The basics: Diagnostic terms, skin anatomy, and stains

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Chapter 1

The basics

Diagnostic terms, skin anatomy, and stains

Tammie Ferringer and Christine J. Ko

Glossary of terms

Normal skin anatomy

Scalp skin

Key features

• Numerous follicles that extend down into the panniculus

• Associated sebaceous glands, arrector pili muscles

Facial skin

Skin of the trunk

Key features

• Very thick dermis, especially in skin from the back

• Scattered hair follicles and sebaceous glands

• Projections of fat extend upward to envelop adnexae

Fig 1-36 Villi, warty dyskeratoma

Fig 1-37 Normal scalp with follicles rooted in the fat

Fig 1-38 Sun-damaged facial skin

Fig 1-39 Sun-damaged skin. Solar elastosis spares papillary dermis

Fig 1-40 Eyelid

Areolar skin

Key features

• Slight acanthosis of the epidermis with basilar hyperpigmentation

• Sometimes there is a central invagination of the epidermis that leads to a follicle and sebaceous glands

• Smooth muscle bundles in the mid–deep dermis

• Apocrine glands in the reticular dermis

Fig 1-41 Eyelid

Fig 1-42 Ear

Fig 1-43 Areolar skin, smooth muscle bundles

Acral skin

Key features

• Compact eosinophilic stratum corneum

• Slight papillomatosis present on dorsal surfaces

Volar skin

Key features

• Compact eosinophilic hyperkeratosis with underlying stratum lucidum

• No hair follicles or sebaceous glands

• Eccrine glands numerous

• Meissner and Pacinian corpuscles may be seen

Fig 1-44 Volar skin, thick stratum corneum and deep Pacinian corpuscles

Fig 1-45 Volar skin, stratum lucidum

Fig 1-46 Volar skin, eccrine glands and Pacinian corpuscle

Mucosa

Key features

• Absent granular layer

• Keratinocytes are large and pale (filled with glycogen)

• Dilated vessels in the submucosa

• Smooth muscle bundles may be present

Fig 1-47 Mucosa

Fig 1-48 Smooth muscle in submucosa

Fig 1-49 Nasal turbinate mucosa: erectile tissue with mucous glands

Fig 1-50 Fetal mesenchyme

Fig 1-51 Fetal periderm

Nasal turbinate

Key features

• Erectile tissue with fibrous septa and vascular sinusoids

• Mucous glands

Fetal skin

Key features

• Stellate and spindled fibroblasts (mesenchyme)

• Densely cellular

Hair anatomy

Infundibulum

Key features

• From epidermis down to insertion of sebaceous gland

• Intraepidermal portion = acrotrichium

• Keratinizes in the pattern of the normal epidermis with a granular layer (keratohyaline granules)

Isthmus

Key features

• From the insertion of the sebaceous gland to the insertion of the arrector pili muscle (bulge)

• Keratin is formed in the absence of a granular layer = trichilemmal keratinization

• The inner root sheath is lost at this level and the outer root sheath develops an inner corrugated, dense pink cornified layer; peripheral palisading of the outer root sheath is seen

Stem

Key features

• From the insertion of the arrector pili muscle (bulge) to Adamson’s fringe

• Only present in anagen hairs

Adamson’s fringe

• The point above which hair cornifies

• Dermatophytes only infect cornified hair above Adamson’s fringe

• Above Adamson’s fringe, Huxley’s layer of the inner root sheath no longer has trichohyalin granules

• Hair tends to retract from the inner root sheath above Adamson’s fringe

• The inner root sheath is fused and blue-gray at this level and trichohyalin granules are not seen

• The outer root sheath is composed of pink cells with peripheral palisading

Bulb

Anagen hairs have a stem and a bulb which produces the hair shaft, whereas telogen hairs lack an inferior segment. Telogen hairs are easily recognized in vertical sections, as the club hair and surrounding trichilemmal keratin give the impression of a flame thrower.

