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To investigate the potential influence of breath holding on the results of peripapillary vessel density (VD) measurement with optical coherence tomography angiography.Three peripapillary VD measurements were made at 2-minute intervals on the right eye of 12 healthy young volunteers using the Angiovue/RTvue-XR OCT (software version 2017.1, Phase 7 update) without changing the head and body positions. During the first 2 measurements the volunteers were breathing normally. Four seconds before the third measurement the participants started to perform the Valsalva maneuver and continued until the completion of the ∼17-second image acquisition. The whole image area, the inside-disc area and the peripapillary area were investigated for capillary VD and all-vessels VD, separately.All volunteers had normal blood pressure and intraocular pressure, and all study eyes were normal with healthy optic nerve head and peripapillary area. No difference for any VD parameter was seen between the baseline images (P≥0.492). The differences between the averaged baseline and the during Valsalva maneuver values did not differ from 0 (P≥0.474), and the mean differences were ≤0.19% for the whole image, ≤0.35% for the peripapillary area, and ≤0.68% for the inside-disc parameters. The mean image quality score was >8 (optimal) in all image sessions.Our results suggest that breath holding during optical coherence tomography angiography image acquisition does not influence the measured peripapillary, whole image and inside-disc VD values, or the quality of the images, in healthy young subjects. Thus, no specific patient instruction regarding breathing is required before image acquisition.