Secondary-ion mass spectrometry (SIMS) is based on the acceleration of high-energy primary ions onto a target. Secondary electrons, neutrals and ions are emitted from the target, reflecting its chemical composition. This enables simultaneous analysis and localization of target molecules, giving valuable information that is difficult or impossible to obtain with other analytical methods. The secondary ions can be extracted and detected by any type of mass analyzer. SIMS is unique in its ability to detect several target molecules simultaneously in small samples and to image their localization at subcellular resolution. The recent development of bioimaging SIMS opens up new possibilities in biotechnology and biological research with applications in biomedicine and pathology. The current development of this technique has the potential to become as important for biotechnology as the advent of the electron microscope, confocal microscope or in situ hybridization.