Fig 1-52 Hair, vertical section

Fig 1-53 Hair, transverse section

Fig 1-54 Telogen hair

Nail anatomy

Key features

• Nail plate

• Nail bed: between distal edge of lunula and the proximal edge of onychodermal band

• Framing portion: proximal nail fold, lateral nail folds, distal nail fold

• Ensheathing portion: “cuticle,” aka eponychium, hyponychium, solehorn, bed horny layer

Cuticle

• “Visible” cuticle (aka eponychium) is the thick keratinous material that borders the proximal nail fold and adheres to the nail plate

• “True” cuticle is located beneath the “visible” portion and is derived from the ventral part of the proximal nail fold

• The true cuticle is generally not seen, but it is sometimes visible as “flakes” of keratinous material parallel to the proximal nail fold

Hyponychium

• The space, epithelium, keratinous material ventral to the nail plate

Solehorn

• Subungual white to colorless keratin, extends from the distal nail bed underneath the onychodermal band to below the free, distal edge of the nail plate

Nail matrix

• Proximal part makes the surface of the nail; distal part makes the ventral nail plate

Lunula

• Visible portion of nail matrix

• Anchoring portion (mesenchyme)

Types of keratinization of the nail

Onychokeratinization (no granular layer)

• Hard keratin of nail plate

Onycholemmal keratinization

• Ventral part of proximal nail fold (+ granular layer), bed epithelium (the cuticle, bed horny layer, solehorn) (no granular layer)

Epidermoid keratinization

• Dorsal proximal nail fold, lateral folds, hyponychium

Types of inflammatory cells

Dermal dendrocyte

Key features

• Macrophage-type cells located in the dermis

• Many are factor XIIIa+, some are S100+

• Likely serve as antigen-presenting cells

Giant cell

Key features

• Cell with multiple nuclei, usually abundant cytoplasm

Types

• Foreign-body: nuclei are arranged haphazardly

• Langerhans: nuclei are arranged in a wreath shape

• Osteoclast-like: nuclei are arranged haphazardly and eccentrically; cytoplasm is deep pink with a scalloped border that molds to adjacent structures

• Touton: nuclei are arranged in a wreath with foamy cytoplasm peripherally

• Ringed siderophage: Touton giant cell with hemosiderin (characteristic of the fibrous histiocytoma type of dermatofibroma)

Fig 1-55 Osteoclast-like giant cell

Fig 1-56 Ringed lipidized siderophage (Touton giant cell with hemosiderin) in dermatofibroma

Fig 1-57 Langerhans cell histiocytosis

Histiocyte

Key features

• Epithelioid cell with a central, round/oval nucleus and surrounding cytoplasm

• Most are derived from a monocyte that takes up residence in tissue

• Functions include phagocytosis and antigen presentation

• Tend to coalesce in tissue with no intervening connective tissue

Langerhans cell

Key features

• Dendritic cells in epidermis and dermis

• CD1a+, S100+, peanut agglutinin, langerin+

• Nucleus is typically eccentric and may be reniform (kidney-shaped)

• Originate in bone marrow and function in presenting antigens to T cells

• Contain Birbeck granules, tennis racket-shaped rod and oval bodies seen on electron microscopy

Lymphocyte

Key features

• Round, dark nucleus, generally with no visible cytoplasm

• B-cell lymphocytes produce antibodies and are important for humoral immunity

• Natural killer (NK) cells are part of the innate immune system

• T-cell lymphocytes mature in the thymus and are important in cell-mediated immunity

• TH1 cells produce interleukin-1 (IL-1), IL-2, IL-12, and interferon-gamma (IFN-γ) and are important for cell-mediated immunity and function in activating macrophages

• TH2 cells produce IL-4, IL-5, and IL-10 and are important for humoral immunity

Mast cell

Key features

• “Fried-egg” appearance with central round nucleus and surrounding oval, bluish cytoplasm

• Contain metachromatic granules composed of heparin, histamine, tryptase, carboxypeptidase, leukotrienes

• Important in immediate-type hypersensitivity reactions

Neutrophil

Key features

• Predominant cell in acute infection

• Multilobulated nucleus

Eosinophil

Key features

• IL-5 induces eosinophil production

• Bilobed nucleus with granular cytoplasm containing major basic protein, eosinophil cationic protein, catalase, and other proteins

Plasma cell

Key features

• Eccentric nucleus with “clock face”

• Perinuclear pale space corresponding to rough endoplasmic reticulum (described by Hopf)

• Pink cytoplasm

Fig 1-58 Mastocytoma

Fig 1-59 Neutrophils in Sweet’s syndrome

Fig 1-60 Eosinophils in dermal hypersensitivity response

Fig 1-61 Plasma cells

Histochemical stains

The affinities of various dyes have been exploited as histochemical “special” stains to aid in identification of cell type, mucopolysaccharides, muscle, lipid, iron, melanin, calcium, elastin, collagen, amyloid, and infectious organisms. Hematoxylin and eosin (H&E) is the routine staining choice for microscopic interpretation resulting in blue nuclei and pink cytoplasm.

