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The use of high-density microelectrode arrays (HD-MEAs) provides a promising approach for electrophysiological studies targeted at understanding of brain functions, pro ling of neurodegenerative diseases, and drug screening. Here we present the protocol for the preparation of various biological samples for the recording of extracellular signals using HD-MEA, including primary cortical neurons, induced pluripotent stem cells (iPSCs)-derived neurons, rodent brain slices, retina, and iPSC-derived neuronal spheroids.