Introduction: It is unclear if sexual orientation is a biological trait that has neurofunctional footprints. With deep learning, the power to classify biological datasets without an a priori selection of features has increased by magnitudes. The aim of this study was to correctly classify resting-state electroencephalogram (EEG) data from males with different sexual orientation using deep learning and to explore techniques to identify the learned distinguishing features. Methods: Three cohorts (homosexual men, heterosexual men, and a mixed sex cohort), one pretrained network on sex classification, and one newly trained network for sexual orientation classification were used to classify sex. Further, Grad-CAM methodology and source localization were used to identify the spatiotemporal patterns that were used for differentiation by the networks. Results: Using a pretrained network for classification of males and females, no differences existed between classification of homosexual and heterosexual males. The newly trained network was able, however, to correctly classify the cohorts with a total accuracy of 83%. The retrograde activation using Grad-CAM technology yielded distinctive functional EEG patterns in the Brodmann area 40 and 1 when combined with Fourier analysis and a source localization. Discussion: This study shows that electrophysiological trait markers of male sexual orientation can be identified using deep learning. These patterns are different from the differentiating signatures of males and females in a resting-state EEG.