Connective tissue stains

Masson’s trichrome stain

• Differentiates collagen (blue-green) from smooth muscle (red)

• Example: scar versus leiomyoma

Important pitfall: Very young collagen can stain red

Fig 1-62 Normal skin with Masson’s trichrome

Verhoeff–Van Gieson stain

• Elastic fibers are black

• Examples: absence or reduction in scar, mid-dermal elastolysis, anetoderma, cutis laxa

• Example: distorted fibers in pseudoxanthoma elasticum

Fig 1-63 Beaded elastic fibers in elastofibroma dorsi (Verhoeff–Van Gieson)

Mast cell stains

Toluidine blue

• Stains mast cell granules metachromatically (i.e. the dye is blue but the granules stain purple)

• Also stains mucin (see below)

Fig 1-64 Metachromatic staining of mast cells in urticaria pigmentosa (toluidine blue)

Leder stain (naphthol ASD chloroacetate esterase)

• Mast cell cytoplasm stains red (not dependent on presence of granules)

• Also stains myeloid cells (example: leukemia cutis)

Fig 1-65 Mast cells in urticaria pigmentosa (Leder)

Carbohydrate stains

PAS (Periodic acid-Schiff)

• Stains glycogen, neutral mucopolysaccharides (such as basement membrane), and fungi red

• Glycogen is diastase labile, i.e. sections exposed to diastase before staining do not stain red with PAS reaction

• Useful in clear cell acanthoma, trichilemmoma

• Fungi and neutral mucopolysaccharides (basement membrane) are diastase resistant, i.e. stain red with PAS after diastase exposure

• Useful in tinea corporis, tinea versicolor, candida, basement membrane thickening of lupus erythematosus, thickened vessel walls in porphyria

• Acid mucopolysaccharides, such as hyaluronic acid, do not stain with PAS

Fig 1-66 PAS-positive basement membrane

Fig 1-67 PAS-positive fungi within the stratum corneum of tinea versicolor (PAS with light green counterstain)

Alcian blue

• Demonstrates acid mucopolysaccharides by staining them blue

• In normal skin, most mucin is sulfated acid mucopolysaccharide (heparin, chondroitin, and dermatan sulfates). In most pathologic states with increased dermal mucin, the mucin is predominantly non-sulfated hyaluronic acid

• Non-sulfated acid mucopolysaccharides (hyaluronic acid) stain with Alcian blue at pH 2.5 but not at pH 0.5

• Examples: follicular mucinosis, granuloma annulare, myxoid cyst, dermal mucin in lupus erythematosus

• Sulfated acid mucopolysaccharides stain with Alcian blue at both pH 2.5 and pH 0.5

• Alcian blue can be used with and without hyaluronidase to differentiate hyaluronic acid from other mucopolysaccharides

Colloidal iron

• Blue color indicates acid mucopolysaccharides

• As with Alcian blue, hyaluronidase digestion can be combined with colloidal iron to differentiate between hyaluronic acid and other mucosubstances

Fig 1-68 Focal mucinosis with colloidal iron

Toluidine blue

• Technically more complicated than Alcian blue and is used less often

• Metachromatically stains mast cells (see above)

Mucicarmine

• Stains acid mucopolysaccharides pink to red

• Stains the mucinous capsule of Cryptococcus neoformans pink to red

Fig 1-69 Pink capsule of Cryptococcus with mucicarmine

Amyloid

Congo red

• Amyloid stains brick red and displays “apple green” birefringence with polarized light

Fig 1-70 Congo red-stained amyloid in a salivary gland viewed with light microscopy

Fig 1-71 Congo red-stained amyloid in a salivary gland viewed with polarized light

Thioflavin T

• Sections must be examined with a fluorescent microscope causing amyloid to have a yellow to yellow-green appearance

Fig 1-72 Lichen amyloid with thioflavin T

Crystal violet

• Metachromatically results in a red-purple color of amyloid

Iron

Prussian blue (Perls stain)

• Ferric ions react to form a deep blue color

• Useful to distinguish melanin from hemosiderin

• Example: hemosiderin in pigmented purpuric dermatosis

• Does not demonstrate iron in intact red blood cells

Fig 1-73 Blue iron deposition from Monsel’s (ferric subsulfate) tattoo with Perls iron stain

