The confocal laser scanning microscopy (CLSM) system has been widely used to analyze early carious lesions with fluorescent ligands in dental imaging. This system can be used to examine the physiological condition of cellular colonization in the tooth structure. However, the undesirable noise in CLSM images hinders accurate activity assessment of early carious lesions. To address this limitation, a total variation (TV)-based noise reduction algorithm with good edge preservation was developed, and its applicability to medical tooth specimen images obtained with CLSM was verified. To evaluate the imaging performance, the proposed algorithm was compared with conventional filtering methods in terms of the normalized noise power spectrum, contrast-to-noise ratio, and coefficient of variation. The results indicate that the proposed algorithm achieved better noise performance and fine-detail preservation, in comparison with the conventional methods.