Mohs Micrographic Surgery: Specimen-Releasing Techniques, Marking, and Orientation

Function

The initial stage of the Mohs procedure involves identifying the biopsy site, which can be challenging due to the small size of many tumors at the time of biopsy, the presence of partially or fully healed scars, and the surrounding actinic damage. Research indicates that 16.6% of patients and 5.9% of physicians misidentify biopsy sites during examinations when preoperative photographs are not used.[1] Once the surgical site is confirmed, the area is cleaned with an alcohol swab, and the visible tumor is outlined with surgical ink, leaving a 1 to 2 mm margin. After marking, the surgical area is disinfected with an antiseptic, ensuring that the ink markings remain intact.[6]

Before administering a local anesthetic, it is essential to identify and mark relevant anatomical boundaries or features that may assist in reconstruction planning. The swelling from the injected anesthesia can make it more challenging to locate landmarks (eg, re-approximating the vermillion lip).[2] Next, the initial stage of tumor removal is performed using a scalpel blade positioned at a 45º angle to the skin. This angle ensures proper alignment of the tissue specimen's peripheral edges during pathological preparation, which is a crucial aspect of MMS.[6]

Before removing the tissue from the patient, orientation marks should be placed on the patient's skin around the excised specimen, ensuring alignment with the marks on the specimen itself. This is typically done by methodically making small nicks or scores in the tissue with a scalpel. This process helps the surgeon correlate histological findings with the exact location on the patient's skin.[6] Additional methods of in vivo orientation involve using sutures, staples, gentian violet, or methylene blue.[7][8] Advocates of tissue scoring highlight its simplicity, precision, speed, cost-effectiveness, resistance to smudging, and minimal risk of causing tattoos. Supporters of gentian violet prefer its complete lack of risk for excessive scarring in adjacent skin. However, many surgeons continue to use the techniques they learned during training.[9]

After positioning the correct orientation markings, the next step is to fully excise the tumor with a narrow margin of normal-appearing skin, ensuring proper orientation for creating an accurate Mohs map. Hemostasis is then achieved using electrosurgery, electrocautery, chemical cautery, or direct pressure.[1] The excised specimen is then measured and drawn to scale on the Mohs map, with particular attention to the orientation markers. The excised tissue and the Mohs map are then transported to the Mohs laboratory for further processing in preparation for freezing. To minimize potential errors and facilitate tissue transport, the Mohs laboratory should be located as close to the surgical suite as possible.[10] 

After the tissue arrives in the laboratory, it may be processed as a single section or divided into multiple sections, depending on the size of the pathology slide relative to the excised tissue.[7] Various techniques have been outlined to ensure a complete margin and epidermal edge in frozen sections, including beveled excision, relaxing incisions (ex vivo or in vivo), mechanical flattening of tissue, and tissue division. The tissue is then carefully scored around the edges to ensure that both the epidermis and deep margin are flat and aligned on the same plane. This process transforms the 3-dimensional (3D) tissue specimen into a 2D pathology slide.[2][10] 

Subsequently, the technician or surgeon then inks the tissue to ensure proper orientation during the microscopic examination of each section. The ink colors and orientation are annotated on the Mohs map for future reference, ensuring they align with the tissue accurately, allowing the surgeon to identify any remaining positive margins.[10] Effective inking patterns are essential, such as using different colors to mark 2 adjacent non-epidermal edges and color-coding the positions at the 12-, 3-, 6-, and 9-o'clock positions.

Consistency in inking patterns is essential, as repeating the same techniques reduces errors and minimizes confusion between the histotechnician and surgeon. Once the specimen is properly flattened and oriented with hash marks and ink, it can be transferred to the histotechnician for sectioning and staining, typically using a nonstick pad, filter paper in a petri dish, glass slide, or gauze pad.[1] In general, adopting a consistent approach for releasing, marking, and orienting specimens is considered best practice, with clear communication of these methods to the entire healthcare team.