Melanin

Fontana–Masson

• A silver stain that results in a black precipitate with melanin

Fig 1-74 Absence of melanin in vitiligo is contrasted with adjacent normal skin (Fontana–Masson)

Calcium

Von Kossa

• A silver stain that stains calcium salts black

• Examples: pseudoxanthoma elasticum, calcinosis cutis, calciphylaxis

Fig 1-75 Calcium deposition in the small vessels of the subcutaneous fat in calciphylaxis (Von Kossa)

Alizarin red

• Binds directly to calcium ions resulting in an orange-red color

Lipids

Oil red O

• Requires fresh frozen tissue, as lipid is removed during routine processing

Sudan black

• Requires fresh tissue

Osmium tetroxide

• Requires fresh tissue

Bacteria

Brown–Hopps

• A modification of the Brown–Brenn technique

• Gram-positive organisms stain blue and Gram-negative organisms stain red

Fig 1-76 Gram-positive cocci in an ulcer bed

Fungi

PAS (Periodic acid-Schiff)

• Fungi are PAS positive and diastase resistant (see above)

Fig 1-77 PAS-positive hyphae of onychomycosis

GMS (Grocott’s methenamine silver)

• Gray-black reaction with fungal wall

• Also stains Nocardia and Actinomyces

Fig 1-78 Aspergillus with GMS

Mycobacteria

Ziehl–Neelsen acid-fast stain; Fite acid-fast stain; Kinyoun’s acid-fast stain

• Mycobacteria appear bright red

• Fite is preferred for “partially acid-fast” organisms such as lepra bacilli, atypical mycobacteria, and Nocardia. Fite preserves color due to use of peanut oil before staining and gentle decolorization

Fig 1-79 Nocardia with Fite stain

Auramine–rhodamine

• Requires a fluorescent microscope

• Mycobacteria fluoresce reddish yellow

Fig 1-80 Mycobacterium with auramine–rhodamine as viewed with a fluorescent scope

Spirochetes

Warthin–Starry

(technically more difficult than the others, so sometimes referred to as the “worthless Starry”)

Dieterle

Steiner (modified Dieterle stain)

• Silver stains resulting in black spirochetes

• Examples: Lyme disease (around vessels and in dermal papillae), syphilis (in lower epidermis)

• Also stains Legionella, Bartonella, and Donovan bodies of granuloma inguinale

Fig 1-81 Spirochetes with Steiner stain

Other “special” stains

Giemsa

• Giemsa has many uses including highlighting myeloid and mast cell granules purplish blue

• Giemsa also stains many types of organisms including bacteria, Leishmania and Histoplasma

Fig 1-82 Metachromatic granules within mast cells of urticaria pigmentosa (Giemsa)

Immunohistochemical stains

Histochemical stains are now being supplemented by immunohistochemical techniques in fixed, paraffin-embedded tissue. Antibodies directed towards the antigen of interest are conjugated to peroxidases. Enzyme histochemical reactions for the peroxidase, such as diaminobenzidine method (DAB), are used to visualize the presence of enzyme–antibody–antigen complex fixed to tissue as a stable brown product. 3-amino-9-ethylcarbazole (AEC) can alternatively be used giving a red reaction, but does not archive as well.

Immunopathologic antibodies are used in dermatopathology to differentiate carcinoma, melanoma, sarcoma, neural neoplasms, and lymphomas. A panel of antibodies is generally recommended rather than a single stain because aberrant staining is common in neoplasms. Independent and internal controls should be assessed to assure proper reactivity.

Epithelial markers

AE1/AE3

• Cocktail of high- and low-molecular-weight monoclonal cytokeratin antibodies

• Expressed in the epidermis and adnexal epithelium

• Stains all epithelial tumors (squamous cell carcinoma and adnexal tumors) and generally excludes mesenchymal, melanocytic, and hematopoietic tumors

• Also stains epithelioid sarcoma, synovial sarcoma and mesothelioma

Fig 1-83 AE1/AE3 positivity of squamous cell carcinoma

CK polyclonal keratin (pankeratin)

• Polyclonal cocktail that offers greater sensitivity than AE1/AE3

p63

• Expressed in basal and spinous cells of the epidermis, germinative cells of sebaceous glands, and myoepithelial cells of the sweat glands

