Table 5.
Stains and alternative methods that can be used with tissue sections
| Stain(s) | Application(s) | Color and fungi stained |
|---|---|---|
| Hematoxylin and eosin (H&E) | Used routinely in pathology to demonstrate tissue morphology; in the case of fungal infections, this stain helps identify the inflammatory host reaction, such as multinucleated giant cells, necrotic material, hemorrhage, and the Splendore-Höeppli phenomenon; most fungi can be observed with this stain, particularly the nuclei of yeast-like cells or if the fungus is naturally pigmented (however, the fungal elements may be difficult to distinguish from the background) | All fungi show pink cytoplasm, blue nuclei, no color for the wall |
| Fungal silver stains (Grocott and Gomori methenamine silver [GMS]) | They highlight the wall of the fungus and thus are useful for screening the tissue sample, can be combined with H&E in such a way that the fungus and the host reaction can be clearly observed | The fungal cell wall appears black or dark brown for all fungi, the surrounding tissue is usually green, the Mucorales may stain very pale |
| Periodic acid-Schiff (PAS) | Detects glycogen in tissues, fungal walls contain large amounts of glycogen and thus PAS can be used for screening for fungal organisms | The fungal cell wall appears pink to red purple; depending on the counterstain used, the nuclei can be blue |
| Gridley fungus | Stains the walls of most fungi | The fungal cell wall appears purplish red, the background is usually yellow |
| Mucine stains (Mayer's or Southgate mucicarmine, Alcian blue) | Stains mucopolysaccharides, including the capsules of a variety of organisms; also stains mucus, which can be present in a variety of human cells | Useful to highlight the capsules of cryptococci, which appear red or blue depending on the stain used |
| Melanin stains (Fontana-Masson) | Stains melanin present in some fungi; also stains melanin present in human tissues, such as in the skin epidermis | Cryptococcus and the dematiaceous fungi will take a black to dark brown color |
| Bacterial stains (tissue Gram stains or acid-fast stains) | Many fungi take bacterial stains; additionally, some filamentous bacteria (actinomycetes and Nocardia) have to be differentiated from fungi | Candida spp. stain purple/blue (Gram positive) with the Gram stain; Blastomyces and Histoplasma can be acid-fast staining (take a red color) |
| Immunohistochemistry (IHC) | Uses antibodies against the different fungi, antibodies can be monoclonal or polyclonal, assays are not FDA approved and require validation by each laboratory | Assays for Blastomyces, Cryptococcus, Histoplasma, Coccidioides, Pneumocystis, Sporothrix, Paracoccidioides, Penicillium, Candida, Aspergillus, and Mucorales have been published in the literature; depending on the colorimetric developer, the fungi can stain dark brown or red; the counterstain is usually hematoxylin; cross-reactivity of the antibodies is a concern |
| In situ hybridization (ISH) | Uses molecular probes for different fungi, probes are generally ribosomal since there are multiple copies of ribosomal genes in each fungal cell, assays are not FDA approved and require validation by each laboratory | Probes published in the literature include those for Blastomyces, Cryptococcus, Histoplasma, Coccidioides, Pneumocystis, Sporothrix, Candida, Aspergillus, Mucorales, Fusarium, and Pseudallescheria; depending on the colorimetric developer, the fungi can stain dark brown or red; sensitivity and specificity of the probes vary |