While previous studies suggest that textual glosses presented along with reading text facilitate deeper textual understanding and vocabulary uptakes, the extent to which such effects vary in different conditions remain obscure. Employing a quantitative research design, the authors specifically focused on the interplay between glossing languages (L1 and L2) and learners’ language proficiency (intermediate and advanced), and the effects of glosses on learners’ textual comprehension and incidental vocabulary learning. In this study, 180 learners were divided into two groups according to their proficiency level (intermediate or advanced). Within each group, the treatment conditions (control, glossing in native language, and glossing in second language) provided the foundation for further division into three sub-group of 60 students. ANOVA and t-test analysis show that the use of glosses led to deeper comprehension and incidental vocabulary learning. Two-way ANOVAs indicate that L1 glosses generated deeper comprehension and recall ability in intermediate learners than L2 glosses, and inversely, advanced learners performed better in reading and vocabulary tests when given L2 glosses. Finally, the authors proposed some implications for vocabulary learning and teaching in accordance with the relative differences in L1 and L2, and intermediate and advanced levels.