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CDE0211

pores and that this is a cause of the transient sen- sitivity experienced by some people when they whiten their teeth. Recently, some companies have added ACP to their formulations. Claims have been made that the addition of ACP reduces tooth sensitivity by decreasing the size of these pores.16 In addition, by fillingminordefectswithintheenamel,theaddition of ACP creates an enamel surface that is smoother and more lustrous. However, in an invitro study on bovineincisors,nosupportinginfluenceoffluoride- containing bleaching gels on remineralisation was observed.17 Thepurposeofthisstudywastoevaluateenamel subsurface structure following application of two bleaching agents to extracted incisor teeth using confocal microscopy. _Materials and method A flattened area (≈ 4 x 5 mm) was created on the labialsurfaceofextractedcentralandlateralincisor teeth(n=10)byserialpolishingwithSiCsandpaper of up to 1,200 grit (Figs. 1a & b). It is true that confo- calmicroscopyholdsgreateradvantagesforsamples thatcannotbepolishedtoaflatsurface.Inourcase, the flattened surface helped in orienting the area of interest as a plane-parallel object held perpen- dicular to the optic axis for sharper image. The teeth were ultrasonically cleaned with dis- tilled water to remove debris. A piece of water- resistant tape was applied to one-half of the tooth and burnished (Fig. 1c). The edge of the tape was sealed by painting it with a transparent nail varnish (Fig. 1d). The exposed area was randomly assigned to one of two groups. The first group (ACP group) was treated with Nite White Excel 3 with ACP (Discus Dental). The second group (OP group) was treated with Opalescence PF 10 % (Ultradent). The untreated control for both groups was the area un- derneath the tape, allowing each tooth to serve as internal control. The composition of the whitening products is described in Table I. Bleaching agents for both groups were applied for seven hours per day for 14 days. The materials wereappliedwithamicro-brush,takingcaretolimit theapplicationtotheappropriateareaonly(Fig.1e). Once the application of bleaching agent had been completed, the teeth were placed inside a plastic box, which acted as a moisture barrier, while keep- ing the bleaching agent undisturbed throughout the procedure. Aftereachdailyapplication,expendedbleaching materialwasfirstremovedwithacleanmicro-brush. The area was then cleansed with water and blotted dry. Finally, the teeth were rinsed with air-water spray for 20 seconds. A cycling treatment method- ology was employed. While not being actively treated, the teeth were stored in artificial saliva (Saliva Substitute, Roxane Laboratories). Up to the point of the microscopic examination, the tape covering the control group area remained in place (Fig. 1f). Before confocal microscopic evaluation, the teethweresubmergedinTexasReddyewithDextran for 24 hours. A two-photon microscope (LSM 510 Meta,CarlZeiss)wasusedtodetectthefluorescence under an Argon 488 laser (Fig. 2a). Each area was examined up to a depth of 100 µm. The flattened enamelsurfacewasorientatedperpendiculartothe laser beam with sticky wax and the whole tooth sample was placed under water contained in a Petri dish (Fig. 2b). Samples were viewed with a 5X/0.16 objective, focusing approximately 5 to 100 µm be- low the surface. Images were relayed to a computer monitor for viewing. Additional images were made Figs. 2a & b_Experimental set up for confocal microscopy (a). Flattened surface of the incisor orientated perpendicular to the laser (b). Tooth sample immersed in distilled water. Figs. 3a & b_OP control area 6 µm subsurface (a). OP-treated 10 µm subsurface (b). Arrow indicates subsurface crack. I 23 research _ bleaching I cosmeticdentistry 2_2011 Fig. 2bFig. 2a Fig. 3a Fig. 3b