• Lack of reactivity in metastatic carcinoma assists in differentiation from primary cutaneous adnexal neoplasms

• Useful in identification of cutaneous spindle cell squamous cell carcinoma from other spindle cell neoplasms

Fig 1-84 p63 in spindle cell squamous cell carcinoma

CAM5.2

• CAM5.2 detects low-molecular-weight cytokeratins present in most glandular neoplasms without staining the epidermis or stratified squamous epithelium

• Marks Paget’s disease and extramammary Paget’s disease

CK7

• Used in determining the origin of metastatic carcinoma

• In general a marker of non-gastrointestinal adenocarcinoma (see Table 1.1)

• Marks Paget’s disease and extramammary Paget’s disease

Fig 1-85 CK7 reactivity of metastatic breast carcinoma

CK20

• Marks Merkel cell carcinoma predominantly in a paranuclear pattern and distinguishes from metastatic oat cell carcinoma of the lung that is typically negative

• Used in determining the origin of metastatic carcinoma (see Table 1.1)

• Marker of gastrointestinal adenocarcinoma

• Highlights sparse Merkel cells within the basaloid islands of desmoplastic trichoepithelioma but not basal cell carcinoma

Fig 1-86 Paranuclear dot with CK20 in Merkel cell carcinoma

Table 1-1

Metastatic carcinoma of unknown origin

	CK20−	CK20+
CK7+	Breast, lung, mesothelioma	Bladder, pancreatic
CK7−	Hepatocellular, prostate, renal, neuroendocrine and squamous carcinoma of lung	Colon

CDX2

• Marker of intestinal adenocarcinoma

• Useful for diagnosis of cutaneous metastatic colon adenocarcinoma and extramammary Paget’s disease associated with an underlying colorectal tumor

Fig 1-87 Metastatic colon carcinoma with CDX2

Renal cell carcinoma (RCC)

• Positive in most cutaneous metastases from renal cell carcinoma and negative in other clear cell tumors of the skin

Thyroid transcription factor (TTF-1)

• Useful in the small blue cell tumor differential diagnosis

• Reactive in metastatic small cell lung carcinoma and negative in Merkel cell carcinoma

Fig 1-88 TTF-1 in metastatic adenocarcinoma of the lung

Epithelial membrane antigen (EMA)

• Highlights normal sebaceous and sweat glands

• Positive in sebaceous carcinoma, Paget’s and extramammary Paget’s

• Positive in squamous cell carcinoma but negative in basal cell carcinoma

Fig 1-89 EMA positivity in sebaceous epithelioma

Carcinoembryonic antigen (CEA)

• Sweat glands are immunoreactive

• Positive in sweat gland neoplasms, Paget’s, extramammary Paget’s, and most adenocarcinomas

Fig 1-90 CEA-positive pagetoid cells of Paget’s disease

Adipophilin

• Expressed in lipid droplets of sebaceous and xanthomatous lesions

• Can help distinguish sebaceous carcinoma from squamous cell carcinoma and basal cell carcinoma

Ber-EP4

• Marks most epithelial cells, but not those undergoing squamous differentiation

• Positive in basal cell carcinoma and negative in squamous cell carcinoma

Fig 1-91 Infiltrating basal cell carcinoma with Ber-EP4

Mesenchymal markers

Desmin

• Positive staining in skeletal and most smooth muscle

• Generally negative in vascular smooth muscle (including glomus cells)

Fig 1-92 Desmin-positive piloleiomyoma

Smooth muscle actin (SMA)

• Positive in smooth muscle, including vascular smooth muscle

• Examples: glomus tumor, leiomyosarcoma

Fig 1-93 Piloleiomyoma with SMA. Note positive staining of normal surrounding vascular smooth muscle in contrast to the absence of reactivity with desmin in Figure 1.92

CD34

• Marker of vascular endothelium and hematopoietic progenitor cells

• Positive in dermatofibrosarcoma protuberans and negative in dermatofibroma

• Positive in spindle cell lipoma, sclerotic fibroma, solitary fibrous tumor, superficial acral fibromyxoma, pleomorphic fibroma, and pleomorphic hyalinizing angiectatic tumor

• Decreased staining in morphea

• Increased staining in nephrogenic systemic fibrosis

• Stains connective tissue around normal hair follicles

• Typically highlights the stroma of trichoepitheliomas but not basal cell carcinomas

