Unsupervised HIV self-testing (HST) has potential to increase knowledge of HIV status; however, its accuracy is unknown. To estimate the accuracy of unsupervised HST in field settings in Uganda, we performed a non-blinded, randomized controlled, non-inferiority trial of unsupervised compared with supervised HST among selected high HIV risk fisherfolk (22.1 % HIV Prevalence) in three fishing villages in Uganda between July and September 2013. The study enrolled 246 participants and randomized them in a 1:1 ratio to unsupervised HST or provider-supervised HST. In an intent-to-treat analysis, the HST sensitivity was 90 % in the unsupervised arm and 100 % among the provider-supervised, yielding a difference 0f −10 % (90 % CI −21, 1 %); non-inferiority was not shown. In a per protocol analysis, the difference in sensitivity was −5.6 % (90 % CI −14.4, 3.3 %) and did show non-inferiority. We conclude that unsupervised HST is feasible in rural Africa and may be non-inferior to provider-supervised HST.