The skin scrape was first employed in the late 1940s by Tzanck in the differential diagnosis of bullous diseases, and is still mainly used for sampling superficial lesions.³

A blunt or sharp curette, a double-ended elevator or a scalpel blade can be used. The skin should be cleansed before the procedure. The keratotic surface and any crusts must be removed completely to obtain a satisfactory representative smear.⁴ Samples from superficial, crusted lesions should be taken from the periphery of the lesion, as the centre may be inflamed and necrotic. It is important to avoid scraping unnecessarily deep as this will lead to bleeding and possibly also scarring. In non-crusted nodular lesions, it is possible to cut into the lesion, either removing the top and scraping the exposed surface, or scraping the incision area to obtain an adequate sample (skin slit smear).⁵

The material is deposited on a glass slide and spread directly with a second slide. If material is abundant, several slides should be prepared. It is important that the cells are evenly spread on the slide so that a thin layer of material is achieved (Fig. 28.2). Small fragments of tumour tissue can be placed on one slide and a second slide then placed directly on top. Firm vertical pressure is applied and the slides are separated horizontally, spreading the cells evenly.²

Fig. 28.2 (A) The cellular material is placed between two glass slides. The slides are then drawn apart horizontally making two smears. (B) The cellular material is placed on a glass slide and smeared with a smaller glass slide.

Brush cytology is also used for sampling superficial lesions.⁶ A gynaecological cytobrush may be used. The material is deposited on the slide avoiding vigorous brushing backwards and forwards which may damage the cells.

This technique may be used successfully in ulcerating lesions and the cut surface of biopsies. A clean, dry microscope slide should be firmly pressed against the lesion after removal of any crusts. Slight abrasion of the surface exposes viable tissue, freeing up tissue fragments and individual cells.

Imprints can also be made from shave or punch biopsies. Excess blood should be carefully removed and the biopsy gently pressed or rolled on the slide.²

Fine needle sampling with aspiration (FNA) or without aspiration (FNS) may be employed to sample skin nodules and deeper lesions. The procedure does not require sterile conditions. Local anaesthetic is not necessary and may, in some instances, make it difficult to obtain adequate cellular material. A narrow gauge needle, <0.7 mm (22–23G) in diameter, is used. The needle is moved back and forth, sometimes almost tangentially to the skin surface, aiming at the raised edges of an ulcer or centre of a nodule. The needle contents are ejected and smeared in a thin layer on a glass slide (Fig. 28.2).⁷ Material may also be deposited in a variety of media for special procedures such as liquid-based cytology (LBC) flow cytometry, polymerase chain reaction (PCR), or microbiology.

Sampling without aspiration, allowing the material to enter the needle by capillary attraction, is particularly suitable for cellular nodules such as lymph nodes or melanoma metastases.

The Papanicolaou stain (PAP) requires immediate fixation in alcohol. Nuclear chromatin patterns are easily discerned and the degree of maturation/keratinisation of squamous cells is readily appreciated. Mucin may not be seen so easily.

May-Grünwald Giemsa (MGG) staining is used on air-dried smears. Nuclear detail is less readily discerned though mucin and other extracellular substances may be easily seen. Modified MGG stains are available for rapid staining.

Wet films may be also stained immediately with a drop of 0.5% methylene blue followed by coverslipping to spread the stain (Tzanck’s stain).³ Ideally, smears should be prepared for PAP and MGG staining and air-dried smears or cell suspensions should be kept for special tests.

Immunocytochemistry can be performed on cytological smears or cytospin preparations, preferably on coated glass slides. Cells from stained or unstained smears may be used for PCR and may be particularly useful in lymphoma diagnostics (see Chs 13, 14, 34). Cell suspensions may be utilised for PCR or flow cytometry. Air-dried smears are eminently suited for in situ hybridisation techniques with either fluorescent or chromogenic markers. ⁸

It is unusual for a pustular lesion clinically considered to be a primary or secondary bacterial infection to be sampled for cytological diagnosis. However, when a scraping or aspirate of pus is taken, samples should be sent for bacterial culture, PCR and extra smears made for special stains such as Gram or Ziehl Neelsen.

Fungal elements are usually a coincidental finding in skin smears (Fig. 28.5). However, cases of fungal cutaneous lesions diagnosed by FNA have been reported in the literature.^17,18

Fig. 28.5 Budding yeast forms in actinic keratosis. Skin scrape (PAP).

At least five groups of viruses can affect the skin; two groups produce lesions with characteristic cytological features that can be reliably diagnosed by skin cytology: the herpes virus group and the poxvirus group. Human papillomavirus causes skin warts. However, these are seldom sampled for cytology except when they occur in the vulvar region. The cytological changes are similar to those of human papillomavirus infection of the lower genital tract (see Chs 21, 23).

Herpes virus infection

The herpes virus group comprises Herpes simplex types 1 and 2 and Varicella zoster. Despite clinical differences, early stages show grouped or isolated vesicles on an inflamed base, that later become covered by crusts. It is impossible to distinguish these entities cytologically; however, a number of diagnostic tests, including PCR, may be used. Samples should be taken from the vesicle at an early stage to avoid degenerative changes and superimposed infection (Fig. 28.6).^2,9

Fig. 28.6 Herpes virus infection. Multinucleated giant cells with ground glass nuclei and nuclear moulding. Skin scrape (PAP).

Cytological findings: herpes virus

• Multinucleated cells measuring between 20 and 30 μm in diameter with up to 30 nuclei

• Nuclear moulding and margination of chromatin

• Pale, eosinophilic, ground glass-like intranuclear inclusion bodies

• Inflammatory cells.

Diagnostic pitfalls: herpes virus

• The large eosinophilic inclusions may be confused with nucleoli

• Multinucleated giant cells in pemphigus vulgaris lack nuclear inclusions and moulding and are accompanied by acantholytic cells.

Molluscum contagiosum

Molluscum contagiosum is caused by a pox-type virus causing isolated, hard, dome-shaped papules with umbilicated centres, occurring mostly in children. Specimens can be obtained either by compression of the lesion to extrude the central keratinous material or, preferably, by removing cells from the top of the papule with a small curette. Vaccinia and Orf share cytological features with molluscum contagiosum (Fig. 28.7).²⁰

Fig. 28.7 Molluscum contagiosum. Squamous cells with eosinophilic ‘molluscum bodies’. Skin scrape (PAP).

Cytological findings: molluscum contagiosum

• Numerous single, round squamous cells with large eosinophilic (PAP), intracytoplasmic aggregates of viral particles (‘molluscum bodies’) giving a mosaic appearance to the cytoplasm

• The nucleus is pushed to the periphery of the cell and is crescent-shaped.

Pemphigus vulgaris can appear in all areas of the skin and mucosa. Bullae form as the cells lose their prickles and intercellular clefts form into which fluid from the underlying corium passes. The disease presents as thin, loose blisters with serous content which easily break leaving an eroded surface with crust formation. Direct immunofluorescence on a Tzank smear has been shown to be comparable to biopsy in diagnosing early pemphigus vulgaris and is particularly useful in lesions in the oral cavity (Fig. 28.9).²²

Fig. 28.9 Pemphigus. Rounded single cells with perinuclear halos. Skin scrape (PAP).

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