Cyborg organoids integrated with stretchable nanoelectronics can be functionally mapped during development.

Organoids are in vitro miniaturized cellular models of organs that offer opportunities for studying organ development, disease mechanisms and drug screening. Understanding the complex processes governing organoid development and function requires methods suitable for the continuous, long-term monitoring of cell activities (for example, electrophysiological and mechanical activity) at single-cell resolution throughout the entire three-dimensional (3D) structure. Cyborg organoid technology addresses this need by seamlessly integrating stretchable mesh nanoelectronics with tissue-like properties, such as tissue-level flexibility, subcellular feature size and mesh-like networks, into 3D organoids through a 2D-to-3D tissue reconfiguration process during organogenesis. This approach enables longitudinal, tissue-wide, single-cell functional mapping, thereby overcoming the limitations of existing techniques including recording duration, spatial coverage, and the ability to maintain stable contact with the tissue during organoid development. This protocol describes the fabrication and characterization of stretchable mesh nanoelectronics, their electrical performance, their integration with organoids and the acquisition of long-term functional organoid activity requiring multimodal data analysis techniques. Cyborg organoid technology represents a transformative tool for investigating organoid development and function, with potential for improving in vitro disease models, drug screening and personalized medicine. The procedure is suitable for users within a multidisciplinary team with expertise in stem cell biology, tissue engineering, nanoelectronics fabrication, electrophysiology and data science.