Despite its advantages, PALM has some limitations. One challenge is the need for specialized fluorescent probes that can be photoactivated or photoswitched. These probes can sometimes be toxic to cells or may not label all targets efficiently. Additionally, PALM requires sophisticated data analysis algorithms to accurately localize each molecule, which can be computationally intensive. Temporal resolution is another limitation, as the sequential activation process can be time-consuming, making it difficult to capture very fast biological events.
