Cd10 clone 56c6

Immunohistochemistry (IHC) was performed with the Dako Autostainer Link 48, according to the manufacturer’s recommendations, for the antibodies CD10 (clone 56C6 from Dako, diluted 1:20), BCL6 (clone PG-B6p from Invitrogen, diluted 1:100), MUM1 (clone MUM1p from Dako, diluted 1:100), BCL2 (clone 124 Dako, diluted 1:80), and IgM (polyclonal, from Dako, diluted 1:500). The markers were scored positive in case of expression in ≥ 30% of the tumor cells for CD10, BCL6, and MUM1, and in ≥ 50% of the tumor cells for BCL2 and IgM.

IHC (Figure 6) was performed on 4 μm sections with formalin-fixed paraffin-embedded (FFPE) specimens. Antibodies applied in the study including CD10 (clone 56C6; Dako, cut-off: 30%), MYC (clone Y69; Abcam, cut-off: 40%), BCL2 (clone 124; Dako, cut-off: 50%), BCL6 (clone LN22; Dako, cut-off: 30%), and MUM1 (clone MUM1p; Dako, cut-off: 30%). The COO was classified according to Hans algorithm [32 (link)].

Antibodies applied in the study, according to the manufacturer’s instructions, included CD5 (clone EP2952, Abcam, cut-off: 30%), CD10 (clone 56C6, Dako, cut-off: 30%), CD30 (clone CON6D/B5, Abcam, cut-off: 30%), Ki-67 (clone Mib-1, Dako), Myc (clone Y69, Abcam, cut-off: 40%), Bcl2 (clone 124, Dako, cut-off: 50%), Bcl6 (clone LN22, Dako, cut-off: 30%), MUM1 (clone MUM1p, Dako, cut-off: 30%), FOXP1 (clone JC12, Abcam, cut-off: 60%), GCET1 (clone RAM341; Abcam, cut-off: 60%) and LMO2 (clone 1A9-1, Santa Cruz, cut-off: 30%). The cell of origin (COO) was classified according to Hans, Choi, Tally and Visco-Young algorithms. The specific cut-off of each antibody used in different algorithms was described previously21 (link)22 (link)23 (link)24 (link).

IHC was performed on 4-μm FFPE sections. The antibodies used were CD10 (clone 56C6, Dako), CD20 (clone L26, Abcam), Bcl6 (clone LN22, Dako), and MUM1 (clone MUM1p, Dako), Myc (clone Y69; Abcam, cut-off: 40%) and Bcl2 (clone 124; Dako, cut-off: 50%) (Figure S1). The cut-off scores for each antibody were described previously9 (link)19 (link). Cases positive for both CD10 and MUM1were classified as DP group. Cases negative for CD10, Bcl6 and MUM1 were defined as TN group. Cases positive for both Myc and Bcl2 or Bcl6 were defined as double expression lymphoma (DEL)19 (link).

Four renal tumours from Fischer male rats given protracted dietary OTA (300 µg/kg body weight daily) [6 (link)] were embedded in paraffin blocks. Animals were from the same lifetime experimental group [6 (link)] as those whose immunoprofiles were previously described [9 (link)]. Ethical review and approval were waived for this study because no new live animals were involved. Sections (3 μm) were mounted on charged slides (TOMO, Matsunami, Japan) and processed for immunohistochemistry in the Cell Pathology Laboratory of South West London Pathology at St George’s Hospital, Tooting, variously applying panels of antibodies in fully automated BenchMark ULTRA immunohistochemistry processing, as required to assist clinical diagnoses. Procedures followed exactly those previously described [9 (link)]. The following antibodies were used: CK MNF 116, clone MNF 116 (Dako); Vimentin, clone V9 (Dako, Novocastra); CD10, clone 56C6 (Dako). After applying DAB chromogen, nuclei were counterstained blue with haematoxylin. The brown immune reaction product of DAB chromogen is cytoplasmic and/or membranar for the listed antibodies. Haematoxylin and Eosin staining was also performed for preliminary standard tissue differentiation of nuclear (blue) and cytoplasmic components (red).

Immunohistochemistry was performed on 4-μm formalin-fixed paraffin-embedded sections. The antibodies used in the study were for CD20 (clone L26, Abcam, cutoff: 30%), CD10 (clone 56C6, Dako, cutoff: 30%), Bcl6 (clone LN22, Dako, cutoff: 30%), MUM1 (clone MUM1p, Dako, cutoff: 30%), and Myc (clone Y69; Abcam, cutoff: 40%). The cutoff values for each antibody have been previously described [7 (link)].