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Continuous wave (CW) and microsecond pulse (MP) laser irradiations were compared regarding cell damage and laser-induced temperature rise at retinal pigment epithelium (RPE). The RPE of porcine RPE-choroid-sclera explants was irradiated with a 577 nm laser in CW or MP mode (5% or 15% duty cycle (DC)) for 20 ms or 200 ms at an average laser power of 20–90 mW. Cell viability was investigated with calcein-AM staining. Optoacoustic (OA) technique was employed for temperature measurement during irradiation. For 200 ms irradiation, the dead cell area (DCA) increased linearly (≈1600 µm2/mW) up to the average power of 40 mW for all modes without significant difference. From 50 mW, the increase of DCA of MP-5% significantly dropped to 610 µm2/mW (p < 0.05), likely due to the detected microbubble formation. OA temperature measurement showed a monotonic temperature increase in CW mode and a stepwise increase in MP mode, but no significant difference in the average temperature increase at the same average power, consistent with the temperature modeling. In conclusion, there is no difference in the average temperature rise between CW and MP modes at the same average power regardless of DC. At lower DC, however, more caution is required regarding mechanical damage due to microbubble formation.