## 2 Materials and Methods  
In this study, a total of 18 drills of zirconia (Dental Tech, Misinto, Milan) and 18 steel drills were used (Dental Tech, Misinto, Milan) (Figure 1).  
Only 2 mm diameter helical conical drills were evaluated in order to analyze their response to disinfecting and cleansing agents, as well as autoclave sterilization. The steel drills presented the following chemical composition: 0.2% sulfur, 0.2% carbon, 0.6% silicon, 0.8% nickel, 1.2% molybdenum, 1.6% manganese, 16% chromium, 22% carbon, and balancing iron. The chemical composition shows the typical characteristics of a stainless-steel alloy. The drills studied were all externally irrigated and had three black laser markings with the function of guiding the surgeon in controlling the depth during osteotomy. In this investigation, chemical agents such as glutaraldehyde 2% (Dimexid, Amedics Ferrara, Italy) and hydrogen peroxide 6% (Sanibios, Noda, Balerna- Switzerland), were taken into consideration, and the effect of sterilization cycles was also observed. The drills were not subjected to the preparation of implant sites, but were immersed for 10 minutes in human blood taken from voluntary subjects and then underwent cycles of immersion in disinfectants without being subjected to autoclaving (approved by Inter Institutional Ethics Committee of UNINGA, No. 72105917.5.0000.5220). Before immersion, decontamination, or sterilization, the drills were rinsed in running water and scrubbed with a toothbrush made of nylon bristles in order to remove gross foreign material (e.g., organic material, clot). A total of 18 steel drills and 18 zirconia drills were used and allotted to 6 groups of 6 drills, with 6 in steel and 6 in zirconia for each group. Then, implant drills were assessed before and after the disinfection by Scanning Electron Microscopy (SEM). A disc of carborundum was used to separate the blades from the grips of the drills to be observed by SEM. Ten flat areas of 200 \(\upmu\)m to 150 \(\upmu\)m in diameter were evaluated for each drill and an image in JPEG format was created (Figure 2).  
Figure 1: Macroscopic aspect of the steel drill (left) and zirconia drill (right) used in the present study.  
The drills were divided into the following groups:  
Group 1: New non-sterilized Steel drill; Steel drills subjected to 50 cycles of 20 min sterilization immersion in glutaraldehyde;  
Group 2: Steel drills subjected to 50 immersion cycles of 20 min immersion cleansing in hydrogen peroxide;  
Group 3: Steel drills subjected to 50 cycles of heat sterilization in an autoclave at 134\({}^{\circ}\) C for 35 min at a pressure of 1.1 bar; New non-sterilized Zirconia drills;  
Group 4: Zirconia drills subjected to 50 cycles of 20 min immersion sterilization in glutaraldehyde;  
Group 5: Zirconia drills subjected to 50 immersion cycles of 20 min for cleansing in hydrogen peroxide;  
Group 6: Zirconia drills subjected to 50 cycles of heat sterilization in an autoclave at 134\({}^{\circ}\) C for 35 min at a pressure of 1.1 bar (A-17 PLUS, ANTHOS, Imola, Italy).  
All the solutions were poured into a plastic container in order to avoid interference between the liquid and metal. In accordance with the Italian legislation DMS of 28/9/90, the drills were immersed for 20 min in a liquid chemical sterilant. After 50 cycles of decontamination and cold sterilization immersion, the drills were rinsed in running water and then in demineralized water. They were then observed by a transmission electron microscope. The method of analysis followed the methodology proposed by [PERSON] et al. [5].  
Previously calibrated examiners compared the micrographs before and after 50 cycles of disinfection and sterilization, respectively, with those obtained for the control group and assessed the percentage of surface covered by damage.