This paper describes the use of confocal microscopy as a new method for in situ analysis of the apatite layer which forms on bioactive substrates after immersion in a simulated body fluid (SBF). Conventional methods such as secondary electron microscopy (SEM) require drying and coating of the samples. This approach causes damage to the structure of the apatite layer of which a common example is surface cracking. With confocal microscopy samples are examined in situ and build a unique optical sectioning of the samples. As shown in this paper, using these sections, a clear image can be constructed of the reactive layer which forms on a sample both in horizontal and vertical cross-sections.