Inhibition of matrix metalloproteinase (MMP) activity is expected to increase the long-term stability and durability of resin-based restorations by preventing degradation of denuded collagen fibrils in the hybrid layer. Endogenous MMP-8, released by adhesive procedures, can hydrolyse collagen fibrils at resin/dentin interfaces compromising adhesive bond durability. In this study, occlusal surfaces of n=4 extracted non-carious human molar teeth (extraction, stored -80°C) were removed using a low-speed diamond saw. 2x4x2mm cavity was prepared on the dentinal surface, leaving surrounding dentinal walls of 1mm thickness. Cavity walls were then acid-etched with 38% phosphoric acid, pulverized using a liquid nitrogen analytic mill, demineralized in 1% phosphoric acid, suspended in extraction buffer (100mM Tris- HCl pH 7.6, 0.1mM ZnCl2, 100mM CaCl2, 200mM NaCl, 1% Triton X-100) and centrifugally concentrated. The results of our study showed that an average of 0.238pg MMP-8/mg dentin is present as measured by ELISA. This study is the first of its kind to identify endogenous levels of MMP-8 present in a single-tooth dentin sample. This information will contribute to the design of specific, clinical inhibition of endogenous MMP activity in dentinal cavity walls to prolong the longevity of resin-based restorations.