Coloring the Canvas: Essential Stains for Pathology (Part 2)

PAS : Periodic Acid-Schiff (PAS) stain

This is used to stain polysaccharides such as glycogen, mucosubstances like glycoproteins, glycolipids, and mucins in tissues.
Here are a few examples of conditions where this stain is utilized:

• Candida: The budding cells and pseudohyphae stains red / purple. 
• Acute Lymphoblastic Leukemia (ALL): PAS-positive blasts.
• Multiple Myeloma: PAS-positive cytoplasmic inclusions.
• Hodgkin’s Lymphoma: PAS-positive Reed-Sternberg cells.
• Mucinous Adenocarcinomas: PAS-positive mucin.
• Ewing’s Sarcoma: PAS-positive glycogen.

ALICIAN BLUE :

This stain is used to stain acidic polysaccharides such as glycosaminoglycans in cartilages and other body structures, as well as some types of mucopolysaccharides.At pH 2.5 = this is the most widely used pH, it stains both carboxylated and sulphated acidic polysaccharides.
At pH 1.0 = the stain becomes more selective for strongly sulphated mucins.
Here are a few examples of conditions where this stain is utilized:

• Adenocarcinomas: To highlight mucin production.
• Mucoepidermoid Carcinoma: To identify mucin-producing cells.
• Adenoid Cystic Carcinoma: To stain the mucinous component.
• Colorectal Carcinoma: To detect mucinous differentiation.
• Salivary Gland Tumors: To identify mucin in various neoplasms.

AB – PAS = Alcian Blue with Periodic Acid Schiff stain

The Alcian blue at a pH of 2.5 will stain all acid mucins deep blue but will not color the neutral mucins. The subsequent application of the PAS technique will stain the neutral mucins bright magenta.

Tissues or cells that contain both neutral and acidic mucins may demonstrate a dark blue or purple coloration.

The combined Alcian blue/PAS technique is perhaps the most sensitive and comprehensive method for detecting mucins, as all mucins should react regardless of the charge nature of the mucin.

MTS : Masson’s Trichrome stain

This is used to differentiate between various tissue components:

• Collagen fibers: Stains blue or green (depending on the variant used).
• Muscle fibers: Stains red.
• Cytoplasm: Stains red.
• Fibrin: Stains bright red.
• Nuclei: Stains black or dark blue.

This staining technique is particularly useful for distinguishing connective tissues from muscle tissues. These applications help pathologists assess the extent of fibrosis, stromal reaction, and the overall architecture of the tumor.
Masson’s Trichrome stain has a few variants, each with different colours for collagen fibres:

• Standard Masson’s Trichrome: Collagen fibers stain blue.
• Lillie’s Modification: Collagen fibers stain green.
• Gomori’s One-Step Trichrome: Collagen fibers stain green.

Here are a few examples of conditions where this stain is utilized:

• Fibrosarcoma: To identify collagen fibers within the tumor.
• Desmoplastic Small Round Cell Tumor: To highlight the desmoplastic stroma.
• Schwannoma: To visualize the collagenous stroma.
• Gastrointestinal Stromal Tumor (GIST): To assess the collagen matrix.
• Liver Cirrhosis with Hepatocellular Carcinoma: To differentiate between fibrotic tissue and tumor cells.

Verhoff’s Stain:

Verhoeff’s stain is used to stain elastic fibers. It is particularly useful in identifying elastic fibers in various tissues and can be employed in the detection of vascular invasion in neoplastic conditions such as colorectal adenocarcinoma. These applications help pathologists accurately identify and assess the extent of vascular invasion, which is crucial for staging and prognosis.
Here are a few examples of conditions where this stain is utilized:

• Colorectal Adenocarcinoma: To detect venous invasion.
• Breast Cancer: To identify vascular invasion.
• Pancreatic Cancer: To assess vascular invasion.
• Thyroid Carcinomas: To detect vascular invasion.

Van Gieson’s Stain:

This stain is used to differentiate between collagen and other connective tissue components. Specifically, it stains:

• Collagen fibers: Red
• Muscle fibers: Yellow
• Cytoplasm: Yellow
• Nuclei: Black (when combined with hematoxylin)

This staining technique is particularly useful for highlighting collagen in various tissues and is often used in conjunction with Verhoeff’s stain to assess elastic fibers and collagen simultaneously.
Here are a few examples of conditions where this stain is utilized:

• Fibrosarcoma: To identify collagen fibers within the tumor.
• Desmoplastic Small Round Cell Tumor: To highlight the desmoplastic stroma.
• Schwannoma: To visualize the collagenous stroma.
• Gastrointestinal Stromal Tumor (GIST): To assess the collagen matrix.

