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Effective biofilm removal from surfaces in the mouth is a clinical challenge. Cavitation bubbles generated around a dental ultrasonic scaler are being investigated as a method to remove biofilms effectively. It is not known how parameters such as surface roughness and instrument distance from biofilm affect the removal. We grew Strepotococcus sanguinis biofilms on coverslips and titanium discs with varying surface roughness (between 0.02–3.15 μm). Experimental studies were carried out for the biofilm removal using high speed imaging and image analysis to calculate the area of biofilm removed at varying ultrasonic scaler standoff distances from the biofilm. We found that surface roughness up to 2 μm does not adversely affect biofilm removal but a surface roughness of 3 μm caused less biofilm removal. The standoff distance also has different effects depending on the surface roughness but overall a distance of 1 mm is just as effective as a distance of 0.5 mm. The results show significant biofilm removal due to an ultrasonic scaler tip operating for only 2s versus 15-60s in previous studies. The technique developed for high speed imaging and image analysis of biofilm removal can be used to investigate physical biofilm disruption from biomaterial surfaces in other fields.