A decrease in sperm motility and thus total motile sperm count (TMSC) over a period of hours may have clinical implications in counseling couples considering intrauterine insemination (IUI), in vitro fertilization (IVF), and intracytoplasmic sperm injection (ICSI). The objective of this study was to identify patients with decreases in sperm motility from 1 hour to 2 hours after collection and examine predictive relationships with semen analysis parameters. Between 2001 and 2005, 2313 semen samples were analyzed. Sperm motility was evaluated at both 1 and 2 hours after time of collection. Relevant seminal parameters were compared between patients with a decrease from the 1-hour to 2-hour motility (n=384) vs. those that showed no change (n=1929). The same analysis was performed in a subset of patients with a TMSC between 10-40 million. In the total patient population, only 16% (384/2313) demonstrated a decrease in the 1-hour to 2-hour motility. In patients displaying a decrease in the 1-2 hour motility, sperm concentration (33.5 vs.79 mil/ml, p<0.0001) and % normal morphology (7% vs. 8%, p<0.0001) were significantly lower. Additionally, a significantly higher incidence of 1-2 hour motility decrease was seen in patients with midpiece anomalies (33.3% vs. 15.9%, p=0.01). Within the subset population of TMSC 10-40 million, the only statistically significant difference was in patients with midpiece anomalies (80.0% vs. 28.2%, p=0.02) who demonstrated a higher incidence of 1-2 hour motility decrease. Overall, patients with a TMSC between 10-40 million showed a significantly higher incidence of 1-hour to 2-hour motility decrease compared to the rest of the patient population (29.0% vs. 14.6%, p<0.0001). Because decreases in the 1 to 2-hour sperm motility affect only a small portion of patients, it is not necessary to check 2-hour motility on all patients. However, because patients with a TMSC between 10-40 million were significantly more likely to show a decrease in sperm motility, a decrease which could have possible clinical implications in couples deciding between IUI, IVF or ICSI, checking 2-hour sperm motility should be considered in this population.