Bioimaging enables the spatiotemporal visualization of biological processes at various scales empowered by a range of different imaging modalities and contrast agents. Upconversion nanoparticles (UCNPs) represent a distinct type of such contrast agents with the potential to transform bioimaging due to their unique optical properties and functional design flexibilities. This review explores and discusses the opportunities, challenges, and limitations that UCNPs exhibit as bioimaging probes and highlights applications with spatial dimensions ranging from the single nanoparticle level to cellular, tissue, and whole animal imaging. Recent advancements in bioimaging applications enabled by UCNPs, including super‐resolution techniques and multimodal imaging methods are summarized, and a perspective on the future potential of UCNP‐based technologies in bioimaging research and clinical translation is provided. This review may provide a valuable resource for researchers interested in exploring and applying UCNP‐based bioimaging technologies.