Instructor Talk—noncontent and nonlogistical language that is focused on shaping the classroom learning environment—is a recently defined variable that may play an important role in how undergraduates experience courses. Previous research characterized Instructor Talk used by faculty teaching in biology lecture classrooms. However, graduate teaching assistants (GTAs) and laboratory classrooms represent critical factors in undergraduate education, and Instructor Talk in this context has yet to be explored. Here, we present findings analyzing Instructor Talk used by GTAs teaching in undergraduate biology laboratory classrooms. We characterized the Instructor Talk used by 22 GTA instructors across 24 undergraduate biology laboratory courses in the context of a single, urban, Hispanic-serving and Asian American and Pacific Islander–serving Institution. We found that Instructor Talk was present in every course studied, GTAs with pedagogical training and prior teaching experience used more Instructor Talk than those without, and GTAs teaching laboratory courses used more Instructor Talk than previous observations of faculty teaching lecture courses. Given the widespread use of Instructor Talk and its varying use across contexts, we predict that Instructor Talk may be a critical variable in teaching, specifically in promoting equity and inclusion, which merits continued study in undergraduate science education.