<p>Frame-based stereotactic biopsy has long been the gold standard for diagnostic brain biopsy, whereas robot-assisted stereotactic techniques have emerged as promising alternatives. However, rigorous methodological comparisons between these approaches remain scarce. To compare the diagnostic yield and complication rates of frame-and robot-assisted stereotactic brain biopsy using a rigorously matched cohort methodology. This retrospective, bicentric, comparative analysis included adult patients who underwent stereotactic brain biopsy between January 2011 and December 2019. Each patient who underwent robot-assisted biopsy (<i>n</i> = 230) was matched in a 2:1 ratio with patients who underwent frame-based biopsy (<i>n</i> = 460). Matching was performed based on clinically relevant criteria including lesion location, size, contrast enhancement, histopathology, and patient age. The primary outcomes were diagnostic yield and complication rates. The diagnostic yield was identical for both techniques 97.4%, with a non-diagnostic biopsy rate of 2.6% in each group (<i>p</i> = 1.0). The symptomatic temporary complication rates were higher in the robot-assisted group (6.5%) than in the frame-based group (2.8%) (OR = 2.40; 95% CI: 1.04–5.58; <i>p</i> = 0.02). No differences were found in symptomatic hemorrhage, infection, seizures, permanent neurological deficits, or mortality between groups. Frame-based and robot-assisted stereotactic biopsy techniques demonstrated equivalent diagnostic efficacies. Although robot-assisted biopsies were associated with a higher incidence of transient symptomatic complications, both techniques exhibited favorable safety profiles with respect to severe complications. The choice of technique should consider the surgeon’s experience, institutional resources, and integration within existing workflows.</p>