For better understanding the functions and mechanism of PMEs and HGs in the growth and development of banana and in the defense of banana to the stresses, we traced the spatial and temporal abundance and distribution of banana pectin methylesterases (PMEs) and five homogalacturonans (HGs) at different methylesterification degrees (MDs) in banana leaves at different developmental stages using immunofluorescence labeling techniques. In addition, enzyme activity of PMEs was also tested. The results indicated that: (1) PMEs located mainly in the mesophyll and guard cells in tissuecultured banana seedlings. Both the level and activity of PMEs decreased with the development of the plant. (2) HGs at different MDs also mainly appeared in mesophyll cells in tissuecultured plants and their contents reduced gradually with the development of plants, though the speed varied with each other. However, the distribution of these HGs in the epidermis, the guard cells and the vein of banana leaves showed differential patterns. The MDs of HGs present in the guard cells was relatively high. These results indicated that PMEs and five tested HGs were mainly present in the mesophyll. During the development of banana leaves, the levels of these HGs showed similar trend to that of PMEs.