mass spectrometer using a kinetic technique (i.e., the DePuy method). Fusion of a cyclobutane ring onto benzene is found to have a slight acidifying effect at the α-position (3.2 ± 1.7 kcal mol-1) and little, if any, influence on the β-site (0.8 ± 1.9 kcal mol-1). Energetic data (ΔH°(acid) = 386.2 ± 3.0 kcal mol-1, EA = 0.84 ± 0.11 eV, and C-H BDE = 92 ± 4 kcal mol-1) for the benzylic position were obtained via the bracketing technique and application of a thermodynamic cycle. Differences in the reactivities of the three conjugate bases also were explored. Ab initio and density functional theory calculations were carried out to provide geometries, energies, and insights into the carbanions' electronic structures.