Background And Objectives : To compare in-vitro, the cleaning effectiveness of pulp space preparation under Scanning electron microscope (SEM) using four different NiTi rotary instrumentation systems – Quantec SC, K3 Endo, RaCe and HERO 642. Materials and Method : Forty-eight freshly extracted palatal roots of human maxillary first molar teeth were procured, stored and surfaced adhering to infection control protocols in the laboratory as prescribed by Centre for Disease Control Global. Random sampling of specimens into four groups of ten teeth each and one group of eight teeth was done. Groups I, II, III & IV served as experimental with Group V as control. Access cavities were prepared in all groups and working length was determined only in experimental groups. Control group remained non-instrumented. Cleaning and shaping was done for experimental groups only. Teeth were prepared to # 40 master apical file using crown down technique. The experimental groups: I. Teeth with Pulp space instrumented with Quantec SC. II. Teeth with Pulp space instrumented with K3 Endo. III. Teeth with Pulp space instrumented with RaCe. IV. Teeth with Pulp space instrumented with HERO 642. V. Control group: Teeth with pulp space not instrumented. Thereafter all teeth were longitudinally sectioned buccolingually and examined under SEM at 20.0 kV and 500 X magnifications in the middle third of the canal. Debris was defined as dentin chips, pulp remnants, and particles loosely attached to the root canal wall. Analysis of SEM images was performed using following five score index:-Numerical Evaluation (Scoring System): Score 1 – Clean root canal walls, only few small debris particles. Score 2 - Few small agglomeration of debris. Score 3- Many agglomerations of debris covering less than 50% of the root canal wall. Score 4- More than 50% of the root canal wall covered with debris. Score 5- Complete or nearly complete root canal walls covered with debris. Statistical Analysis : Kruskal Wallis test was used to determine whether a statistical difference existed amongst the four group of instruments, with P < 0.001, considered to be statistically very highly significant. Whitney Mann -U test for individual intergroup comparison and “CHI-square” test was used to detect frequency of individual debris score in each tested groups followed by Kendell’s tau-b test to measure observer agreement. Results : Teeth with pulp space instrumented with RaCe system exhibited maximum canal cleanliness compared to the other experimental groups. The differences observed within the groups were statistically very highly significant (p<0.001). Conclusion : Post pulp space preparation with four different NiTi rotary systems, statistically very highly significant differences in canal wall cleanliness at the midroot level was achieved. Under the conditions of this study, best canal cleaning efficacy was achieved by RaCe, followed by HERO 642, K3 Endo and Quantec SC.