## 2 Materials and Methods  
The present in vitro study compared IH3 DBs with three different commercial 0.022'' \(\times\) 0.028'' brackets: ceramic Clear Damon bracket (Ormco Corporation, Glendora, CA, USA), ceramic Synovate C bracket (Genesis Orthodontics, Castle Rock, CO, USA) and composite Crystal bracket (Genesis Orthodontics, Castle Rock, CO, USA).  
The IH3 DBs, with a slot size of 0.022 \(\times\) 0.028 inches and an MBT prescription, were printed using the desktop SprintRay PRO 95 printer (SprintRay Inc., Los Angeles, CA, USA), which relies on DLP (Digital Light Projection) technology. [21] The resin for 3D-printing of temporary crowns BEGO Verseosmile Temp (BEGO GmbH & Co. KG, 28359 Bremen, Germany) certified IIa with a flexural strength of \(\pm\)80 Mpa and colour A2 on the Vita scale calibrated for the SprintRay Pro 95 printer was used and a print-layer thickness of 50 \(\upmu\)m on the \(Z\) axis was adopted.  
There were ten brackets for each type to a resin bar with cyanoacrylate adhesive, for a total of four bars, and a first colour measurement (T0) was taken by a spectrophotometer (Vita Easyshade Compact, Bad Sackingen, Germany), which was calibrated by the white light spectrum before each measurement. The digital spectrophotometer provides emission of all tooth colours in the two VITA Classical A1-D4 scale and VITA System 3D-master colour standards scale, and in tooth brightness according to the American Dental Association scale. Colour values were recorded for each of the ten brackets for each bar and subsequently averaged.  
In order to test the repeatability of the analysis, two measurements were taken under the same initial conditions.  
Subsequently, the four bars were immersed individually in a plastic box with artificial saliva, Oral Balance (Biotene Oral Balance, Biopharm Sas, Peschiera Borromeo, Italia) and brown food colouring [18]. The latter solution was prepared by mixing 1.5 g of Brooke Bondtea with 150 mL of boiling water. The solution was cooled to 37 \({}^{\circ}\)C (room temperature) and subsequently was filtered out using a filter paper to eliminate any residues, thus obtaining a homogenous solution [22].  
A constant temperature was maintained by simulating the condition of the oral cavity at 37 \(\pm\) 1 \({}^{\circ}\)C. The brushing cycle of about 65 min corresponding to two years of orthodontic treatment was calculated to be equivalent to 8.5 years of brushing, based on a brushing time of 120 s twice daily for all teeth. For the brushing simulation, a manual toothbrush connected to an electric motor (24 vdc 60w 250 rpm brush type electric machinery reducer motor) and 16,250 cycles were performed setting the cleaning force to 2.5 N [23] (Figure 1).  
At the end of the brushing procedure, the colour values of all brackets were re-measured with the same spectrophotometer by the same operator (T1) (L.L.).  
In order to perform the statistical analysis, it was necessary to convert the results of the VITA scale and Master scale into absolute values. Firstly, the Master scale's values were transformed into the VITA scale's colours [24] so that there were two VITA scale values for each bracket analysed. Then, both values were transformed into absolute values by means of the conversion table (Figure 2) [25].