A3534

IHC was performed with antibodies against BCL-2 (M0887, Dako, Glostrup, Denmark), BCL-XL (SC7195, Santa Cruz Biotechnology, Santa Cruz, CA, USA) and MCL-1(A3534, Dako, Glostrup, Denmark) on paraffin-embedded tissue sections after antigen retrieval (pH 6, pH 9, and pH 6, respectively). Staining was visualized using HRP-labeled secondary antibodies (Dako) and 3,3′-diaminobenzidine (Sigma–Aldrich, St Louis, MO, USA). Appropriate positive and negative controls were performed for each staining. The cases were stained and scored on a tissue micro-array (TMA). In the TMA, each case was represented by three tissue cores.

Immunohistochemistry was performed with antibodies against BCL-2 (M0887, Dako, Glostrup, Denmark), BCL-XL (SC7195, Santa Cruz Biotechnology, Santa Cruz, CA, USA), BCL-W (4G12E6, Novus biologicals, Bio-Techne, UK), MCL-1 (A3534, Dako) and ERβ (CWK-F12, 1:160, Developmental Studies Hybridoma Bank, Iowa, USA) on paraffin-embedded tissue sections after antigen retrieval (pH 6,pH 9, pH 9, pH 6, and pH 9, respectively). Staining was visualized using HRP-labeled secondary antibodies (Dako) and 3,3′-diaminobenzidine (Sigma Aldrich, St Louis, MO, USA). Appropriate positive and negative controls were performed for each staining. The cases were stained and scored on a tissue micro-array (TMA). The TMA contained a total of 84 HL cases with 67 cases of NS HL, 7 cases of MC, 1 NOS HL, and 9 NLPHL. Patient material was acquired in accordance with international regulations and professional guidelines (the Declaration of Helsinki and the International Conference on Harmonization Guidelines for Good Clinical Practice). This project has been approved by the internal review board of the UMCG under RR#201800551.