AbstractLabel-free in situ confocal Raman microscopy has been used to investigate the differences in cell morphology and components distribution between opposite wood (OW) and tension wood (TW) in Populus nigra. In the Raman images showing the ratio of the 1657 cm-1 to the 1603 cm-1 band, the spatial heterogeneity in coniferyl alcohol and coniferyl aldehyde (Lignin-CAA) was visualized within both samples. Enrichment of Lignin-CAA was visualized in the CC and CML regions where there was the highest lignin concentration. In fiber S2, which had the highest content of cellulose, there was apparent deficiency in the Lignin-CAA concentration. Raman spectra analysis revealed that the band at 1097.0 cm-1 in OW fiber S2 shifted to lower wavenumbers in TW fiber S2 (1094.4 cm-1) and GL (1094.0 cm-1), which resulted from the stretch of the glycosidic C-O-C bond of the cellulose molecular. The negative band shift at 1097 cm-1 indicated that the microfibrils in the TW fiber S2 and GL were stretched during TW formation and the microfibrils still kept the tensional deformation even after the fibers were transversally cut. The sub-cellular localization of cellulose and lignin together with the variation in molecular deformation will contribute to understanding the morphological and chemical properties of TW as a desirable woody biomass, as well as the development of its high tensile strength.