Benign and malignant tissue specimens from a consecutive series of 448 patients undergoing radical prostatectomy for localized prostate cancer at Malmö University Hospital, Sweden, between 1988 and 2003 were mounted in 17 tissue microarray blocks using a manual tissue arrayer (Beecher Inc ,Wisconsin WI) as previously described[39].
Immunohistochemical stains were performed on 5 μm unstained sections from the tissue microarray blocks using the antibodies listed in Table 1.
From the selected regions, 1000 μm tissue cores were mounted into tissue microarray blocks using a custom made instrument (Beecher Instruments, Silver Spring, MD, USA).
Histologically representative tumor regions were selected by a neuropathologist (HH) and the samples from these areas were applicated in tissue microarray blocks using a custom-built instrument (Beecher Instruments, Silver Spring, MD, USA).
A dual-probe hybridisation was performed on 3- μm-thick sections from tissue microarray blocks using the LSI EGFR SpectrumOrange/CEP7 SpectrumGreen probe set (Vysis, IL, USA) as described elsewhere (Cappuzzo et al, 2005).
Immunohistochemistry was performed on 4 μm sections from the tissue microarray blocks using a Lab Vision Autostainer LV-1 (LabVision/Neomarkers, Fremont, CA, USA), according to the manufacturer's protocol.
Epidermal growth factor receptor protein expression was evaluated by immunohistochemistry (IHC) on 4- μm-thick sections from tissue microarray blocks using mouse anti-human EGFR, clone 31G7 monoclonal antibody (Zymed laboratories, CA, USA) was used with the labelled streptavidin-biotin complex staining method (LSAB kit, DAKO, CA, USA).
Representative core tissue sections of strong immunoexpression of EGFR were taken from paraffin blocks from each patient and arranged in a new tissue microarray (TMA) block using the Manual Tissue Arrayer (Beecher Instruments, Silver Spring, Maryland, USA).
Two tissue cores (2 mm in diameter) were obtained from each selected specimen and were precisely deposited into a recipient paraffin block using a tissue microarray workstation (tissue microarray builder ab1802; Abcam, Cambridge, UK) as described elsewhere [ 28, 29].
Three cylindrical tissue cores (2 mm in diameter) were obtained from each selected specimen and transferred to a recipient paraffin block, using a microarray instrument (TMA Builder, Histopathology Ltd., Hungary).