Fig 1-94 Dermatofibrosarcoma protuberans with CD34

Fig 1-95 Decreased CD34 expression in the interstitial dermis of morphea

Factor XIIIa

• Highlights a population of dermal dendritic cells

• Positive in dermatofibroma and negative in dermatofibrosarcoma protuberans

• Positive in fibrous papule of the face

Fig 1-96 Dermatofibroma with factor XIIIa

CD31

• Helpful in confirming vascular origin of tumors

• More specific vascular marker than CD34

Fig 1-97 CD31-positive angiosarcoma

D2-40 (podoplanin)

• Lymphatic endothelial marker

• Increases detection of lymphovascular invasion

• Lack of reactivity in metastatic carcinoma assists in differentiation from primary cutaneous adnexal neoplasms

Fig 1-98 Intralymphatic invasion of melanoma with D2-40

UEA-1 (Ulex europeus agglutinin)

• Binds vascular endothelium

GLUT1 (glucose transporter)

• Expressed in endothelial cells with blood–tissue barrier function as in placenta

• Positive in infantile hemangiomas (except RICH and NICH) and negative in vascular malformations

• Also stains perineurial cells and perineurioma

Fig 1-99 GLUT1 in infantile hemangioma

Vimentin

• Stains mesenchymal cells, endothelial cells, fibroblasts, melanocytes, lymphocytes, macrophages but does not react with keratinocytes or other epithelium

• General marker of sarcomas

• Excludes most carcinomas except rare spindle cell carcinomas and synovial sarcoma

Neuroectodermal markers

S100

• Reactivity is observed for neural crest-derived cells and some mesenchymal lines

• Stains melanocytes, Langerhans cells, sweat glands, nerves, schwann cells, myoepithelial cells, fat, muscle, and chondrocytes

• Useful in differential diagnosis of spindle cell neoplasms

• Examples: desmoplastic melanoma, Langerhans cell histiocytosis, granular cell tumor, Rosai–Dorfman disease

Fig 1-100 Granular cell tumor with S100

S100A6 (calcyclin)

• Member of the S100 protein superfamily

• Stains melanocytes, Schwann cells, Langerhans cells, and dermal dendrocytes thus expressed in nevi, particularly those with type C Schwann cell-like features, and some neural and fibrohistiocytic tumors

• Positive in cellular neurothekeoma while S100 is negative

• Reactive in most atypical fibroxanthomas but is not specific and also stains other entities in the cutaneous spindle cell tumor differential diagnosis

• Has been reported to stain Spitz nevi strongly and diffusely but spitzoid melanomas have weak or patchy staining in the dermis only

Fig 1-101 S100A6 in atypical fibroxanthoma

HMB-45

• Premelanosome marker

• Loss of staining of melanocytes with descent into the dermis is a manifestation of loss of premelanosomes. As such, it serves as a marker of normal maturation

• Loss of staining in deep dermal component of most benign nevi, but uniform staining of blue nevi

• Does not stain desmoplastic melanoma reliably

Fig 1-102 HMB-45-positive staining throughout a blue nevus

Melan-A and Mart-1

• Two different antibodies that stain the same epitope

• Stain melanocytic lesions

• Do not stain desmoplastic melanoma reliably

Fig 1-103 Obscured nevus cells in the lymphocytic infiltrate of halo nevus with Mart-1

p75 (nerve growth factor receptor)

• Early neural crest marker

• Expressed in type C (spindled) melanocytes and schwann cells

• Sensitive marker for spindle cell and desmoplastic melanoma

Fig 1-104 p75 in desmoplastic melanoma

Microphthalmia-associated transcription factor (MITF)

• Essential in development and survival of melanocytes

• Nuclear melanocytic marker

• Positive in cellular neurothekeomas

Fig 1-105 Nuclear reactivity of melanocytes in melanoma in situ with MITF

Sox-10

• Nuclear marker of Schwann cells and melanocytes

• Sensitive marker of melanoma, including conventional, spindled, and desmoplastic types

Fig 1-106 Nuclear reactivity of melanocytes in desmoplastic melanoma with Sox-10

Neuroendocrine markers

Neuron-specific enolase (NSE)

• Positive in neuroendocrine cells, neurons, and tumors derived from them

• Positive in so many other cell lines that it is sometimes referred to as “non-specific enolase”

