## 4 Discussion  
TotalFill BC Sealer(r) is one of the new calcium silicate-based sealers that has appeared on the European market. In Canada it is called iRoot BC Sealer, and in the USA, EndoSequence BC Sealer [14]. This bioceramic sealer has been reported by several studies because of its favorable properties as a root canal filling material, including good sealing ability [15], biocompatibility [16], antibacterial activity [15], and dentine adhesive properties [17]. It has proven to have higher tubular penetration data compared to other bioceramic sealers [18], and even in comparison with the resin-based sealer AH Plus [19; 20]. In the present study, TotalFill BC Sealer(r) was evaluated, and regardless of the fluorescent staining method evaluated, TotalFill BC Sealer(r) demonstrated, in most of the images acquired with CLSM, that it can infiltrate the dental tubules. One of the performance measures of endodontic sealers is its ability to penetrate the dentinal tubules. According to [PERSON] et al. [21], there are no correlations between sealer penetration and the sealing of filling materials. Nevertheless, it has been shown that the penetration of sealers into the dentinal tubule forms a physical barrier [22] that improves the retention of the filling material [23], and residual microorganisms are encapsulated within the canal [24]. To evaluate this, many authors have used Rhodamine B as a marker in their methodology to visualize the sealer inside the tubules [19; 20]. Rhodamine B has a strong affinity for humidity and less affinity for the calcium in the sealer composition [8]. As a result, it can separate from its mixture with the sealer, and emit sealer-independent fluorescence, showing a deeper penetration into the dentine tubules [8]. Thus, we found it necessary to check the predictability of Rhodamine B and ensure that, in the present study, the method of evaluating tubular infiltration with this stain is the most appropriate. Fluo-3 is a more appropriate fluorescent stain for tracking calcium ions under CLSM because in the presence of calcium ions, it increases the intensity of its fluorescence [25]. However, Fluo-3 cannot measure the number of calcium ions in the tooth structure [8]. Consequently, the fluorescence produced in CSLM images acquired in this study may be related absolutely to the sealer. With its unique property of generating light in response to the specific wave range of the argon laser (488 nm), Fluo-3 can be used to detect the presence of calcium silicate-based sealers under CLSM [26]. In this study, the results indicated that, in terms of maximum penetration and the percentage of sealer penetration (%), Rhodamine B had higher values throughout, demonstrating significant differences with respect to Fluo-3 when evaluating the maximum penetration at the apical level. Apart from this, it is important to emphasize that the interquartile range (IQR) was always higher in the Rhodamine B group, particularly at these apical levels. This shows the presence of greatly \begin{table}
\begin{tabular}{c c c c} \hline \hline  & & \multicolumn{2}{c}{**Interquartile Range (IQR)**} \\ \cline{3-4} Percentage of tubular penetration & & Fluo-3 & Rhodamine B \\ \cline{2-4} Maximum penetration & & Apiel & 17.8 & 31.1 \\ \cline{2-4} Maximum penetration & & Apiel & 21.5 & 24.5 \\ \cline{2-4}  & & Fluo-3 & Rhodamine B \\ \cline{2-4} Maximum penetration & & Apiel & 199.8 & 757.6 \\ \cline{2-4}  & & Medial & 694.1 & 780.4 \\ \hline \hline \end{tabular}
\end{table} 
Table 1: Interquartile range of each group. variable and different data in the Rhodamine B group compared to the Fluo-3 group, where all data were similar (Table 1). Thus, it could be observed that Fluo-3 presents a better performance and a lower variability, showing more homogeneous and more reliable data.