*4.3. Semi-Quantitative Assessment*

BMI-1 expression has been individually quantified in the epithelial and in the stromal components. The semi-quantitative assessment of the BMI-1 in tumor cells was done by using adapted scores based on literature reports [27,51] that took into account the staining intensity (I) and the percentage of positive cells (P). BMI-1 showed a double immunostaining, nuclear, and cytoplasmic/membrane [27,51]. The intensity of BMI-1 immunoreaction was scored as: 0—absent, 1—weak, 2—moderate, and 3—strong. The percentage of BMI-1 positive cells was scored as follows: 1—< 10%, 2—10–50%, 3—> 50%. The final BMI-1 score was obtained by multiplying P by I. BMI-1 score values < 3 were considered as a low score, and score values ≥ 3 were considered as a high score.

For the semi-quantitative assessment of stromal BMI-1, we used a standard 2-point scale scoring system. The immunoreaction was considered negative when ≤ 10% of the tumor stromal area had a positive immunostaining of BMI-1, and positive when > 10% of the stromal area showed BMI-1 immunostaining, regardless of the level of staining intensity.

BMI-1 expression has been independently evaluated and scored by three histopathologists with experience in immunohistochemistry interpretation and scoring differences have been revised in the evaluation panel in order to reach a consensus.
