Accurate diagnosis of oral blistering diseases is predicated on obtaining a satisfactory biopsy for both hematoxylin and eosin (H&E) and direct immunofluorescence (DIF) evaluation. To be diagnostic, the biopsy specimens must contain intact epithelium over the underlying connective tissue. However, in the oral cavity these lesions are often fragile and separation between the epithelium and connective tissue is common, rendering the specimen non-diagnostic. A biopsy should be performed by a dentist or dental specialist experienced in performing biopsies of vesiculobullous lesions.
Here are some general guidelines:
- Do not sample the bed of an ulcer because there is no epithelium there, resulting in a non-specific diagnosis.
- As a general rule, biopsies must contain intact epithelium and should be taken from perilesional (within 1 cm) or normal appearing tissue rather than the directly ulcerated tissue because an ulcer, by definition, is missing the epithelium. Without the epithelium, it is not possible to detect destruction between epithelial cells (as in PV) or between epithelial cells and the underlying connective tissue (as in MMP).
- During the biopsy procedure, avoid any action that may cause separation of the epithelium from underlying connective tissue, e.g. rubbing the area with gauze to remove saliva or blood.
- Two specimens must be taken with samples submitted for BOTH routine hematoxylin and eosin (H&E) stain (storing specimen in 10% formalin) AND for direct immunofluorescence testing (MUST submit sample in Michel’s transport medium and send to pathology laboratory as quickly as possible to preserve antibody detection).
- When performing DIF biopsies, have Michel’s solution on hand. If planning to perform DIF biopsies, order and store in advance the Michel’s transport medium. Identify IN ADVANCE a commercial laboratory equipped to perform DIF studies.
See these instructions from Dr. David Sirois, DDM, PhD, for more information about H&E and DIF biopsies, as well as IIF serum studies, to evaluate PV/MMP.
The below list includes 11 guidelines provided by Dr. Terry Rees, DDS, MSD, Professor of Periodontics, Director of Stomatology Center, Texas A&M Baylor College of Dentistry.
Many authorities have addressed this issue and the following general guidelines may apply:
- If lesions are located in several mucosal sites including the gingiva, it is usually preferable to avoid a gingival biopsy. This is true because gingiva is thin and tends to be fragile. Most authorities report a relatively low diagnostic success rate with gingival biopsies.1-4
- If a gingival biopsy is required it should be taken apical to the marginal gingiva to avoid the inflammation found in gingival crevices which may mask the autoimmune disease histologic features.
- Avoid direct infiltration of local anesthetic into the biopsy site to prevent hemorrhage or artificial separation of epithelium from the connective tissue.
- The cutting force should be directed internally only, to avoid lateral stresses that may disrupt the epithelium/connective tissue interface.4
- As a general rule, biopsies should be taken from perilesional (within 1 cm) or normal appearing tissue rather than directly from an erosive lesion since the lesion epithelium has potentially been lost.5
- In cases of extreme erosive lesions caused by suspected autoimmune diseases it may be preferable to take the biopsy from distant normal tissue such as buccal mucosa and submit it only for DIF analysis.6
- When possible, two or more biopsies should be obtained to submit for H&E and DIF analysis. This avoids having to split a single biopsy into two components since this may disrupt the epithelial/connective tissue interface. On occasion, only a single biopsy specimen can be obtained so it must be split for analysis. In this event be certain that the direction of the separation cut is only internal without applying any lateral force.
- Early oral manifestations of autoimmune diseases may be confined to the gingiva (desquamative gingivitis). In this event, gingival biopsies are required and special surgical techniques may improve the likelihood of obtaining diagnostic specimens (Punch biopsy, stab and roll technique).
- Several authorities have recommended a punch biopsy for intraoral lesions. However, this technique, as recommended includes the application of a rotational and internal force, which may create lateral stresses on the tissue and dislodge the epithelium. To date there are no studies reporting on the success of this technique when obtaining gingival biopsies. Limiting the cutting direction of force to only an internal direction appears likely to insure more predictable success.
- Recently a “stab and roll” biopsy technique has been described for gingival biopsies, which limits the cutting force to an internal direction only. In this technique a #15 scalpel blade is used. The point of the scalpel is gently applied until the underlying bone is reached. Then the blade is rolled from its tip along the entire cutting edge resulting in application of only internal forces. In larger specimens the scalpel tip is reapplied into the incision line and the procedure is repeated. Using this technique it was possible to obtain biopsy specimens with intact epithelial surfaces in 51 of 52 biopsies (98.1%). Perhaps most importantly, intact epithelium was retained in 26 of 27 lesion or perilesional biopsies.4
- It is essential that biopsy tissues be appropriately transported to the laboratory. For H&E evaluation formalin is the required medium but for DIF a special transport solution (Michel’s transport medium) is used. The DIF specimen should only be sent to a laboratory that performs direct immunofluorescence studies.
- Siegel Ma. Intraoral biopsy techniques for direct immunofluorescence studies. Oral Surg Oral Med Oral Pathol 1991; 72:681-684.
- Daniels TE, Quadra-White C. Direct immunofluorescence in oral mucosal disease: A diagnostic analysis of 130 cases. Oral Surg Oral Med Oral Pathol 1981; 51:38-47.
- Casiglia J, Woo SB, Ahmed AR. Oral involvement in autoimmune blistering diseases. Clin Dermatol 2001; 19:737-747.
- Endo H, Rees TD, Allen EP, Kuyama K, Aoki S, Yamamoto H, Ito T. A stab-and-roll biopsy technique to maintain gingival epithelium for desquamative gingivitis. J Periodontol 2014; 85:802-809.
- Sano SM, Quarracino MC, Aquas SC et al. Sensitivity of direct immunofluorescence in oral diseases. Study of 125 cases. Med Oral Pathol Oral Cir Bucal 2008; 13:E287-E291.
- Mutasim DF, Adams BB. Immunofluorescence in dermatology. J Am Acad Dermatol 2001; 45:803-822.
- Suresh L, Neiders ME. Definitive or differential diagnosis of desquamative gingivitis through direct immunofluorescence studies. J Periodontal 2012; 83: 1270-1275.
- Eisen D. The oral mucosal punch biopsy. A report of 140 cases. Arch Dermatol 1992; 128:815-817.
- Seoane J, Varela-Centelles PI, Limeres-Posse J, Seoane-Romero JM. A punch technique for gingival incisional biopsy. Laryngoscope 2013; 123:398.