The Multidimensional Acculturative Stress Inventory (MASI), a 36-item measure that assesses acculturative stress among people of Mexican origin living in the United States, was tested on 331 adolescent (14–20 years of age) high school students (204 female, 127 male) of Mexican origin. Exploratory factor analyses yielded 4 factors: bicultural practices conflict (9 items), Spanish competency pressures (8 items), English competency pressures (8 items), and bicultural self-consciousness (2 items). These factors accounted for 59.5% of the variance and correlated in the expected directions with criterion measures of acculturation and the Psychological General Well-Being Schedule. Bicultural practices conflict and bicultural self-consciousness emerged as the first and fourth factors for adolescents, which differed from the last 2 factors observed in a previous study of adults by Rodriguez, Myers, Mira, Flores, and Garcia-Hernandez (2002)—pressure to acculturate and pressure against acculturation. Comparisons of the MASI factor structures between adolescents and adults also revealed that English competency pressures and Spanish competency pressures played a prominent role for both adolescents in this study and adults in the study by Rodriguez et al. (2002). The congruence and difference in factor structure of the MASI between adolescents and adults indicates that both groups experience acculturative stress because of English- and Spanish-language competency pressures, but adolescents differentially experience difficulties in negotiating between American and Latino practices and identities. The results highlight the importance of assessing acculturative stress from both Latino and American culture and recognizing the varying levels of these sources of acculturative stress by generation.