(1) Background: This study aimed to validate the Greek version of the Autism Parenting Stress Index (APSI) among parents of children with ASD. (2) Methods: The translated version was administered to 113 parents (Male: 12, Female: 101, 39.24 years old, SD 6.70, age range, 25–58) of children diagnosed with ASD and 127 parents (Male: 24, Female: 103, 41.08 years old, SD 6.22, age range: 27–56) of typically developing children. (3) Results: Significant differences between the APSI total scores and three domains between groups were observed. Although the initial factor structure could not be replicated, the APSI’s internal consistency was excellent (a = 0.914), with a high positive item–total correlation (0.900–0.917). The APSI’s test–retest reliability was excellent, showing an ICC equal to 0.922 [95%, CI: 0.900–0.940]. The APSI’s total score cut-off point was equal to 12.00 (AUC 0.845, p < 0.001) with a sensitivity of 0.839 and 1-specificity of 0.220. A principal component analysis of the 13 items, using varimax rotations, identified three factors, which explained approximately 45.8% of the overall variance. (4) Conclusions: The Greek version of the APSI exhibited discriminant validity for measuring parents of children with ASD. Greek health professionals can use it to assess the stress experienced by parents of children with ASD.