Issues of Concern

Wrong-Site Surgeries

Wrong-site surgeries pose a significant risk to patients' well-being and can lead to legal liabilities. Capturing images of the biopsy site, along with surrounding anatomical markers, is critical for accurate tumor localization. Skin cancers often blend with normal tissue, making precise identification challenging. Mistakes in identifying the biopsy site can result in unnecessary tissue removal, patient distress, and potential legal complications.[2] As such, photographic documentation is essential for effective communication among the healthcare team and for maintaining accurate medical records. These images act as a visual reference that can be easily shared among healthcare providers involved in the patient's care, ensuring consistency and clarity.

All Mohs surgeons should utilize biopsy site photographs, anatomical landmarks, and diagrams to guide their procedure and reduce the risk of misidentifying the biopsy site.[1] Many individuals now have personal cell phones with photo capabilities, making it helpful for patients to take photos of their biopsy sites and input them into their medical records. In cases where outside referrals to Mohs surgeons are needed, the anatomical location of the biopsy site, along with specific measurements from surrounding landmarks, should be documented, especially if photographs are not readily available or accessible. 

Hash Mark Placement

Accurate tissue mapping and orientation are essential for achieving clear margins in MMS. Hash marks are applied to both excised tissue and adjacent tissue to aid in orientation. Incorrect orientation can complicate the identification of positive margins, requiring additional tissue for reevaluation and potentially resulting in a larger defect. It is crucial to place hash marks clearly and avoid positioning them exactly 180 degrees apart, as this could lead to inaccurate tissue orientation if a processing error occurs. While practices vary, many surgeons use a single hash mark at the 12-o'clock position or 3 hash marks at the 12-, 3-, and 6-o'clock positions for initial stages. This approach, or other forms of asymmetric hash mark placement, allows for proper tissue reorientation if a handling error arises during tissue processing.[2] Surgeons often adjust this approach as needed for subsequent incomplete circumferential stages or specific anatomical considerations. Once again, precise marking of the orientation on the Mohs map is crucial to prevent confusion.

Tissue Debulking

Although many tumors do not require debulking before processing, larger tumors may require in situ (on the patient) or ex vivo (off the patient) debulking to facilitate proper flattening of the deep and peripheral margins for sectioning.[11] Debulking large exophytic tumors can make the staging process easier for the surgeon and the tissue processing more manageable for the histotechnician. Debulking a Mohs tissue specimen from the bottom is not advisable, as this can compromise the accuracy of margin evaluation. The bottom of the specimen contains the critical deep margin, which must be thoroughly examined for residual tumor cells. Removing tissue from this area may distort the natural anatomy and potentially eliminate part of the margin that needs to be assessed, increasing the risk of leaving behind cancerous cells. Therefore, preserving the integrity of the bottom margin is vital for a comprehensive and accurate evaluation. Tumor debulking is commonly performed with a scalpel or curette. Surgeons often recommend cleaning or changing instruments when taking the initial stage to prevent false-positive "floaters" from contaminating the processed tissue. 

Releasing Techniques

One of the fundamental principles of MMS is converting a 3D tissue specimen into a 2D slide, enabling assessment of both peripheral and deep margins on the same plane.[2] When a stage is taken without prior debulking, the tissue is carefully prepared to ensure accurate analysis by making shallow, parallel cuts to the surface. This technique relaxes the tissue centrally, allowing the edges to lie flat on the slide alongside the deep margin.

If the tumor is too large, dense, or fibrous for a single slide, it is typically divided into smaller sections for proper processing and placement on glass slides.[2][11] This division of tissue increases the time and cost required for processing and reading the slides.[12] The surgeon and histotechnician must employ all available techniques to ensure thorough assessment of both the peripheral and deep margins, while avoiding the loss of any part of the true margin or compromising further tissue processing.

