Measuring the Senescence-Associated Secretory Phenotype.

Cellular senescence is a fundamental hallmark of aging, contributing to tissue dysfunction and chronic disease through the senescence-associated secretory phenotype (SASP). The SASP encompasses a diverse and dynamic collection of secreted cytokines, chemokines, growth factors, and proteases that vary depending on cell type, senescence trigger, and microenvironmental context. Accurate quantification of SASP components is critical to understanding the mechanisms linking senescence to pathology and for advancing senotherapeutic strategies. However, measuring the SASP presents significant technical and biological challenges due to its complexity, heterogeneity, and context dependence. This review provides a comprehensive overview of the principal methodologies used to measure SASP components across different biological levels-transcriptional, translational, and functional-and sample types, including cell cultures, tissues, and systemic fluids. We discuss the advantages and limitations of widely used RNA-level techniques (e.g., qRT-PCR, RNA sequencing, in situ hybridization), protein-level assays (e.g., ELISA, Western blotting, mass spectrometry, Luminex, MSD), and spatial detection methods (e.g., immunohistochemistry, immunofluorescence). By organizing current SASP detection strategies by molecular level and sample source, this review highlights the importance of multiparametric approaches to capture the full spectrum of senescent cell activity. We also identify key methodological gaps and propose directions for refining SASP biomarker discovery in aging and disease research.