Background: Hands are the most common medium for initiation and spread of infection in clinics. Hand hygiene is the simplest and most economic method for reducing the healthcare-associated infections (HCAIs). Aim: The aim of the present study was to compare the efficacy of hand sanitizer, liquid soap, and their combination for reducing the microbial colonies on hands. Materials and methods: The study was a single-blinded randomized controlled trial executed in a dental college on 90 participants, which were randomly assigned into three intervention groups of 30 each. The participants were instructed to contaminate their hands followed by a hand hygiene protocol for the liquid hand-wash group, the alcohol-based hand sanitizer (ABHS) group, and the combination group according to WHO guidelines. The swabs were collected pre- and post-intervention and mean colony-forming units were determined for each group. Results: Median percentage reduction was highest for the combination group (100%), followed by the ABHS group (94.29%) and lowest for the liquid soap (92.31%). This difference in the amount of colony-forming units (CFU) among all the groups was nonsignificant (p = 0.114). Conclusion: The results of the study show that there was no significant difference in hand disinfection potential among hand sanitizer, liquid soap, or their combination. All the three interventions were equally effective with the reduction of total bacterial contamination from participants’ hands.