DVM, MS, DACVD
Sarah Bartlett, DVM, MS, DACVD, graduated from the University of Florida College of Veterinary Medicine and completed an internship at Affiliated Veterinary Specialists in Maitland, Florida. She completed her dermatology residency at Animal Dermatology Clinic in Marietta, Georgia. She has recently returned to Animal Dermatology Clinic in Marietta after working in Florida for 2 years. She is a member of the ACVD Credentials Committee, and her clinical interests include allergic and immune-mediated skin disease.Read Articles Written by Sarah Bartlett
Biopsy is often an important diagnostic step in determining the cause of cutaneous disease. An accurate diagnosis requires appropriate timing of the biopsy, careful site selection and biopsy technique, selection of a dermatopathologist, good communication between the clinician and the pathologist, and correlation of the results with the clinical picture.
In general, biopsy is indicated when a dermatosis is acute and severe, is unusual in appearance, or is not responding to appropriate therapy after 3 weeks. Persistent ulcerations, suspected neoplasms, or vesicular dermatitis should always be biopsied. Biopsy should also be chosen for suspected conditions easily diagnosed by biopsy, such as follicular dysplasia, zinc-responsive dermatosis, sebaceous adenitis, and immune-mediated disease. Biopsy is also indicated in any suspected condition for which therapy is expensive, dangerous, or time consuming.1
At least 3 to 4 biopsy sites should be chosen, unless only a focal lesion is present. A variety of types of lesions should be sampled. Primary lesions, such as pustules, nodules, papules, vesicles, bullae, and tumors, are more likely to provide an answer. However, secondary lesions may also be diagnostic, especially crusts. Scale, comedones, alopecic areas, the margins of ulcers, and hypopigmentation (especially at the nasal planum or mucocutaneous junctions) may also be helpful. Typical locations for a suspected disease should be biopsied, but any atypical locations that are affected should also be included.
Timing of the biopsy is important. Active disease is required in order to obtain an accurate diagnosis, and in some cases lesions may be transient. If pemphigus foliaceus is suspected and pustules were previously present, it may be helpful to wait for pustule recurrence. In general, it is better to consider biopsy before the development of secondary changes that may cloud the diagnosis. Withdrawal of corticosteroids and treatment of bacterial infections for several weeks prior to taking biopsy samples is usually necessary to avoid masking of primary pathology.
Biopsies can often be performed without sedation. However, sedation may be necessary to minimize patient movement and to increase patient comfort. Biopsies from the nasal planum and paws almost always necessitate deep sedation or anesthesia. I often use a combination of dexmedetomidine and butorphanol for biopsy in these sensitive areas.
Biopsy sites are typically outlined with a permanent marker as a guide for where lidocaine should be injected (Figure 1). The same marker may be used to draw a line in the direction of the hair growth to ensure proper orientation of the hair follicles when the sample is cut. This is most important for alopecic lesions, where the pathologist cannot determine the direction of hair growth by sight. Biopsy sites should never be scrubbed because diagnostic material may be removed. Gentle clipping may be necessary in long-haired patients; care should be taken not to let the clippers touch the skin surface.
Lidocaine 2% is typically used for local anesthesia. This may be mixed with sodium bicarbonate in a 10:1 ratio to reduce the stinging sensation that lidocaine can cause. Inject lidocaine subcutaneously with a 25-gauge needle into the skin surrounding the lesion. Avoid injection into the dermis because it can cause an artifactual appearance of dermal edema. Do not inject lidocaine directly into lesions of the panniculus or if tissue cultures are to be performed. In these cases, use regional or general anesthesia. The total dose of lidocaine should not exceed 5 mg/kg in dogs or 2.5 mg/kg in cats.1 In small patients with many biopsy sites, the lidocaine may be mixed 1:1 with sterile saline to reduce the amount used, creating a 1% solution, which has also been shown to be effective.2
The sample may be obtained with a punch, incisional, or excisional biopsy.
Punch biopsies are most often performed, and typically a 6-mm punch is used. Reserve 4-mm punches for pinnae, the nasal planum, or footpads of small dogs and cats. Obtain punches from the center of a lesion unless it is an ulcer, and center small lesions within the punch. Do not include any substantial amount of normal skin within the specimen because when the tissue section is cut in half at the lab the lesion may be missed. Use a fresh punch for each animal, and if the punch dulls during sample collection use a new one for subsequent punches. Dull punches cause tissue compression and artifact. Rotate the punch in one direction to prevent shear artifact. Decreased resistance is felt once the punch has reached the subcutis. Use a gauze square to blot any excess blood from the edge of the excised tissue; do not blot on top of the lesion. Once the subcutis is reached, remove the punch and gently grasp the tissue section (do not squeeze) with tissue forceps or a 25-gauge needle, and cut the attachment with iris scissors (Figure 2). A single cruciate suture easily closes the defect left by a 6-mm biopsy punch.
