The usefulness of 1-naphthol as substrate for horseradish peroxidase (HRP) in immunohistochemistry was studied using the peroxidase-antiperoxidase (PAP) and avidin-biotin-complex (ABC) methods in the demonstration of glial fibrillary acidic protein (GFAP), vimentin, carbonic anhydrase C (CA.C), and factor VIII-related antigen (FVIII/RAg) in central nervous tissue and cerebral tumors. In the presence of ammonium carbonate, 1-naphthol is oxidized by HRP and hydrogen peroxide, producing a fine gray-violet precipitate. The oxidation product of 1-naphthol proved capable of binding a great number of basic dyes. For each stain the final reaction product had a characteristic color that was different from the spontaneous color of the dye and from the color displayed by nuclei. The final color obtained with this procedure was alcohol resistant and could be mounted in solvent-based mounting media. The results obtained with the 1-naphthol basic dye (1-NBD) method were compared with those obtained using the diaminobenzidine (DAB) reaction in the demonstration of GFAP-positive astrocytes. The DAB reaction produced a more intense staining but also a coarser precipitate than the 1-NBD reaction. The 1-NBD procedure showed more morphological detail of fine structures and did not obscure nuclei and mitosis. The very low toxicity of 1-naphthol compared with DAB (a suspected carcinogen) is an important advantage of the 1-NBD method, as is its high specificity and sensitivity.
1-Naphthol basic dye (1-NBD). An alternative to diaminobenzidine (DAB) in immunoperoxidase techniques.
MAURO, Alessandro;GIORDANA, Maria Teresa;
1985-01-01
Abstract
The usefulness of 1-naphthol as substrate for horseradish peroxidase (HRP) in immunohistochemistry was studied using the peroxidase-antiperoxidase (PAP) and avidin-biotin-complex (ABC) methods in the demonstration of glial fibrillary acidic protein (GFAP), vimentin, carbonic anhydrase C (CA.C), and factor VIII-related antigen (FVIII/RAg) in central nervous tissue and cerebral tumors. In the presence of ammonium carbonate, 1-naphthol is oxidized by HRP and hydrogen peroxide, producing a fine gray-violet precipitate. The oxidation product of 1-naphthol proved capable of binding a great number of basic dyes. For each stain the final reaction product had a characteristic color that was different from the spontaneous color of the dye and from the color displayed by nuclei. The final color obtained with this procedure was alcohol resistant and could be mounted in solvent-based mounting media. The results obtained with the 1-naphthol basic dye (1-NBD) method were compared with those obtained using the diaminobenzidine (DAB) reaction in the demonstration of GFAP-positive astrocytes. The DAB reaction produced a more intense staining but also a coarser precipitate than the 1-NBD reaction. The 1-NBD procedure showed more morphological detail of fine structures and did not obscure nuclei and mitosis. The very low toxicity of 1-naphthol compared with DAB (a suspected carcinogen) is an important advantage of the 1-NBD method, as is its high specificity and sensitivity.
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