This study evaluated acceptability, engagement in prevention, and efficacy of a primary care screening-and-referral-to-prevention program to reduce substance use in early adolescence. Screening tools were the Youth Risk Index and Transmissible Liability Index and prevention consisted of the Family Check-Up (FCU). Three hundred sixty-one 10- to 13-year-olds from low resource neighborhoods (85.9% African American; 52.4% female) screened “at risk” during primary care visits and were randomized to the FCU (n = 123) or usual care (n = 238). Screening was acceptable to parents and youths: nearly 95% of each rated it as important, about 90% of each were happy with or did not mind it, and only 2.4% of parents did not want their child to be screened at their next check-up. Of parents who had a chance to receive the FCU (or waitlist-control), 87.5% followed through with researchers while 93.5% who were offered FCU engaged in it. FCU efficacy primarily involved interactions such that youth with greater risk at baseline experienced larger benefits. At 12-month follow-up, FCU was associated with 11% reduced risk of initiating a new substance per substance that had been initiated before baseline; greater reductions in tolerance of deviance among those with higher tolerance of deviance at baseline; and a main effect of reduced anxiety, but no effect for conduct problems. Pediatric well-child check-up screening can identify high-risk youth before, or in the initial stages of, problematic SU; engage families in a preventive intervention; and reduce rates of substance use and related risk factors.