Abstract
ISH for SSTR2 mRNA was performed using the RNAscope 2.0 FFPE Assay (Advanced Cell Diagnostics, Inc., Hayward, CA, USA) on 4-μm formalin fixed and paraffin embedded tissue sections. After deparaffinization they were pretreated with heat and protease prior to hybridization with a target probe to the SSTR2. A horseradish peroxidase-based signal amplification system was then hybridized to the target probes followed by color development with 3,3 -diaminobenzidine. Control probes for the bacterial gene DapB (negative control) and for the POLR2A gene (positive control – evidence of adequate RNA) were also included in each case. The study slides (and corresponding controls) were read independently by two study pathologists (A.K. and J.L) and classified in a binary manner as either positive or negative. Positive cases had to have granular cytoplasmic and/or nuclear brown staining that was higher than the signal on the DapB negative control slide. Positive cases were subsequently semi-quantitatively assessed according to the manufacturer s scoring guideline as follows: Methods
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