Ata 100

For each paraffin embedded tumor, optimal sites of the formalin-fixed paraffin embedded blocks for core sampling were identified. Characteristics of optimal sites included the following features: high cellularity, minimal necrosis, minimal hemorrhage and minimal matrix or bone deposition. The construction of the tissue microarray (TMA) was based upon a 2009 review by Parsons and Grabsh [25 (link)]. An Advanced Tissue Arrayer (model ATA-100, Chemicon International) was used to cut and insert canine OSA tissue core samples, in triplicate, into pre-cast paraffin blocks (Paraplast Plus, Sigma Aldrich). For the 33 canine cases, four canine OSA TMAs were constructed using a 9 × 6 grid pattern. Three, 2 mm diameter core biopsies from each donor tissue block, along with 2 positive control core biopsies (canine glioblastoma tissue) were semi-randomly arranged in the paraffin block while the outer circumferential border was generally comprised of negative control tissue (normal canine cerebrum or renal cortical tissue). To generate unstained paraffin sections, each assembled TMA was cut into 4 μm thick sections with a microtome and placed on positively charged glass slides. TMA sections were stained with either hematoxylin and eosin stains according to standard protocols or were further processed for P16 immunohistochemistry.

The advanced tissue arrayer (ATA-100, Chemicon International, Tamecula, CA, USA) was used to create holes in a recipient paraffin block and to acquire cylindrical core tissue biopsies with a diameter of 1 mm from the specific areas of the ‘donor’ block. The tissue core biopsies were transferred to the recipient paraffin block at defined array positions. The tissue microarrays contained tissue samples from 75 formalin-fixed paraffin-embedded cancer specimens with known diagnosis, and corresponding ANCT from these patients. The block was incubated in an oven at 45°C for 20 min. to allow complete embedding of the grafted tissue cylinders in the paraffin of the recipient block, and then stored at 4°C until microtome sectioning.

Nine tissue microarrays (TMAs) were constructed using a manual tissue arrayer (Chemicon ATA-100). Tissue cores of 1 mm in diameter were obtained from the most representative tumor areas of FFPE tissue blocks and were arranged in recipient TMA blocks. The TMAs contained cores from 220 radical prostatectomy specimens. For each case, 1–2 cores of the different Gleason score tumor areas were selected from haematoxylin-eosin (H&E) stained sections of donor blocks. From the nine resulting TMAs, sections of 3 μm were transferred to glass slides. H&E staining of the TMA sections were performed for diagnostic and grading confirmation and assessed by two expert pathologists.