Diagnostic Procedures
Biopsy of the nail unit
The nail is made up of many individual components. Each has a particular anatomical and physiologic function and characteristic. Therefore, appropriately, each of these components is affected by specific disease processes. Several specific indications for performing biopsy on the nail unit exist. Overall, the primary purpose of nail biopsy is to help the clinician establish or confirm a definitive diagnosis and, thus, initiate early effective treatment to prevent further nail destruction. Biopsy of the nail unit should be avoided if possible in patients with known peripheral vascular disease, collagen vascular disease, or diabetes mellitus and in those who are immunocompromised.
Biopsy of the nail unit may be performed for the following indications:
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To determine the pathogenicity of fungal organisms in recalcitrant fungal infection [41]
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To establish the presence of a psoriatic nail versus a mycotic nail or other underlying nail disease causing long-standing nail plate deformity (eg, onycholysis) [42]
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To identify and treat the cause of pain (eg, ingrown nails, tumors)
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To facilitate the early diagnosis of longitudinal pigmented streaks; malignant subungual and periungual tumors; and benign tumors, such as subungual exostosis of the nail unit [8]
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To differentiate an inflammatory etiology (eg, lichen planus) from an infectious etiology involving the nail unit
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To obtain diagnostic and prognostic information in presentations of isolated nail pathology when routine diagnostic procedures are unsuccessful [43]
To obtain an acceptable tissue specimen for pathologic evaluation and to ensure optimal cosmetic success when performing a biopsy of the nail apparatus, several issues, described below, should be considered. [13, 44]
Before proceeding with nail biopsy, the nail surgeon must determine the procedure that will be most beneficial to the patient, while offering minimal risks and complications.
The nail surgeon should perform a biopsy on the smallest amount of nail tissue necessary to make a diagnosis, leaving the healthy tissue undisturbed.
When possible, biopsy of the nail bed should be performed, and the anatomical area marking the nail matrix should be avoided.
Biopsy samples of the nail bed should be longitudinal, while samples of the nail matrix, when necessary, should be transverse or horizontal. Depending on the location of the pathology, longitudinal biopsy samples are best obtained at the lateral margin of the nail where the risk of scarring is reduced.
All incisions should be extended to the underlying bony phalanx. Excisions should be fusiform, approaching a width of less than 3 mm. After successful biopsy, if the area excised is more than 2 mm, it should be carefully undermined and closed with nylon sutures.
Anesthesia must be properly administered before performing any invasive procedure on the nail unit. After biopsy, the specimen should be placed in periodic acid-Schiff (PAS) or Gomori methenamine-silver stain. In onychomycosis, a few fungal elements may not be seen if the specimen is stained with only hematoxylin and eosin (H&E). [2]
Biopsy of the nail plate
Biopsy of the nail plate and the hyponychium is performed to help diagnose mycotic nail dystrophies, such as onychomycosis and other dermatoses affecting the nail plate. Biopsy of the nail plate usually involves removing a piece of the distal plate along with a portion of the underlying hyponychium by using scissors, a bone rongeur, or a scalpel. Alternatively, in cases where mycotic disease is suspected to extend beyond the distal nail plate, a 3- or 4-mm trephine punch may be used. [8] At the target site, the punch is applied with moderate pressure against the nail plate, passing through the nail bed until it contacts the periosteum. The punch is withdrawn, and the specimen is removed with fine scissors for histopathologic examination.
A technique that can be used to rule out onychomycosis is the noninvasive keratin biopsy. This biopsy technique has the added benefit of helping differentiate between hemosiderin versus melanic pigment in cases in which the nail has black or brown discoloration. [40]
The 2-punch technique is used when both the nail plate and the nail bed are involved in a specific disease. In this technique, the larger punch is used first to remove a circular defect in the nail plate. This punch is followed by a smaller punch that is advanced through the defect created in the nail plate to the nail bed underneath. [8, 14] When the nail plate is the primary target for sampling, a quick punch biopsy can be performed without anesthesia. Some surgeons prefer disposable punches because of their sharpness and ease of penetration. After biopsy, hemostasis is achieved by the direct application of pressure and the placement of Oxycel or oxidized cellulose at the biopsy site.
Biopsy of the nail bed
Biopsy of the nail bed may be performed to facilitate the accurate diagnosis of a tumor, to diagnose an unknown isolated lesion in the nail bed, or to diagnose a disease [8, 14] Multiple ways to perform biopsy of the nail bed exist.
