Fig. 4

Heterogeneous PD-L1 expression patterns in the 3D space. (A) In 7 of 20 cases (35%), PD-L1 SP142 at different depth levels illustrated absence immunofluroescence change to signal expression (left). Case #18 showed no PD-L1 expression at a 20-µm depth (middle) and change into positive PD-L1 expression at a 100-µm depth (right). (B) In 6 of 20 cases (30%), PD-L1 SP142 at different depth levels illustrated PD-L1 category (cross 1%) change. Case #17 showed above 1% PD-L1 expression at a 15-µm depth (middle) and change into negative expression at a 140-µm depth (right). (C) Eight of 20 cases (40%) illustrated immune cell scores for all levels of 3D fluorescence images were difference from original IHC. Case #5 showed the different PD-L1 expression level in IHC and 3D fluorescent image. Scalebar, 1000-µm (whole specimen image), scale bar = 100-µm (region of interest image)