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Caldas Immunohistochemistry Protocol

Tissue microarrays were prepared using duplicate 0.6mm cores extracted from formalin-fixed paraffin-embedded blocks containing material from patient tumors and xenografts. These were run using Leica’s Polymer Refine Detection System (DS9800) on their automated Bond – III platform. The HIER’s (sodium citrate and tris EDTA pre-treatments) are all run at 100°C. The de-waxing and re-hydration prior to IHC are done on the automated Leica ST5020, as is the post-IHC de-hydration and clearing. Finally, the mounting is done on Leica’s CV5030. The slides were reviewed by a pathologist.

Antibody Manufacturer Catalogue No. Dilution/conc.
Mouse monoclonal (ER 6F11) anti-ER Novocastra NCL-ER-6F11/2 1.071 ug/m
Mouse monoclonal (PgR 636) anti-PR Dako M3569 1:50
Rabbit Monoclonal (4B5) anti-HER-2/neu Roche 790-2991 Unknown

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