A randomized controlled clinical study was set up to assess caries removal following tunnel preparation (test group) and class II cavity preparation (control group). Sixty approximal lesions in adult posterior teeth, visible in the outer third of dentine on bite–wing radiographs, were referred for operative treatment. Initial dentine samples were taken on entry to the lesions in both groups. Following cavity preparation dentine samples were taken from beneath the marginal ridge in the tunnel group and at the cervical floor in both groups. Microbiological analysis was carried out to establish the level of infection of the dentine. The bacterial counts were high on entry to the lesions with a median log10 (CFU + 1) per sample of 3.07 (±1.24). Following cavity preparation bacterial counts at the cervical floor were significantly reduced in both amalgam and tunnel groups (p<0.00001). In the tunnel group, however, slightly increased bacterial counts were found beneath the marginal ridge compared to the cervical floor (p<0.01).