Anthers of male-sterile mutant (ms₃) and normal soybean [Glycine max (L.) Merr.] were compared by using a triple-staining procedure for DNA, polysaccharides and proteins, and a single-staining procedure for sporopollenin in conjunction with brightfield and fluorescent microscopy. The purpose of this study was to determine at what stage of anther development histological differences were detectable between male-sterile (ms₃) and normal soybean. At the tetrad stage, a prominent greenish-blue material, visible with the triple-staining procedure, filled some of the enlarged tapetal cells of anthers of ms₃ plants. This material was also autofluorescent and had an affinity for the fluorochrome primuline which identified sporopollenin. By the early microspore stage, microspores of ms₃ plants were unusually enlarged and the tapetum surrounding them prematurely degenerated. This abnormality of tapetal behavior seems to be associated with microspore abortion and serves as an early marker for sterility. Other differences noted between the anthers of ms₃ and normal plants with the staining procedures used, were variations in the endothecium and the thickness and degree of staining of the microspore and pollen walls. All of these differences suggest that the effect of the ms₃ gene in the homozygous condition is expressed during early anther development, probably beginning at meiosis and directly influences the malfunctioning of the tapetum. The results of this study support numerous other published reports on both genetic and cytoplasmic male sterility implicating the tapetum as the primary tissue responsible for male sterility.