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Summary: The isolated perfused kidney is a classic ex vivo preparation for studying renal physiology in general and vascular function. Here, we present a protocol for assessing myogenic tone in isolated mouse kidneys as well as vasodilatory and vasoconstrictive responses, expressed as perfusion pressure. We describe steps for pre-operative preparation, kidney and renal artery isolation, and connection of renal artery with glass cannula. We then detail how to measure pressure changes in perfused kidneys and the myogenic tone.For complete details on the use and execution of this protocol, please refer to Cui et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.