Alexithymia, a difficulty identifying and communicating one’s own emotions, affects socio-emotional processes, such as emotion recognition and empathy. Co-occurring alexithymia is prevalent in Autism Spectrum Disorder (ASD) and underlies some socio-emotional difficulties previously attributed to autism. Socio-emotional abilities are examined during behavioural diagnostic assessments of autism, yet the effect of alexithymia on these assessments is not known. In this study, two previously collected samples of Autism Diagnostic Observation Schedule (ADOS) assessments were used to examine the relationship between alexithymia and ADOS score. Participants included 96 women with anorexia, and 147 adolescents who were either high in autistic symptoms, or whose twin had high autistic symptoms. We examined 1) the impact of alexithymia on meeting the criteria for autism/ASD, 2) correlations between alexithymia and ADOS subscales, and 3) whether alexithymia predicted scores on specific ADOS items, selected a priori based on the literature. In the adolescent group, parent-reported (but not self-reported) alexithymia predicted autism/ASD diagnosis, correlated with both sub-scales, and predicted scores on ADOS items. Fewer associations were observed in the anorexic sample, but the presence of alexithymia predicted a near fourfold increase in the likelihood of meeting diagnostic criteria for autism/ASD in this sample.