HIV-1 serodiscordant couples may experience increased risks of relationship dissolution; however, longitudinal stability of these relationships is poorly understood. We determined rates and correlates of separation among 469 serodiscordant couples in Nairobi and found that 113 (24 %) separated during 2 years of follow-up. Couples with a female HIV-1 infected partner (F+M−) and no income were more likely to separate than M+F− couples without income (HR = 5.0; 95 % CI 1.1–25.0), and F+M− and M+F− couples with income (HR = 2.4; 95 % CI 1.3–4.5 and HR = 2.3; 95 % CI 1.2–4.8, respectively). High separation rates may be important for couple support services and for conducting discordant couple studies.