Immunohistochemical Staining for L1CAM, Notch, Jagged, p53, p21, and p38

Immunohistochemical staining for L1CAM using sections of paraffin-embedded tissues was carried out as previously described [13 (link)] [19 (link)]. To increase staining intensity, L1CAM staining for the inintially L1CAM low melanoma cells MV3 was modified in that the K5005 AP/RED Rabbit/Mouse staining kit (Dako, Copenhagen, Denmark) was used after the primary antibody. Other stainings were carried out as follows: Notch 1 (Abcam, Cambridge, UK; cat. no. ab52627; Clone EP1238Y) at 19.2 μg / ml; pretreatment steamer at 121°C for 10 min; citrate buffer pH 6. Jagged 1 (Jag1) (Sigma-Aldrich, St. Louis, MO, USA; cat. No. HPA021555) at 6 μg / ml; water bath at 99°C for 20 min; S1699 retrieval solution pH 6 (Dako). p53 (Dako; cat.no. M7001; clone DO-7) at 1.37 μg / ml, microwave 2X 4 min at 800 W, citrate buffer pH 6. p21 Thermo-Fisher, Waltham, MA, USA; cat.no. MS-891-P; clone CP74400) at 1 μg / ml; steamer at 121°C for 10 min; S1699 pH6. p38 (Abcam; cat.no. ab38238; clone pohosoha T180 + Y1820) at 2 μg / ml, steamer at 100°C for 20 min, S1699 pH6 with K5005 AP/RED treatment after primary antibody.
Stained slides were scanned by a Mirax microscope (Zeiss, Jena, Germany) and the Panoramic Viewer software (3D Histech, Budapest, Hungary) was used to take images.