Semi automated tissue arrayer

65 non-consecutive, unselected primary ovarian cancer specimens were included in the tissue microarray. The tumor samples were collected within one hour after resection from the primary site. Formalin-fixed paraffin-embedded (FFPE) tissue blocks were prepared according to the standard procedure. Tissue cylinders of 2 mm in diameter were punched from representative areas of each block with regard to the matching H&E staining control by a MiniCore Control Station (Alphelys Sarl, France). The Selected tissue cylinders were re-arranged and brought into three paraffin blocks by a semi-automated tissue arrayer (Beecher Instruments, Sun Prairie, WI, USA). 4 µm section slides were prepared for further use.

ALDH1A1 and ALDH1A3 protein expression was detected and quantified using immunohistochemistry (IHC). Fresh frozen FFPE tissue blocks (donor blocks) were used to create tissue microarrays (TMA). Three representative cores per sample from donor blocks were placed into a TMA recipient using a semiautomated tissue arrayer (Beecher Instruments, Sun Prairie, WI, USA). Immunohistochemistry was performed after deparaffinization, following treatment with a primary anti-ALDH1A1-antibody (Thermo Fisher Scientific, PA5-11537) or anti-ALDH1A3-antibody (Atlas Antibodies, HPA046271) on the Ventana BenchMark (Roche, Basel, Switzerland) by using the IView DAB Detection Kit. Expression levels were evaluated by two pathologists (AO, SP) and categorized according to negative, low to moderate, and high staining intensity. The protein expression in all three replicates per sample was considered, and the highest expression in a single core was used for further analysis in cases of heterogeneous staining levels. Androgen receptor (AR) expression was detected and evaluated as described before 93 (link).