Matrix metalloproteinases are a family of zinc-dependent proteinases which are involved in the breakdown and remodeling of extracellular matrix. As children grow and adolescents reach pubescence, their bodies undergo changes that require age-related morphogenesis of the extracellular matrix, possibly requiring unique patterns of matrix metalloproteinase (MMP) expression during periods of rapid tissue growth (i.e., childhood) or accelerated tissue remodeling and expansion (i.e., adolescence). Therefore, we have characterized age-specific and gender-specific differences in circulating concentrations of MMPs (specifically MMP-1, -2, -3, -8 and -9) in 189 serum samples obtained from healthy subjects, aged 2−18 years. MMP concentrations were measured using Fluorokine® MultiAnalyte Profiling kits and a Luminex® Bioanalyzer, as well as by commercial ELISA. Serum levels of MMP-1, -2, -3, -8, and -9 in healthy pediatric subjects represent log-normal distributions. MMP-2 was significantly negatively correlated with age (r=−0.29; p<0.001), while MMP-3 was significantly positively correlated with age (r=0.38; p<0.001). Although plasma, not serum, is considered the appropriate blood sample for measurement of MMP-8 and -9, serum levels of MMP-8 and -9 were also found to be highly positively correlated with each other (r=0.76; p<0.01). MMP results obtained by Fluorokine® MultiAnalyte Profiling methods correlated well with conventional ELISA methods and use of this technology provided several advantages over ELISA.