This study investigated differentiated instruction in EFL classrooms in higher education, particularly in a university English preparatory program in Turkey. The existing differentiation practices were determined using Tomlinson's (1999) framework that involves the differentiation of content, process, product, and learning environment based on students' readiness, interest, and learning profile using various instructional strategies. Fifty-one instructors teaching at the English preparatory program participated in the study, and the data were collected through a questionnaire adapted from Santangelo and Tomlinson (2012). The findings revealed that the instructors in the English preparatory program considered students' readiness level more than their interests and learning profile when differentiating instruction. It was also found that instructors preferred to differentiate the learning environment more than the content, process, and product. Finally, the study showed that the most frequently used differentiation strategies were providing supplemental materials for slow learners, using a variety of grouping formats, and supporting weak students to complete assignments, while the least frequently used ones were providing students with multiple text options, grouping students based on their interests, and allowing students to produce tasks in different forms.