Fig 1-107 NSE in Merkel cell carcinoma

Chromogranin

• Positive in Merkel cell carcinoma

Synaptophysin

• Positive in Merkel cell carcinoma

Hematopoietic markers

CD45Ra (LCA)

• Leukocyte common antigen (LCA) is a general marker of hematolymphoid differentiation

• LCA is present on all hematopoietic cells and their precursors with the exception of maturing erythroids and megakeratocytes

CD45Ro (UCHL-1)

• Mature T cells

CD20

• B-cell antigen (often absent in plasma cells)

• Positive in B-cell lymphomas and negative in T-cell lymphomas

• Target for rituximab. Loss correlates with rituximab resistance

Fig 1-108 CD20 positivity in intravascular lymphoma using amino-9-ethylcarbazole (AEC) as the enzyme in the enzyme–antibody–antigen complex, resulting in a red product

CD10 (CALLA)

• Common acute lymphoblastic leukemia antigen (CALLA) is an early marker of B-cell differentiation

• Useful in differential diagnosis of B-cell lymphoproliferative disorders (see Table 1.2)

• Positive in periadnexal mesenchymal cells, staining only the stroma of trichoblastomas but the epithelial cells of basal cell carcinoma

• Expressed in most atypical fibroxanthomas but not uncommonly seen in the other tumors in the differential diagnosis of cutaneous spindle cell tumors

• Marker of renal cell carcinoma but also expressed in other cutaneous clear cell lesions including: balloon cell nevi, clear cell hidradenoma, and sebaceous tumors

Fig 1-109 CD10 in atypical fibroxanthoma

CD79a

• Plasma cell and B-cell marker

CD138 (syndecan-1)

• Plasma cell marker

Fig 1-110 CD138 in plasmacytoma

CD3

• Pan-T-cell marker

• Positive in T-cell lymphomas but negative in B-cell lymphomas

Fig 1-111 CD3-positive lymphocytes of subcutaneous T-cell lymphoma

CD4

• T-helper lymphocytic marker

Fig 1-112 CD4-positive epidermotropic cells of mycosis fungoides

CD8

• T-cell cytotoxic/suppressor marker

CD5

• Pan-T-cell marker like CD3 but aberrant loss in CTCL is common

• Positive in mantle cell lymphoma and infiltrates of chronic lymphocytic leukemia

CD30 (Ki-1, BERH2)

• Originally identified on Reed–Sternberg cells of Hodgkin’s disease

• Positive in activated lymphocytes of anaplastic large cell lymphoma and lymphomatoid papulosis

• Many positive cells may be seen in scabies nodules and chronic tick bites

Fig 1-113 CD30-positive anaplastic large cell lymphoma

CD7

• Immature T-lymphocyte antigen

• Most commonly lost antigen in T-cell lymphoma

Fig 1-114 Aberrant loss of CD7 in epidermotropic cells of mycosis fungoides (compare with Figure 1.112)

CD56

• Marker of NK cells and subsets of T cells

• Stains blastic plasmacytoid dendritic cell neoplasm (formerly known as blastic NK/T-cell lymphoma or CD4+/CD56+ hematodermic neoplasm)

CD68 (KP-1)

• Reactive in virtually all monocyte/macrophage cells

Fig 1-115 CD68-positive histiocytes of granuloma annulare

CD163

• Reactive in monocytes and macrophages

CD123

• Marker of plasmacytoid dendritic cells

• Positive in blastic plasmacytoid dendritic cell neoplasm

Myeloperoxidase

• Major constituent of granules of neutrophilic myeloid cells

• Marker for acute myeloid leukemia

Fig 1-116 Myeloperoxidase reactivity in leukemia cutis using amino-9-ethylcarbazole (AEC) as the enzyme in the enzyme–antibody–antigen complex, resulting in a red product

ALK-1

• Anaplastic lymphoma kinase expressing chromosomal translocation t (2,5)

• Positive in most systemic anaplastic large cell lymphoma and negative in primary cutaneous anaplastic large cell lymphoma

• Those few patients with ALK-1-negative systemic anaplastic large cell lymphoma have a poor prognosis

Kappa/lambda

• Normally expressed in a ratio of two-thirds kappa to one-third lambda

• 10-fold deviation from this ratio suggests a clonal B-cell proliferation

CD117 (c-Kit)

• Expressed in mast cells and melanocytes

• In nevi and primary melanoma there is a decrease in expression in the dermal component

