Introduction: The study aimed to compare the efficacy of a single application of plain nano-hydroxyapatite (n-HA) paste in reducing dentine hypersensitivity. Materials and Methods: Sixty-three patients were recruited to participate in the study and randomized to be allocated into three groups (n-21): (1) Group 1 – n-HA paste, (2) Group 2 – fluoride paste, and (3) Group 3 – placebo. The patient's response to dentin hypersensitivity (DH) was evaluated at baseline (T0), immediately after application (T1), and after 1 week (T2). Tactile and cold air sensitivity (CAS) stimulus tests and visual analog scale subjective tests were used to measure the patient's response to DH. The data analysis was performed using the Statistical Package for the Social Sciences statistical software. Results: All three groups showed a statistically significant reduction in tactile sensitivity (TS), cold air sensitivity, and visual analog scale scores from T0–T1 and T0–T2 (P 0.005) except for Group 1, which showed a significant reduction (P = 0.033) in visual analog scale scores from T1–T2. However, the intergroup comparison at T2 demonstrated a significant difference in tactile scores (P = 0.005), CAS (P Conclusion: n-HA paste was the most effective desensitizing paste compared to fluoride and placebo pastes. The single application of the n-HA paste demonstrated a significant reduction in visual analog scale scores after 1-week.