Verhoeff-Van Gieson (VVG) stain

This is a histological staining technique that combines Verhoeff’s elastic stain with the Van Gieson counterstain. It is particularly effective for differentiating and studying elastic fibers in tissue sections. Here’s what it stains:

• Elastic fibers: Black (Verhoeff’s stain)
• Collagen fibers: Red (Van Gieson’s stain)
• Muscle fibers and cytoplasm: Yellow (Van Gieson’s stain)
• Nuclei: Blue-black (Verhoeff’s stain)

The VVG stain is primarily used for identifying and assessing elastic fibers in tissues such as skin, lungs, and blood vessels. It is also useful in diagnosing vascular diseases like arteriosclerosis and examining connective tissue disorders.

Myeloperoxidase (MPO)

This stain is used to identify myeloid cells by staining the enzyme myeloperoxidase, which is present in the primary granules of neutrophils, eosinophils, and monocytes. It is particularly useful for differentiating between acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL), as MPO is positive in myeloid blasts (AML) and negative in lymphoid blasts (ALL)
Here are a few examples of conditions where this stain is utilized:

• Acute Myeloid Leukemia (AML): MPO-positive blasts help differentiate AML from other types of leukemia.
• Chronic Myeloid Leukemia (CML): MPO can be used to identify myeloid cells in CML.
• Myelodysplastic Syndromes (MDS): MPO staining helps in the assessment of myeloid cell differentiation.

Giemsa stain

This is used to stain blood cells, bone marrow cells, and microorganisms. It is particularly useful for identifying parasites, bacteria, and other microorganisms in blood smears.

Here are a few examples of conditions where this stain is utilized:

• Leukemias: To identify abnormal white blood cells.
• Lymphomas: To examine lymphoid cells.
• Multiple Myeloma: To identify plasma cells.
• Mast Cell Tumors: To stain mast cells.
• Ewing’s Sarcoma: To identify small round cells.

Congo Red Stain

This is used to identify amyloid deposits in tissues. It is particularly useful for diagnosing amyloidosis, where it shows apple-green birefringence under polarized light.

Perls’ Prussian Blue stain

This is used to detect iron deposits in tissues. It stains iron in the ferric state, such as ferritin and hemosiderin, blue. This stain is particularly useful for diagnosing conditions related to iron metabolism and storage.

Here are a few examples of conditions where this stain is utilized:

• Hepatocellular Carcinoma: To identify iron deposits in liver tissue.
• Myelodysplastic Syndromes (MDS): To detect ring sideroblasts, which are erythroblasts with iron-loaded mitochondria.
• Hemochromatosis: To assess iron overload in various organs.
• Refractory Anemia with Ring Sideroblasts (RARS): To identify ring sideroblasts in bone marrow.

Grocott-Gomori’s Methenamine Silver (GMS) stain

This is used to detect fungal organisms and carbohydrates in tissue sections. It is particularly useful for identifying fungal infections, such as Pneumocystis jiroveci and Histoplasma spp.

In conclusion, the critical role of stains in pathology cannot be overstated. These essential stains transform the invisible into the visible, revealing the subtle intricacies of cellular structures and tissue components. By enhancing contrast, stains enable pathologists to diagnose diseases with greater accuracy and provide invaluable insights into the underlying mechanisms of various conditions. 

From basic stains like Hematoxylin and Eosin (H&E) to specialized stains such as Periodic Acid-Schiff (PAS) and Immunohistochemistry (IHC), each serves a unique purpose, contributing to the rich tapestry of diagnostic pathology.

As we advance in the field, the development of new staining techniques and the refinement of existing ones will undoubtedly continue to enhance our understanding of disease processes. By embracing the power of these colorful tools, pathologists can continue to paint a clearer picture of health and disease, ultimately improving patient outcomes.

Remember, each stain is a brushstroke that adds depth and detail to the diagnostic canvas, making pathology a truly vibrant and dynamic field.