Reactive oxygen species (ROS) are a family of molecules continuously generated, transformed, and consumed in all living organisms. While traditionally viewed as harmful, emerging data show that ROS can also contribute to physiology and increased fitness. This Perspective discusses the factors that determine whether ROS act as signals or stress agents, highlighting the importance of understanding their chemistry in biology. ROS are involved in various biological processes, including redox modifications of biomolecules, and their overproduction leads to oxidative stress. However, organisms have evolved mechanisms to harness ROS for essential physiological processes. ROS are produced in various cellular locations, including mitochondria, the endoplasmic reticulum, and cell membranes. ROS can mediate redox modifications on proteins, lipids, and DNA, and their reactivity is influenced by their chemical properties and the proximity of target molecules. The study of ROS chemistry is crucial for understanding their diverse roles in biology. Chemical tools are being developed to detect and study ROS, enabling a better understanding of their functions in physiological and pathological processes. The regulation of ROS production and signaling is essential for maintaining cellular homeostasis and health. This review highlights the importance of ROS in various physiological processes, including cell migration, circadian rhythm, stem cell proliferation, and neurogenesis. The development of new chemical tools and methods for studying ROS is essential for advancing our understanding of their roles in biology.Reactive oxygen species (ROS) are a family of molecules continuously generated, transformed, and consumed in all living organisms. While traditionally viewed as harmful, emerging data show that ROS can also contribute to physiology and increased fitness. This Perspective discusses the factors that determine whether ROS act as signals or stress agents, highlighting the importance of understanding their chemistry in biology. ROS are involved in various biological processes, including redox modifications of biomolecules, and their overproduction leads to oxidative stress. However, organisms have evolved mechanisms to harness ROS for essential physiological processes. ROS are produced in various cellular locations, including mitochondria, the endoplasmic reticulum, and cell membranes. ROS can mediate redox modifications on proteins, lipids, and DNA, and their reactivity is influenced by their chemical properties and the proximity of target molecules. The study of ROS chemistry is crucial for understanding their diverse roles in biology. Chemical tools are being developed to detect and study ROS, enabling a better understanding of their functions in physiological and pathological processes. The regulation of ROS production and signaling is essential for maintaining cellular homeostasis and health. This review highlights the importance of ROS in various physiological processes, including cell migration, circadian rhythm, stem cell proliferation, and neurogenesis. The development of new chemical tools and methods for studying ROS is essential for advancing our understanding of their roles in biology.