Purpose: To compare the antimicrobial activity of the tricalcium silicate-based Biodentine (BD) and mineral trioxide aggregate (MTA)-Angelus cement with the aid of agar diffusion test. Materials and Methods: Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, and Enterococcus faecium were inoculated in the Brucella liquid medium and were incubated at 37°C for 24 h. Thereafter, 100 >μl of the liquid culture of bacteria inoculated in the Mueller-Hinton agar with spread plate technique. Petri plates were dried in room temperature. For every microorganism, 3 petri plates were prepared (12 in total). In the medium, in every petri plate, 2 holes with 5 mm diameter and 2 mm depth were made. Afterward, BD and MTA-Angelus were filled into these holes under aseptic conditions according to the instructions of the manufacturing company. Then, the plates were kept in the incubator at 37°C for 24 h, and the diameters of the inhibition zones were measured with a digital caliper. Results: Inhibition zones formed by BD against E. coli and S. aureus were significantly larger than the zones formed by MTA-Angelus (P < 0.05). However, the inhibition zones formed by MTA-Angelus against P. aureus and E. faecium were larger than the zones formed by BD (P < 0.05). Conclusion: Within the limits of the present study, tricalcium silicate-based MTA-Angelus and BD have antimicrobial activity against E. coli, S. aureus, P. aureus, and E. faecium.