Incisional and Excisional Biopsy
An incisional elliptical biopsy is preferred for ulcers, vesicles, and bullae. The shearing force of a punch may damage vesicles and bullae, and the adjacent unaffected skin must be included for these lesions because the diagnostic tissue resides in the margin between affected and unaffected areas. Keep in mind that samples are cut in half longitudinally during processing. Incisional or excisional biopsies are performed for suspected disease of the panniculus and for tumors. A punch often does not reach deep enough to obtain the diagnostic sample for diseases of the panniculus.
Biopsy for Culture
This article is primarily focused on biopsy for histopathology. However, in some cases, such as nodular disease, cellulitis, and diseases with fistulous tracts, biopsy for aerobic, anaerobic, and/or fungal culture may be needed. When fungal or unusual bacterial diseases (eg, mycobacteriosis, bacterial pseudomycetoma, actinomycosis, actinobacillosis, nocardiosis) are suspected, tissue biopsy specimens should be submitted and the lab should be notified of the suspected disorder. In these cases, disinfect the surface to avoid contamination with surface bacteria and observe sterile technique. Submit tissue in a sterile container, typically a plain blood collection tube with a small amount of sterile saline added. Do not use lidocaine if a culture is going to be obtained from the biopsy specimen. Lidocaine inhibits various gram-positive and gram-negative bacteria, mycobacteria, and fungi.3
Blot tissue samples to remove blood and immediately place the samples into the biopsy jar to avoid the desiccation that can rapidly ensue. Ten percent buffered formalin is the most commonly used fixative and must be used with a minimum of 10 parts formalin to 1 part tissue to ensure adequate fixation. Place elliptical or large punch specimens dermis side down on a piece of wooden tongue depressor or cardboard to prevent curling artifact, allow them to adhere for 30 to 60 seconds, and then place them upside down in the biopsy jar.
Samples from different locations can be placed into different jars or may be marked with ink or suture to differentiate them if needed. This is important when removing multiple tumors or when multiple processes are suspected. Separate crusts may also be included in the biopsy jar wrapped in lens paper. Notify the pathologist that additional crusts have been submitted. If samples are mailed in the winter months, allow fixation for at least 12 hours before cold exposure.1 Contact the laboratory to determine if further fixation or additives are required.
Send samples to a veterinary pathologist specializing in dermatology or a veterinary dermatologist with expertise in dermatopathology. Ask your local dermatologist whom they would recommend in your area. Make sure to provide signalment, comprehensive history (including previous medications), examination findings, and differentials. Always include a lesion description and location on the animal. Good-quality pictures can be helpful as well.
A biopsy is merely part of a clinical workup and may not give a definitive diagnosis. However, even without a definitive diagnosis, some of the differential diagnoses may be eliminated or the results may establish a group of diseases to consider (eg, hormonal or allergic disorders). This information must then be combined with the patient’s history, clinical signs, and possibly additional testing to establish the diagnosis. If the description provided by the dermatopathologist does not correlate with the clinical appearance of the lesions, consider that the clinical interpretation has been incorrect or that the biopsy specimens are not representative of the disease.4 Some cases will require repeat biopsy procedures because diagnostic lesions may not be present at the initial biopsy.
- Miller WH, Griffin CE, Campbell KL. Diagnostic methods. In Muller & Kirk’s Small Animal Dermatology, 7th ed. St. Louis: Elsevier, 2013, pp 92-95.
- Henfrey JI, Thoday KL, Head KW. A comparison of three local anaesthetic techniques for skin biopsy in dogs. Vet Dermatol 1991; 2(1):21.
- Williams BJ, Hanke CW, Bartlett M. Antimicrobial effects of lidocaine, bicarbonate, and epinephrine. J Am Acad Dermatol 1997; 37(4):662.
- Shearer D. Dermatopathology. In Foster AP, Foil CS (eds): BSAVA Manual of Small Animal Dermatology, 2nd ed. Gloucester: BSAVA, 2003, pp 31-36.