A standard punch biopsy of the nail bed is one of the most commonly performed surgical procedures on the nail unit. [2] Before biopsy of the nail bed, radiography of the involved digit is indicated if extensive tissue distortion is present or if the type of pathology is questionable. [30]
When the nail bed is sampled, a partial avulsion of the distal half of the plate may be performed, or a complete avulsion may be performed to remove the entire nail plate, allowing direct visualization of the disease process involving the underlying nail bed. Avulsion is followed by 3-mm punch biopsy of the nail bed extending to the periosteum. Fine iris scissors are used to release the tissue specimen. The surgeon may choose to perform a longitudinal elliptical excision of the nail bed, advancing fully to the periosteum to remove a narrow wedge of tissue for sampling. Proximal avulsion of the nail plate is performed when a glomus tumor is suspected. In cases where nail avulsion is not performed, thinning of the nail plate may be required before obtaining tissue from the nail bed.
Alternatively, a decision might be made to leave the nail plate intact. The double punch technique is performed to obtain tissue from the nail bed. First, a larger 4- or 6-mm punch is applied to remove a circular defect in the nail plate. [8, 14, 45] A second smaller 3-mm punch is used to obtain a specimen of the nail bed through the nail plate defect created by the first larger punch. [8, 14, 45]
In situations where PAS staining is indicated to detect hyphal elements for diagnosing chronic onychomycosis from other nail dystrophies, a punch through the nail plate into the distal nail bed and the hyponychium is useful. [13]
If a standard punch biopsy of 3 mm or smaller is used, the surgical site does not need to be closed, and it is allowed to heal by secondary intention. Granulation tissue develops shortly after biopsy.
For longitudinal excisions larger than 2 mm, the surgical site should be undermined first, [14] especially if the nail bed epithelium is greatly bound down. This step is followed by the placement of 4-0 or 5-0 Dacron or nylon sutures. After biopsy, to achieve hemostasis, 35% aluminum chloride in 50% isopropyl alcohol (Monsel solution) is applied to the site. Oxycel or absorbable collagen may also be used as an aid to hemostasis. Excellent healing with minimal nail scarring and distortion is normal after biopsy of the nail bed. Occasionally, nail splitting at the sampling site may occur.
Biopsy of the nail matrix
The primary indications for biopsy of the nail matrix include (1) diagnosing tumors originating in the matrix; (2) determining the cause of full length nail plate malformations and nail dystrophies arising from a matrix abnormality; and (3) establishing a diagnosis of melanonychia striata versus malignancy (eg, melanoma) in presentations of longitudinal, dark pigmented streaks in the nail plate. [1, 13, 33, 46, 47] Although junctional nevi in the matrix most often account for the brown- or black-pigmented streaks found in the nail plate, melanoma involving the matrix is an important part of the differential diagnosis and must be excluded by matrix biopsy.
When biopsy of the nail matrix is performed, the normal curvature of the distal matrix or lunula must be maintained to avoid distortion and scarring of the nail plate. [1, 8, 13] Additionally, all biopsy samples of the nail matrix should be transverse rather than longitudinal, as samples taken from the nail bed.
Before biopsy of the nail matrix is performed, the extremity is cleansed with derma surgical soap, followed by a 1:1000 aqueous solution of Zephiran. A sterile surgical field must be maintained throughout the procedure.
In presentations with suspicious pigmented lesions involving the matrix, a mark should be placed on the PNF to identify where the pigmented lesion was first found.
Anesthetic is administered through a digital or local block, and a wide Penrose drain is used for hemostasis.
The nail plate may be completely avulsed, but complete avulsion is not always indicated, especially if a punch 3 mm or smaller is used. A partial avulsion of the proximal one third of the plate may be performed, leaving the distal two thirds of the nail plate intact (eg, when a focal abnormality is confined to the proximal one third of the matrix). Zaias recommends performing a complete nail avulsion to avoid the throbbing pain that develops from edema occurring in the traumatized area in partial avulsions. After avulsion, the matrix is fully exposed, and the lesion is identified.
At this point, biopsy of the matrix can be completed in 1 of 2 ways. All or part of the lesion may be removed with a 3-mm trephine punch biopsy or by a crescent-shaped (transverse) elliptical wedge excision, both extending down to the bony phalanx. The distal edge of the ellipse should be parallel to the curvilinear shape of the distal matrix. Maintaining the integrity of the distal lunula border minimizes the formation of a new dystrophic nail plate. [1, 8, 13, 14]
In proximal nail plate avulsions performed for matrix biopsy, after standard preparation, a Freer elevator is inserted beneath the PNF disrupting the association between the cuticle and the nail plate. A small incision is made at the junction of the PNF and the LNFs on both sides of the digit. The incision is extended posteriorly and approximately 5 mm proximally and laterally toward the DIP joint. The PNF and its free edges are reflected with skin hooks, allowing full exposure of the matrix. At this juncture, the matrix may be sampled by performing 3-mm trephine punch biopsy, incisional biopsy, or elliptical wedge excision. When complete exposure of the matrix is required, the nail plate may be partially or completely avulsed prior to biopsy.
