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Background: Impaired perfusion indices signal potential microvascular dysfunction preceding atherosclerosis and other cardiometabolic pathologies. Post-occlusive reactive hyperemia (PORH), a vasodilatory response following a mechanically induced ischemia, is a transient increase in perfusion and can assess microvascular function. The greatest blood flow change corresponding to the first minute of hyperemia (represented by time-to-peak, hyperemic velocity, AUC within 1st min) has been shown to indicate microvascular dysfunction. However, the reproducibility of these temporal kinetic indices of the PORH response is unknown. Our aim was to examine the inter- and intra-day reproducibility and standardization of reactive hyperemia, with emphasis on the kinetic indices of PORH, using laser speckle contrast imaging (LSCI) technique., Methods and Results: Seventeen healthy adults (age = 24 ± 3 years) completed three PORH bouts over two lab visits. LSCI region of interest was a standardized 10 cm region on the dominant ventral forearm. A 5-min brachial artery occlusion period induced by inflating an arm cuff to 200 mmHg, preceded a 4-min hyperemic period. Inter- and intra-day reliability and reproducibility of cutaneous vascular conductance (LSCI flux / mean arterial pressure) were determined using intraclass correlation (ICC) and coefficient of variation (CV%). Maximal flow and area under the curve standardized to zero perfusion showed intra- and inter-day reliability (ICC > 0.70). Time to maximal flow (TMF) was not reproducible (inter-day CV = 18%). However, alternative kinetic indices such as 1-min AUC and overshoot rate-of-change (ORC), represented as a piecewise function (at 5s, 10s, 15s, and 20s into hyperemia), were reproducible (CV< 11%). Biological zero was a reliable normalization point., Conclusion: PORH measured with LSCI is a reliable assessment of microvascular function. However, TMF or its derived hyperemic velocity are not recommended for longitudinal assessment. Piecewise ORC and 1-min AUC are reliable alternatives to assess the kinetic response of PORH., Competing Interests: The authors have declared that no competing interests exist.