Ck19 rck108

The tissue specimen was fixed in formalin and processed routinely for histopathology. Immunohistochemistry (IHC) was performed on 3-µm sections cut from paraffin blocks using a fully automated system (“Benchmark XT System”, Ventana Medical Systems Inc, 1910 Innovation Park Drive, Tucson, Arizona, USA) and the following antibodies: CK7 (OV-TL, 1:1000, Biogenex), p63 (SSI6, 1: 100, DCS), CK5 (clone XM26, 1: 50, Zytomed), EMA (E29, 1:20, Dako), CEA (polyclonal, 1:400, Dako), CK19 (RCK108, 1:300, Dako), p16 (clone JC8, 1:100, Santa Cruz Biotechnology), TP53 (DO-7, 1:50, Dako) and anti-NUT antibody (clone C52B1, 1:45, Cell Signaling).

All 13 patients included in this study were analyzed by IHC. The immunohistochemical analysis was performed on paraffin-embedded sections on a DAKO Autostainer. The primary antibodies used in the study included synaptophysin (SP11, dilution 1 : 100; Thermo Fisher Scientific), chromogranin A (DAK-A3, dilution 1 : 100; DAKO), Ki-67 (MIB-1, dilution 1 : 200; DAKO), cluster of differentiation protein 56 (123C3, dilution 1 : 100; DAKO), CK7 (OV-TL12/30, dilution 1 : 400; DAKO), CK19 (RCK 108, dilution 1 : 100, DAKO), and cytokeratin (AE1/AE3, dilution 1 : 100; DAKO). Appropriate positive and negative controls were used for all antibodies tested. For each immunohistochemical procedure, antigen retrieval was performed in a citrate buffer, and detection was amplified with the DAKO EnVision System. Mitoses were counted in at least 50 HPFs (1 HPF = 2 mm²), and the Ki-67 index was defined using the MIB antibody as the percentage of 500-2000 cells counted in areas of the strongest nuclear labeling (“hot spots”) [6 ].