Immunofluorescence Analysis of FFPE Tumor Samples

Formalin-fixed, paraffin-embedded (FFPE) sections from primary MCC tumours were purchased from Origene and analysed as previously described [32 (link)]. Primary antibodies were: FITC-conjugated anti-CK20 (Dako, dilution 1:50), MCPyV LT CM2B4 (Santa Cruz Biotechnology, dilution 1:125) and ADAM 10 and 17 (Abcam, dilution 1:250). An isotype-matched irrelevant antibody was used as a negative control on sections of tissues in parallel, a rabbit polyclonal isotype control antibody (Abcam) was used to match the ADAM 10 primary antibody. Sections were incubated with appropriate secondary antibodies labelled with different fluorochromes (Alexa Fluor 546 IgG2B, 643 IgG2A, Invitrogen, and IgG (H+L)-TRITC, Jackson ImmunoResearch). All slides were mounted with Immuno-Mount and images were captured with a Zeiss LSM880 confocal laser scanning microscope.