Optical coherence tomography (OCT) and T2 mapping are emerging clinical imaging technologies with potential to detect subsurface changes in cartilage retaining a macroscopically intact articular surface. This study tests the hypothesis that OCT correlates with magnetic resonance imaging (MRI) T2 values, and that OCT signal is sensitive to cartilage matrix degeneration. Forty-five osteochondral cores were harvested from five human tibial plateau explants after MRI T2 mapping. Cores underwent OCT imaging and were graded as follows: A, obvious birefringence; B, no birefringence; C, subsurface voids and/or irregular surface. Extracellular matrix content was determined and cores underwent histologic and polarized light microscopy (PLM) evaluation. Grade B and C cores had 25% higher superficial T2 values (p = 0.047) and 50% higher deep T2 values (p = 0.012) than grade A cores. Grade B and C cores had 36% higher glycosaminoglycan (GAG) content compared to grade A cores (p = 0.009). Histology and PLM demonstrated increased surface irregularity and structural disorganization with increasing OCT grade. OCT grade and T2 value increased with increasing collagen disorganization, suggesting that MRI T2 mapping and OCT are sensitive to changes in collagen structure. Our results demonstrate the ability of OCT and T2 mapping to detect early cartilage degeneration in clinically normal appearing cartilage.