ABSTRACT: The actin-based cell-cell adherens junction (AJ) between the Sertoli cell and the germ cell in the mammalian testis is important not only in mechanical adhesion of the cells, but in the morphogenesis and differentiation of the germ cells. The Sertoli ectoplasmic specialization (ES), a specialized type of AJ, is associated with Sertoli-spermatid binding and is important in cell-cell adhesion in the seminiferous epithelium. Abnormal or absent Sertoli ESs have been associated with step-8 spermatid sloughing and oligospermia in conditions associated with reduced fertility potential. The reproductive hormones, follicle stimulating hormone (FSH), and testosterone (T) have also been shown to play a role in the regulation of binding of spermatids at the Sertoli-spermatid junctional complex (STJC). Adjudin [1-(2,4-dichlorobenzyl)-1H-indazole-3-carbohydrazide] is a potential male contraceptive and is thought to exhibit its contraceptive effects by interrupting the STJC. It has been shown that this compound induces reversible germ cell loss from the seminiferous epithelium, particularly elongating/elongate/round spermatids and spermatocytes. Using a micropipette pressure transducing system (MPTS) to measure the force needed to detach step-8 spermatids from Sertoli cells, this study examined the strength of the STJC in Sertoli-spermatid cocultures in the presence of Adjudin (1 ng/mL, 50 ng/mL, 125 ng/mL, or 500 ng/mL in EtOH) and hormones [FSH (0.1 μg/mL, NIDDK-oFSH-20, AFP7028D, 175 × NIH-FSH-S1), T (100 nM)] to optimize in vitro binding. The average forces required to detach the spermatids from the underlying Sertoli cells in the presence of 1 ng/mL, 50 ng/mL, 125 ng/mL, and 500 ng/mL Adjudin were 18.2 × 10−10 pN, 14.3 × 10−10 pN, 7.74 × 10−10 pN, and 6.51 × 10−10 pN, respectively. The average force required to detach step-8 spermatids in the presence of vehicle only (control) was 19.0 × 10−10 pN. A significant difference for Adjudin concentrations at or above 125 ng/mL was determined by one-way ANOVA (P < .05). These data confirm that Adjudin is effective in reducing the strength of the STJC, identifying Adjudin as a potential contraceptive agent in the male by inducing spermatid sloughing and therefore oligospermia.