Measuring single-cell water content non invasively and with high precision

SUMMARY Water is the medium in which biochemistry operates. The amount of water inside a cell defines the concentration of all biomolecules and the degree of molecular crowding. Consequently, water content is predicted to influence virtually all cell functions from immune surveillance to stem cell renewal in vivo. There are also severe genetic diseases caused by mutated water channels on cell membranes. Despite the fundamental role of cellular water content in cell physiology and diseases, there are no direct and non-invasive methods that measure how much water a single cell or a cluster of cells, such as an organoid, contains. This lack of water content measurements has prevented us from understanding cell physiology in normal and disease states, and from discovering drugs that can modulate cellular water content in diseases where water content is perturbed. Here, we propose to develop a new method that will directly, non-invasively and precisely measure the absolute and fractional (v/v) water content of a single cell or an organoid. This method will enable the long-term monitoring of the same cell or organoid during growth, differentiation or drug exposure. To achieve this, we will integrate total volume and dry volume measurements obtained using two independent approaches that we have previously developed. Our proposed method will enable water content measurements in complex and biomedically relevant samples, such as immune cells and organoids, with high throughput and in conjunction with detection of fluorescent markers. This method will enable novel basic and biomedical research that will increase our understanding of water content regulation in health and disease, and provide a new platform for drug discovery.