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PURPOSE To evaluate the biomechanical changes occurring after LASIK flap creation and rapid corneal cross-linking (CXL) measured with Brillouin light microscopy. METHODS Porcine eyes (n = 11) were evaluated by Brillouin light microscopy sequentially in the following order: virgin state, after LASIK flap creation, and after rapid CXL. Each eye served as its own control. Depth profile of the Brillouin frequency shift was computed to reveal the depth-dependent changes in corneal stiffness. RESULTS There was a statistically significant reduction of Brillouin shift (reduced corneal stiffness) after LASIK flap creation compared to virgin corneas across total corneal thickness (-0.035 GHz, P = .0195) and within the anterior stromal region (-0.104 GHz, P = .0039). Changes in the central (-0.029 GHz, P = .0391) and posterior (-0.005 GHz, P = .99) stromal regions were not significant. There was a small increase in Brillouin shift after rapid cross-linking that was not statistically or clinically significant across total corneal thickness (0.006 GHz, P = .4688 for any specific stromal region; 0.002 to 0.009 GHz, P > .46 for all). CONCLUSIONS LASIK flap creation significantly reduced Brillouin shift in the anterior third of the stroma in porcine eyes. Rapid corneal cross-linking had no significant effect on Brillouin shift after LASIK flap creation in porcine eyes. With further validation, non-contact, non-perturbative Brillouin microscopy could become a useful monitoring tool to evaluate the biomechanical impact of corneal refractive procedures and corneal cross-linking protocols. [J Refract Surg. 2017;33(6):408-414.].