Indiana University-Purdue University Indianapolis (IUPUI) / Objective: The objectives of this in-vitro study were: 1) to evaluate the ability of Near-Infrared Digital Imaging Transillumination (NIDIT) to detect non-cavitated approximal caries lesions; and 2) to compare the performance among NIDIT, Digital Radiography (DR), Digital Imaging Fiber-Optic Trans-Illumination (DIFOTI) and International Caries Detection and Assessment System (ICDAS). Methods: Thirty human extracted premolars were selected. The approximal surface status ranged from sound to surfaces with non-cavitated caries lesions into the outer one-third of the dentin. Lesion depth was determined by micro-computed tomography (μ-CT) and used as a gold standard. Teeth were mounted in a custom-made device to simulate approximal contact. ICDAS, DR, DIFOTI and NIDIT examinations were performed and repeated by three trained and calibrated examiners. Sensitivity, specificity, area under ROC curve (Az), inter- and intra-class correlation coefficients (ICCs) of each method, and correlation among the methods were determined. Results: ICCs for intra-/inter-examiner agreement were almost perfect for DIFOTI (0.85/0.83), substantial for ICDAS (0.79/0.72) and NIDIT (0.69/0.64), and moderate for DR (0.52/0.48). Sensitivity/specificity for DIFOTI, ICDAS, DR, and NIDIT were 0.91/0.69, 0.89/0.83, 0.50/0.64, and 0.68/0.93, respectively. Az of DR (0.61) was significantly lower than that of DIFOTI (0.91, p = 0.002) and ICDAS (0.90, p = 0.005), but was not significantly different from NIDIT (0.81, p = 0.052). DIFOTI, ICDAS, and NIDIT were not significantly different from each other (p > 0.13). Spearman correlation coefficients for DIFOTI (0.79, p < 0.001), ICDAS (0.74, p < 0.001), and NIDIT (0.65, p < 0.001) demonstrated a moderate association with μ-CT, while that of DR suggested no association (0.19, p = 0.289). Conclusion: Within the limitations of this in-vitro study, NIDIT system demonstrated a potential for early approximal caries detection. ICDAS, DIFOTI, and NIDIT were superior to DR in terms of validity and reliability.