A mosquito hemostat is used to free the proximal one third of the nail plate from its attachments at the proximal and lateral nail walls. The blunted hemostat is used to separate the proximal plate and the associated nail root from the underlying nail bed. Fine iris scissors are used to dissect the proximal one third of the nail plate from the distal two thirds, allowing the proximal nail plate to be released. Usually, bleeding is controlled with direct pressure. Some permanent scarring or defect is expected to form in the nail plate, especially when biopsy samples larger than 3 mm are obtained from the distal matrix. [48, 49] However, the defect is barely noticeable if the matrix is appropriately sampled.
Transverse biopsy of the nail matrix may change the thickness of the nail plate, producing a thinner plate only if the proximal matrix is undisturbed and no fissures are present. [8] Distal onycholysis and an abnormal curvature of the distal free edge of the nail plate may result if the convex border of the distal lunula is accidentally bisected.
When closing the surgical defect created in the matrix, several guidelines should be followed to prevent long-term nail disfigurement and dysfunction. After biopsy, the matrix is sensitive to tension. [1] Therefore, the defect in the matrix should first be carefully undermined and approximated to avoid tension during closure. [1, 8] The surgical site is closed with interrupted absorbable 4-0 or 5-0 Dexon sutures placed superficially. Sutures should not be buried deeply into the dermis because it is here that the matrix epithelium gives rise to the nail plate. At times, the surgical defect is allowed to heal by secondary intention. When sutures are appropriately threaded into the wound, they are seen emerging from the matrix epithelium into the newly formed plate as the nail plate grows distally. [8]
If the PNF is retracted, it is placed into its original position, and the incisions are closed with interrupted 5-0 nylon sutures. Xeroform gauze is usually inserted between the matrix and the ventral surface of the PNF to avoid the subsequent formation of adhesions. Pterygium results when the PNF and the open matrix adhere to each other and form a scar. The outcome is a new nail that is onycholytic. When compared with fusiform biopsies and excisions of the matrix, crescentic excisions are less favored because they may provide samples of inadequate width. [8] In conclusion, the likelihood of eventual nail distortion is decreased if the defect is properly sutured.
Haneke recommends a practical approach to the management of large pigmented lesions of the nail unit instead of total nail avulsion in all cases. He suggests performing a shave biopsy of the complete pigmented layer, removing the upper layers of the matrical epidermis. If the lesion is found to be malignant after a pathology evaluation, excision of the entire nail apparatus is required. Alternatively, if the lesion is found to be benign, the patient is spared the complication of significant nail dystrophy. [50]
En bloc biopsy of the nail complex
The lateral longitudinal en block biopsy of the nail is performed to sample the PNFs and the LNFs, the nail plate, the nail bed, the nail matrix, and the hyponychium. This procedure should be reserved for cases suggestive of longitudinal melanonychia, for dystrophy present throughout the entire length of the nail, and for determining the extent of involvement of the nail complex by malignancy (eg, to exclude localized vs invasive SCC or basal cell carcinoma of the nail unit). [23, 46, 48]
Lateral longitudinal biopsy of the nail complex is also performed for the purpose of surgically correcting a lesion in the lunula–nail bed area producing onycholysis and for ablation of the lateral matrix in the treatment of an ingrown nail. [1, 23, 51, 52] Standard preparation includes performing a digital block and soaking the extremity in a warm antiseptic solution, followed by scrubbing of the area beneath the distal edge of the nail plate.
If biopsy of the nail plate is not necessary, a complete avulsion is performed. If the surgeon desires to sample the nail plate, nail avulsion is performed on both sides of the area selected for biopsy. Partial hemiavulsion is performed with a mosquito hemostat or, preferably, a Freer elevator, the latter being less traumatizing to the nail. First, the PNF is separated from its attachments. Then, the nail plate is separated from the nail bed, the nail matrix, and the hyponychium.
