Fig 2: Comparison of large BCL6+ secondary follicular TLS in human melanoma to tonsil tissue. (A) Melanoma: CD20+ lymphocyte aggregates with a germinal center surrounded by CXCL13 expressing cells. A CD21+ and CD23+ FDC network covers the complete germinal center area with randomly interspersed CD4+ T cells and BCL6+ cells. No detectable germinal center polarity, no identifiable light and dark zone areas in routine light microscopy (see Pathology View, bottom left). (B) Close up of BCL6+CD20+ B cells and BCL6+CD4+ T cells from a BCL6+ secondary follicular TLS in human melanoma. (C) Tonsil: note canonical germinal center polarity with polarized spatial distribution of the CD21+ and CD23+ FDC network together with CD4+ T cells (arrowheads) and a complementary localization of BCL6+ cells (arrow). Identifiable light (arrowhead) and dark zone (arrow) areas in routine light microscopy (see Pathology View, bottom left). (D) Tonsil (upper row): polarized distribution of BCL6+, Ki67+ (arrows) and BCL6+Ki67+ cells in germinal centers with canonical germinal center polarity and the presence of light (arrowhead) and dark zone (arrow) areas in routine light microscopy (see Pathology View, left). Melanoma (lower row): note decreased numbers and absence of polarized distribution of BCL6+, Ki67+ and BCL6+Ki67+ cells in melanoma germinal centers. Lack of canonical germinal center polarity and absence of light and dark zone areas in routine light microscopy (see Pathology View, left). Images for each of the individual markers and their composites (for clarity without DAPI staining) are shown, together with the corresponding Pathology View (respective (bottom) left). Scale bars represent 100 µm (A, C, D) and 5 µm (B).

Fig 3: Correlation analysis between TIL-B and mature tertiary lymphoid structures (TLS). (A) Among the 200 patients included in the BRCA dataset from the TCGA research consortium, a subset of 18 patients who exhibited TLS positivity were identified. H&E images of characteristic TLS in these patients. (B) Co-immunofluorescence subimages depicting the co-localization of CD23 and CD21 within CD23-positive TLS were obtained from a collection of 70 BC samples that we compiled. (C) In 9 TLS-positive patients of 70 BC patients, representative images of H&E staining and IHC staining of CD4, CD8, CD20, CD23 and BCL6 expression in TLS-positive BC patients. (D) Analysis of DEGs in 200 TCGA BRCA cases categorized as TLS-negative and TLS-positive. Source data are available at: https://portal.gdc.cancer.gov. (E and F) H&E (E) and CD23 IHC (F) images of the typical TLS-positive case in 70 BC patients. (G) Different tumour types exhibit FCER2 expression, according to TIMER2. *p < .05;**p < .01;***p < .001. (H) Utilizing the EPIC and TIDE algorithms within TIMER2, the relationship between FCER2 expression and tumour infiltrating immune cells in BC was computed. (I-J) Kaplan–Meier Survival curve of FCER2(I), co-acting with FCER2 and B cells(J) in BC by TIMER2. TIL-B, tumor-infiltrating B cells.