• Typically lost in metastatic cutaneous melanoma

Fig 1-117 CD117 in urticaria pigmentosa

CD1a

• Stains Langerhans cells

• Examples: Langerhans cell histiocytosis

Fig 1-118 CD1a in Langerhans cell histiocytosis

Langerin (CD207)

• Surrogate marker for presence of Birbeck granules in Langerhans cells

CD43 (Leu-22)

• Pan-T-cell marker

• Aberrant coexpression with B-cell marker CD20 is strongly suggestive of B-cell lymphoma

BCL2

• An oncogene that inhibits apoptosis

• Useful in differential diagnosis of B-cell lymphoproliferative disorders (see Table 1.2)

• Most basal cell carcinomas reveal diffuse BCL2 staining, whereas trichoepitheliomas only show staining of the outermost epithelial layers of the tumor islands

Table 1-2

Cutaneous B-cell lymphoproliferative disorders

PCMZL, primary cutaneous marginal zone lymphoma

PCFCL, primary cutaneous follicle center lymphoma

2° CFL, secondary cutaneous follicular lymphoma

PCLBCL-leg, primary cutaneous large B-cell lymphoma, leg type

NOTES: Immunoreactivity refers to the malignant cells in all cases except the reactive germinal center. BCL2 also stains background normal T cells.

^a Plasma cells are positive with MUM-1 but the neoplastic B cells are negative in PCMZL.

^b Only 10–20% of PCFCL are BCL-2 positive.

^c Most PCFCL have a diffuse histologic pattern and are CD10 negative but cases with a follicular pattern are CD10 positive.

^d The BCL6-positive malignant cells are outside the follicle.

Multiple myeloma oncogene-1 (MUM-1)

• Expressed in plasma cells, activated T cells, and subset of germinal center cells

• Distinguishes primary cutaneous diffuse large B-cell lymphoma, leg type from diffuse follicle center lymphoma

• Reactivity reported in anaplastic large cell lymphoma, lymphomatoid papulosis, and blastic plasmacytoid dendritic cell neoplasm

• Also expressed in benign and malignant melanocytic cells

Fig 1-119 MUM-1 in diffuse large B-cell lymphoma, leg type

CD21

• Follicular dendritic cell marker

• Highlights residual follicle in lymphoma

• CD23 has a similar staining pattern

BetaF1

• Identifies αβ T cells

Table 1-3

Spindle cell neoplasms

Table 1-4

Small blue cell tumor

Table 1-5

Tumors with intraepidermal buckshot scatter (pagetoid spread)

Infectious disease markers

Specific immunohistochemical antibodies to viral, bacterial, fungal, and parasitic antigens are available for detection and identification of the causative agents in many infectious diseases including HHV8, HSV, VZV, CMV, EBV, Bartonella, Rickettsia, Treponema, Borrelia, Aspergillus, Leishmania, and others.

Fig 1-120 Immunohistochemical identification of Treponema pallidum in syphilis

Fig 1-121 Immunohistochemical identification of HHV8 in Kaposi sarcoma

Fig 1-122 Immunohistochemical identification of VZV in multinucleate giant cell

Proliferation markers

Mib-1(Ki-67)

• Nuclear proliferation marker

• Not cell type specific

• Expressed in all active phases of the cell cycle (G1, M, G2, and S)

• Pattern and number of reactive melanocytes can be helpful in diagnosing melanocytic lesions

Fig 1-123 Mib-1 positivity in malignant melanoma. Positive background reactive lymphocytes must be distinguished from Mib-1-positive melanocytes by cytology or double staining technique

pHH3

• Mitotic marker that only stains cells in the M phase of the cell cycle

• Helps differentiate mitoses from apoptotic or hyperchromatic nuclei

Fig 1-124 pHH3-positive mitotic figures in malignant melanoma

Transport media

Routine

For most purposes, 10% buffered formalin is the recommended fixative. The length of time for fixation depends on the specimen size: 1–2 h per mm of thickness is required.

Electron microscopy

Glutaraldehyde is the preferred fixative for electron microscopy.

Immunofluorescence

For tissue that is not flash-frozen, Michel’s medium (ammonium sulfate) is the preferred transport media for immunofluorescence. Normal saline also performs well.

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The basics: Diagnostic terms, skin anatomy, and stains

The basics

Diagnostic terms, skin anatomy, and stains