En bloc excisional elliptical biopsy is performed by using a scalpel and curved iris scissors. The incision should be parallel to the lateral edge of the nail plate, and it must be advanced to the periosteum to ensure that a full-thickness sample of the matrix is included in the specimen. A rectangular block of tissue about 2-3 mm in diameter that contains the nail bed, the nail matrix, the PNF, and the hyponychium, with or without the nail plate, is removed. After biopsy, the edges of the lunula must be carefully approximated and sutured in a manner that recapitulates its original shape. A new LNF is formed by joining the remaining LNF to the lateral nail plate. All suturing should be directed away from the incision site to reduce tension and to provide better maneuverability; 3-0 or 4-0 nylon sutures should be used. The first suture is placed into the LNF, followed by the nail bed, and then the nail plate. The suture knot is tied on the nail plate away from the incision.
In some cases, the biopsy site is allowed to heal by secondary intention. After this procedure, the nail's width is permanently reduced, and the angle between the PNF and the LNF becomes more acute. Onycholysis rarely develops after lateral longitudinal nail biopsy. Removing the specimen from the lateral margin of the nail reduces the risk of functional impairment and nail dystrophy. Central longitudinal nail biopsies are associated with significant scarring, permanent fissuring, and poor cosmetic outcome. [32, 48, 49] If the only choice is to perform a midline biopsy, scarring is reduced if the PNF is reflected.
En bloc excision of the PNF
The PNF may be completely excised to assist in diagnosis and treatment of recalcitrant chronic paronychia, refractory myxoid cysts, collagen vascular disorders, and benign and malignant tumors. [13, 49]
Separation of the nail plate from the cuticle is accomplished with a dental spatula or Freer elevator. The elevator or spatula is advanced until it reaches the proximal nail groove. The PNF is excised with a scalpel by making a crescent-shaped excision 3 mm beyond the DIP joint, starting at the junction of the PNF and the LNF and ending at the proximal-lateral fold junction on the contralateral side. When biopsy of the PNF is performed, the matrix may be shielded from accidental incision by placing a Freer elevator between the PNF and the matrix. Healing occurs by secondary intention.
Biopsy of the PNF and the LNF
Incisional biopsy is the preferred surgical approach when sampling the PNF or the LNF for diagnostic purposes. [13] The most common indication for biopsy of the PNF is the presence of focal disease, such as chronic paronychia, focal tumor, myxoid cyst, collagen vascular disease, or dermatosis. [8, 23] Biopsy of the PNF may be completed in one of three ways.
One technique involves performing a 2-mm punch biopsy that reaches and samples the dorsal surface of the nail plate but leaves the distal margin of the PNF intact. When a larger amount of tissue is required for sampling, a transverse incisional biopsy of the PNF is performed. The incision extends to the periosteum and is parallel to the relaxed skin tension lines. This technique provides adequate tissue for histology and staining (eg, PAS, acid-fast, H&E). It also permits easy primary closure without undermining the wound, while maintaining the anatomy of the nail fold. Areas that should be avoided during the incision are the distal free margin of the nail to prevent notched scarring and the extensor digitorum tendon near the DIP joint. [13] Suturing is performed by using 4-0 or 5-0 Prolene sutures. Sutures are removed in 1 week.
When surface biopsy is indicated, a razor blade is used (shave biopsy). The blade is first split into two parts, and the half blade is used in a flat plane to obtain a sample of the PNF tissue at the desired depth. [13, 49] Hemostasis at the surgical site is achieved with Monsel solution and direct pressure. The biopsy site usually heals by secondary intention with negligible scarring. Biopsy of the LNF is most commonly performed in presentations of hyperkeratotic plaques previously treated as warts or fungal infection. [13]
Excisional biopsy is performed to exclude Bowen disease or SCC. This procedure is also used to excise the hypertrophic tissue occurring in onychocryptosis. Except for a longitudinally directed fusiform excision, biopsy of the LNF follows the same surgical approach as that used on the PNF. Frequently, the lateral sulcus and the ventral nail plate may harbor pathology requiring paring back of the nail plate to allow full visualization of all areas.
All biopsies of the nail bed should extend to the periosteum. When suturing, the suture is first passed through the LNF skin, then across the nail bed, and finally through the nail plate. In all biopsies, a digital block is performed, and a tourniquet is used for hemostasis.
Imaging Studies
Ultra‒high-frequency ultrasound
More recently, the development of ultra‒high-frequency ultrasound introduces new diagnostic, noninvasive imaging modalities. The main applications include assessment of skin layers and hair follicles, as well as the nail unit. [53] This has the potential to delineate anatomy preoperatively, differentiate solid and cystic lesions, [54] and aid in therapeutic decisions when clinical examination is equivocal. However, there is still a need for comprehensive validation studies and assessment of long-term outcomes based on the ultrasound examinations.
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Longitudinal section of a digit showing the dorsal nail apparatus (top). Component part of the nail apparatus (bottom).