This study aimed to validate the use of two-step Modified Checklist for Autism in Toddlers (M-CHAT) screening adapted for a Thai population. Our participants included both high-risk children with language delay (N = 109) and low-risk children with typical development (N = 732). Compared with the critical scoring criteria, the total scoring method (failing =3 items) yielded the highest sensitivity of 90.7%; specificity was 99.7%, positive predictive value 96.1%, and negative predictive value 99.4%. The two-step M-CHAT screening is a promising instrument that can be utilized to detect ASD in Thai children in both primary and clinical settings. Moreover, socio-cultural context should be considered when adopting the use and interpretation of the M-CHAT for each country.