Dual Immunohistochemistry-Aniline Blue Stain: The Trichrome Stain Revisited

摘要

The degree of fibrosis in a liver biopsy provides important information about the stage and prognosis of liver disease and helps to guide therapy. The trichrome stain, by highlighting fibrous tissue (collagen), is therefore essential for the interpretation of liver biopsies. The goal of this project was to combine a collagen stain with an immunohistochemical stain. Ki-67 (a proliferation marker) was used as an example of a nuclear immunohistochemical stain. Once optimized, the procedure has the potential to be extended to cytoplasmic markers. A similar procedure has been previously published, but all three dyes of the trichrome were used. Because we were interested in staining only collagen and combining it with a nuclear immunomarker, we sought to eliminate the cytoplasmic and nuclear dyes of the trichrome stain. In simple terms, we were interested in combining immunohistochemistry with an aniline blue stain. We carried out experiments to determine whether other steps of the trichrome stain such as postfixation with Bouin's fluid, treatment with phosphotungstic/phosphomolybdic acids, and acetic acid, are essential for the staining of collagen with aniline blue. We optimized the dual staining procedure by eliminating or shortening various steps which resulted in a more efficient procedure.