Background/Objectives: Endodontic success in maxillary first molars is often complicated by their complex anatomy, increasing the risk of procedural errors in "danger zones" where dentine is thin. While global data on root morphology exists, population-specific information for South Africans is lacking. This study aims to quantify root dentine thickness in a Black South African sample using a novel software program to enhance accuracy for preoperative planning and improved clinical outcomes. Methods: Micro-CT scans of 97 maxillary first molars (57 individuals) were analyzed. Den-tine thickness was measured at 0.1 mm intervals using a novel surface-to-interface software tool, which enabled automated, high-precision quantification of complex curved geometries. Data were standardized into 1 mm segments for analysis. Reliability trials confirmed high precision (within 0.0001 mm). Results: Buccal and lingual surfaces of mesiobuccal and distobuccal roots were consistently thicker than mesial and distal surfaces. All roots showed progressive thinning toward the apical third. Conversely, palatal roots exhibited an opposing pattern with mesial and distal surfaces that were thicker than buccal and lingual aspects. Age correlations were not statistically significant. Conclusions: The study identifies specific anatomical patterns in root dentine thickness within a South African sample. The thinner mesial and distal walls of buccal roots represent critical "danger zones" for clinicians. The novel software proved highly effective for precise morphometric mapping and offers a versatile framework for broader endodontic research. These findings provide essential data for optimizing endodontic treatment strategies and the development of population-specific dental instruments.