## 4 Discussion
### Study Methodology  
Standardized and updated guidelines must be followed when performing in vitro studies [59]. Variations in the sample preparation methods could alter the results obtained. For example, bovine and swine substrates allow higher marginal leakage than human substrates [60]. Therefore, to maintain consistency in the findings, animal teeth studies were excluded from this study to reduce methodological heterogeneity. The use of human substrates was considered more relevant to the study's objectives, and this decision contributes to the consistency of the results. Furthermore, primary teeth differ from permanent ones because they have thinner enamel, dentin, and longer pulp horns [40]. Therefore, the qualitative results were synthesized into two parts, primary and permanent teeth, to analyze the difference in the results.  
Determining the appropriate sample size is crucial for achieving scientifically and statistically valid outcomes [61]. It is worth noting that among the studies reviewed, there was a lack of reporting or incomplete reporting of sample size calculations, and only two reported calculating the necessary sample size for the teeth used [40; 41]. Additionally, two studies with small sample sizes produced different results from other included studies and demonstrated no statistical differences in their outcomes [43; 51]. However, inadequate sample sizes can compromise the ability to detect differences between groups, leading to potentially falsely adverse outcomes and an elevated risk of a type II error [62].  
The results of microleakage studies can be affected by cavity design. Notably, the U-shaped Class V cavity demonstrated superior performance in minimizing microleakage compared to the V-shaped design [63], a finding not mentioned in the studies.  
Penetration may also be affected by the chosen dye material, the dye material's concentration and diffusion coefficient, the dentin's thickness, and the dentin's available surface area for diffusion [63]. For example, rhodamine B dye has a molecular size smaller than the diameter of a denital tubule. It presents greater diffusion on human dentin than methylene blue, resulting in a higher leakage score [47]. The tested teeth were immersed in dye material for 24 h in all selected studies except for one study, in which the sample was immersed in 2% methylene blue dye for 4 h [43].  
Thermocycling is a widely used method in laboratory dental studies to simulate temperature changes in the oral environment. The International Organization for Standardization (ISO) recommends thermal cycling between 5 and 55 \({}^{\circ}\)C as an accelerated aging test [64]. Accordingly, most authors chose cycling temperatures between 5 and 55 \({}^{\circ}\)C except in one study by [PERSON] et al. [43], where no thermocycling was applied. Concerning the methods used to detect microleakage, the authors opted for semi-quantitative assessment by different scores and quantitative assessment as a percentage in millimeters, allowing for precise measurements of the extent of leakage. The microleakage assessment using stereomicroscope methods featured high intra- and inter-examiner reproducibility levels [64]. However, a few studies referenced the level of inter-examiner agreement [39; 40; 42; 45; 48], indicating almost perfect inter-examiner reliability.