Cytology of lumps and bumps (Proceedings)

Lumps and bumps that are cutaneous or subcutaneous often lend themselves very well to cytologic evaluation. They are easy to get to and most animals don't require sedation or anesthesia for you to obtain these samples. Although a definitively diagnostic sample isn't always obtained, the investment of time and equipment is minimal, and may give you the answer quickly and inexpensively.

Obtaining the sample

Proper sample collection and handling are key to getting a diagnostic sample.

What do you need?

• 21-25 gauge needles

• 3-20 ml syringes

• Microscope slides

• Stain – can be Wrights stain, Wrights-Giemsa, Diff-Quick, etc.

• A good microscope with an oil-immersion objective, immersion oil, and lens cleaner to clean it all off with!

• A good cytology book for reference

The skin is prepped as for a venipuncture. With a needle connected to a syringe (often 22 g needle with a 3-6 ml syringe), immobilize the lesion and insert the needle, apply suction, release suction, and withdraw the needle. Depending on the mass, the needle may be redirected within the lesion (without pulling back out through the skin. Don't expect to see material in the barrel of the syringe-it's collecting in the needle.

After removing the needle, disconnect it from the syringe, fill the syringe with air, reconnect the needle and shoot out (gently!) the contents of the needle onto the middle of a glass slide.

An alternative method involves "tattooing" a needle into the mass (making many quick jabs). A syringe filled with air is then attached to the needle and the sample within the needle is expelled. This technique may lead to less blood contamination and cell rupture in the sample.

After expelling the sample on a slide, smears are made by placing a clean glass slide on the surface of the sample, either end to end or at right angles to each other, and gently pulling the slides apart, allowing the material to spread from the weight of the slide. NOTE-you're not trying to squish the cells (they are fragile-treat them nicely), just spread them out into a layer that is one cell thick so they stain evenly and you can evaluate individual cells. Make several slides if possible.

All slides should be allowed to air-dry before staining. No other fixation is needed and may actually damage the sample if used. Don't heat fix cytology slides (that's for gram-staining only!) If you're submitting samples to an outside laboratory, it is best to send unstained, air-dried smears (as well as any stained samples) separately from any formalin-fixed samples you may also be submitting as the formalin fumes can destroy cytologic detail. If you're staining them to review in-house, remember that you can use the same quick stains used for blood smears, but if the material is thick, you may need to both fix and stain for longer.

Slide interpretation

Slides should first be reviewed with the microscope on low (scanning) power to determine whether there are enough cells present, whether they are in good condition, and to identify areas of good cellularity for closer review. Further viewing is usually performed on high dry or oil immersion.

Most lumps and bumps may initially be classified as inflammatory or neoplastic. Then, a more specific diagnosis can be sought based on type of inflammation or neoplasia. In addition, the presence or absence of microorganisms or specific cell type in a neoplastic lesion can be extremely useful in deciding treatment and prognosis.

Inflammatory lesions

Inflammatory lesions may be classified as due to infectious agents, or as non-infectious. Infectious agents that may cause lumps or bumps include bacteria, fungi, and protozoa.

Bacterial infections are associated with a predominantly neutrophilic inflammatory response. Neutrophils often appear degenerate (swollen nuclei) when bacteria have been phagocytized. Bacteria are most often seen within neutrophils, though there may also be free bacteria. They can include rods, cocci, filamentous bacteria and negative-staining Mycobacteria spp. If bacteria are present, but large numbers of neutrophils aren't, or if all the bacteria are extracellular, be aware that the bacteria may be contaminants of the sample, rather than pathogens.

Fungal infections are usually associated with neutrophils and macrophages, or pyogranulomatous inflammation. Lymphocytes, plasma cells, and occasionally eosinophils may also be present. Fungal agents that may be associated with cutaneous infections include Sporothrix schenckii, Histoplasma capsulatum, Blastomyces dermatitidis, Cryptococcus neoformans, and Cocciocioides immitis. There are also some fungi that are saprophytes (common in soil or vegetation), that may form hyphae when introduced into or under the skin.

Protozoal agents that may cause subcutaneous lesions include Leishmania spp. and Toxoplasma. Both have a very characteristic appearance in cytologic samples.

Non-infectious inflammatory lesions are caused by a number of processes such as vaccination (usually dominated by lymphocytes, plasma cells and macrophages), foreign bodies (commonly neutrophils, macrophages, eosinophils) and fat necrosis (macrophages, +/- neutrophils and lymphocytes). Eosinophil-predominant lesions include eosinophilic granulomas and allergic reactions

Neoplastic lesions

Neoplasms that can cause cutaneous and subcutaneous lesions include those of epithelial and mesenchymal origin as well as discrete cell tumors. It's important to try to classify which of these types a lesion falls into, and then to determine whether the tumor is malignant or benign.

Cutaneous and subcutaneous lesions that are neither inflammatory nor neoplastic

Some of these lesions are fluid-filled, such as seromas, hematomas, hygromas, sialoceles, and cysts. In these cases, cytologic examination of the cells in the fluid is often definitively diagnostic. Sometimes, the wall of the lesion, rather than the fluid it contains, must be aspirated to find diagnostic cells.

Cytologic features of these lesions are summarized briefly below:

• Seromas/hygromas: low cellularity, few mononuclear cells or macrophages

• Hematomas: erythrocytes and macrophages containing RBC's or hemosiderin, a breakdown product of hemoglobin; leukocytes may also be present

• Sialoceles: large, round, highly vacuolated (foamy) cells and pale blue amorphous material (mucin); sometimes associated inflammatory cells

• Cysts: often no cells are seen

Epidermal cyst is an example of a non-inflammatory and non-neoplastic lesion that does not contain fluid. Cytologic findings in an aspirate of an epidermal cyst generally include moderate numbers of superficial squamous epithelial cells, lots of amorphous blue material, and sometimes large, angular, negative-staining cholesterol clefts. These lesions sometimes stimulate surrounding inflammation, so inflammatory cells may also be seen.