We examined the differentiation of the generative and the vegetative cells of tobacco (Nicotiana tabacum SR-1) by a newly developed cytological method using fluorescence microscopy after embedding of samples in Technovit 7100 resin. The extent of condensation of the chromatin in generative and vegetative nuclei was measured semi-quantitatively in sections of uniform thickness by microphotometry. The extent of chromatin condensation in the generative nucleus was about 5 to 10 times as high as the vegetative nucleus throughout pollen maturation which lasted about 5 days. Such a difference was clear immediately after PM I, indicating that the differentiation of the two cells had already started as soon as PM I has finished. Maturing generative cells had almost as much condensed chromatin as chromosomes of PM I. We also examined whether the generative cell has the potential for gene expression. The localization of RNA was visualized by staining with acridine orange. The results revealed the cytoplasm of the generative cell contained RNA and the density of RNA was as high as in the vegetative cell. This result suggests that the generative cell has the potential for gene expression in spite of the highly condensed chromatin in the nucleus. Furthermore, the generative nucleus in the mature pollen retained small nucleoli. Their presence implies that rRNA is synthesized consistently in generative cells.