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As a natural polymer material, bamboo is regarded as a super material with lightweight but excellent mechanical properties. With the increasing application of bamboo in structures, high-strength composite materials, bridges, musical instruments, and damping materials, the study of Poisson’s ratio is of great significance to the structural design and stability of these materials. In this work, Poisson’s ratio of bamboo for different ages, growth heights, and bamboo wall radial positions were determined by electrical measurement. The relationships between intrinsic factors (density, chemical composition, crystallinity, and microfibril angle) and Poisson’s ratio were also analyzed. The results showed that Poisson’s ratio of bamboo for different ages and locations ranged from 0.180 to 0.334, decreased with the increasing bamboo age, growth height, and the radial position from the outer layer to the inner layer. While the peak values of Poisson’s ratio were obtained at bamboo of 2 years old, lower growth height, and near the outer layer of the bamboo. Furthermore, the Poisson’s ratio had a positive correlation with the increasing density, cellulose content, and MFA, but a negative correlation with lignin content. There is no direct relationship between the crystallinity and Poisson’s ratio, but it affects the axial and transverse deformation of bamboo, which can be used as an indirect index to judge Poisson’s ratio.