Summary Different cell types, each with unique composition, properties, and functions, are essential to sustain life in an organism. Bulk analysis of such heterogeneous mixtures of cells does not distinguish the characteristics of a given cell type or differences between cells within the same type, missing information that is key to understanding molecular mechanisms foundational to the rational design of therapies for disease and ailments. The goal of this proposal is to develop single-cell and imaging methods using metal-bearing probes to monitor metabolic processes in live cells such as siRNA delivery, protein synthesis, and post-translational modification. The cells will then be fixed and processed for subsequent detection of protein and mRNA markers. There are three aims: (1) Delivering molecular probes to viable cells to explore molecular pathways in single cells by CyTOF. (2) Barcoding prenylation probes to explore molecular pathways in mixed samples of single cells. (3) Exploring the effect of aging in murine primary cells by CyTOF and murine tissue by MIBI-ToF. The proposed work will expand the use of multi- parametric capabilities of mass cytometry (i.e. cytometry by time-of-flight, CyTOF, and multiplexed ion beam imaging by time-of-flight, MIBI-ToF) in the biotechnological and biomedical fields. The development of these methodologies will explore the causal relationship between autophagy and senescence in murine liver models, exploring the hypothesis that the mevalonate pathway provides a link between the two. The ultimate goal is to understand the relationship between autophagy, mevalonate, and senescence pathways in aging and inform emerging treatments to counteract the detrimental effects of senescent cells.