BACKGROUND: Some previous research has suggested that testosterone prenatally contributes to gender differences in autistic traits, but little is known about the role of testosterone during early postnatal development (mini-puberty). Two prior studies found no sex difference in testosterone postnatally in saliva samples and detected little to no relationship between testosterone postnatally and autistic traits in toddlers. These findings may reflect late measurements of testosterone at 3 to 4 months of age, after the peak of mini-puberty at 1 to 3 months of age. The present study examined the relationship between testosterone at 1 to 3 months of age and autistic traits at 18 to 30 months of age. FINDINGS: Testosterone was measured in saliva samples collected from children at 1 to 3 months of age. When the children (40 boys, 47 girls) reached 18 to 30 months of age, parents completed the Quantitative Checklist for Autism in Toddlers (Q-CHAT). Boys had higher concentrations of testosterone postnatally and higher Q-CHAT scores than girls. However, testosterone did not correlate with Q-CHAT scores in boys, girls, or the entire sample. CONCLUSIONS: The current results suggest that testosterone during the early postnatal period does not contribute to later autistic traits. Given our relatively small samples and therefore limited power, however, further research could usefully examine if testosterone in saliva samples collected during the peak of mini-puberty in larger groups predicts autistic traits or other traits that show gender differences.