Bamboo has excellent mechanical properties compared to wood and other plant materials, due to its multilayered structure and polytropic microfibril angle (MFA). The micro/nano scale structure and MFA of fibers, parenchyma cells, and vessels from 4-year-old Moso bamboo (Phyllostachys Heterocycla Var. Pubescens) were investigated by a novel LC-PolScope imaging system and transmission electron microscopy. At the nanoscale, the numbers of layers and accurate MFA for each layer especially thin layers could be obtained quickly using this novel LC-PolScope imaging system. Based on the differences of structure and shape, fibers and parenchyma cells in the vascular bundle were divided into FI, II, III and PI, II cells, respectively. The former class of FI, II, III included 2, 6–8, and 6–8 secondary cell wall layers in turn. The latter class exhibited 9 secondary cell wall layers, with a maximum of 16 layers. To our knowledge, this is the first report of accurate MFA measurement based on the differences of structure and shape for every layer of single fibers, parenchyma cells and vessels in the vascular bundle. For all three cell types, the results also showed that the MFA of sub-layers in secondary walls followed the same changing law: alternating smaller and then bigger MFA. This structural form may be the consequence of natural selection and optimization indicating the long-term mechanical adaptation of bamboo.