A change of z-directional structural and surface properties by calendering has a great influence on liquid penetration into a sheet. It could be also important for multi-ply sheet because it contacts liquid dunhg coating or converting process. Therefore, this study was aimed to evaluate of a change of z-directional structure in multi-ply sheet by calendering. To do this, multi-ply sheets were prepared with various raw materials and calendered at the different pressure and temperature conditions. In multi-ply sheet which consisted of one kind of pulp fiber, thickness reductions were higher in top and bottom plies than in middle plies. And in the case of soft nip calender treatment with high temperature, top layer which was in contact with heating roll showed the highest reduction of thickness. Hard nip calender treatment showed U-shaped density profile in z-direction, but compression profile by SNC treatment was dependent on calendering condition. To examine z-directional structure of multi-ply sheet which was composed of different raw material for each layer, CLSM (Confocal Laser Scanning Microscopy) analyses were carried out on cross direction of sheet. It turned out to be a useful tool for investigating z-directional analysis. As a result, variation of thickness reduction in z-direction is dependent on ply structure, compressibility of pulp fiber, and calendering condition.