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Molecular composition is intricately intertwined with cellular function, and elucidation of this relationship is essential for understanding life processes and developing next-generational therapeutics. Technological innovations in capillary electrophoresis (CE) and liquid chromatography (LC) mass spectrometry (MS) provide previously unavailable insights into cellular biochemistry by allowing for the unbiased detection and quantification of molecules with high specificity. This chapter presents our validated protocols integrating ultrasensitive MS with classical tools of cell, developmental, and neurobiology to assess the biological function of important biomolecules. We use CE and LC MS to measure hundreds of metabolites and thousands of proteins in single cells or limited populations of tissues in chordate embryos and mammalian neurons, revealing molecular heterogeneity between identified cells. By pairing microinjection and optical microscopy, we demonstrate cell lineage tracing and testing the roles the dysregulated molecules play in the formation and maintenance of cell heterogeneity and tissue specification in frog embryos (Xenopus laevis). Electrophysiology extends our workflows to characterizing neuronal activity in sections of mammalian brain tissues. The information obtained from these studies mutually strengthen chemistry and biology and highlight the importance of interdisciplinary research to advance basic knowledge and translational applications forward.
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