Mohs Map Markings and Orientation

Accurate Mohs mapping is essential for the safety and success of MMS. Errors in mapping, such as incorrect orientation, sectioning, inking, miscommunication with the histotechnician, or specimen mislabeling, can result in inaccurate outcomes. As the number of cases and stages increases, so does the risk of mapping errors, highlighting the importance of strict quality control measures in MMS. Please see StatPearls' companion resource, "Mohs Micrographic Surgery Mapping Techniques," for more information.

The methods of tissue mapping and processing for MMS may vary, but ensuring accuracy and consistency at each stage is crucial for achieving reliable and repeatable cure rates for tumors. Effective cooperation and communication among everyone involved in handling, mapping, processing, and reading tissues are essential to maintain maximum control and ensure the efficiency and integrity of MMS.[12]

Errors in Tissue Processing

One of the key advantages of MMS is the ability to perform real-time histological evaluation of the entire surgical margin, ensuring the detection of any remaining cancer cells.[13] Accurate and efficient frozen section analysis is essential for reliable tumor control, making meticulous evaluation of surgical margins crucial.[14][15] Common sources of error in MMS include specimen preparation, slide interpretation, map marking, tissue mounting, and processing. The most significant risk factor for tumor recurrence after MMS is an unexcised tumor resulting from a surgeon's interpretation error.

Additionally, tissue dropout on histological slides due to processing is the second-highest risk factor for tumor recurrence.[3] For instance, if the slide showing the true margin is missing any part of the epidermis or deep margin, the surgeon should ask the histotechnician to recut the sample until the complete margin is visible. If the entire peripheral or deep margin cannot be observed on the slide, the surgeon cannot confidently confirm that all tumor cells have been removed. These factors highlight the importance of having a well-trained and skilled histotechnician to process tissue specimens.

Clinical Significance

Accurate orientation, consistent marking, and effective specimen-releasing techniques are critical for the success of MMS. They enable precise tumor localization, reliable mapping, and thorough histopathological analysis. These meticulous practices reinforce MMS as the gold standard for treating most skin cancers by preserving healthy tissue, ensuring clear margins, and minimizing the risk of recurrence. While continuous refinement of these methods is encouraged, Mohs surgeons consistently achieve optimal outcomes by maintaining a methodical and focused approach to these fundamental techniques.

Enhancing Healthcare Team Outcomes

MMS is a highly effective technique for treating skin cancer on the head, neck, genitalia, hands, and feet.[16] The methods for releasing, marking, and orienting specimens in MMS are crucial for ensuring the accuracy and efficiency of the procedure. Before surgery, capturing photographs and documenting the patient's condition can visually demonstrate any preexisting asymmetries, scars, muscle weakness, or other anatomical irregularities the patient may be unaware of. This documentation helps prevent the misattribution of these conditions to the surgical intervention.[2]

Marking specimens with colored dyes or other indicators is essential for maintaining the orientation of tissue sections. This practice ensures the surgical team can accurately map tumor margins and identify areas with remaining tumor cells. Physicians and histotechnicians should regularly evaluate their practices for efficiency, safety, and potential areas for improvement to minimize errors in the multistep MMS process. The medical community must prioritize transparency, accountability, and continuous quality assessment to reduce human error.[17]

Proper specimen orientation is equally crucial, as it provides a spatial reference for the surgeon to accurately assess the margins. Proper orientation ensures that the microscopic examination of the tissue accurately reflects its in vivo position. This enables the surgical team to precisely locate any residual tumor, which is crucial for achieving clear margins and reducing the risk of recurrence. Additionally, it minimizes the need for further surgical interventions, improving patient outcomes and lowering healthcare costs. 

A detailed Mohs map incorporating anatomical landmarks and skin notches is essential for accurate orientation, enhancing communication among the surgical team, and providing clear documentation for future reference. This mapping ensures a shared understanding of the tumor's location, the extent of tissue removal, and any areas requiring further attention. Additionally, it serves as a valuable record for follow-up care, facilitating the accurate assessment and management of recurrences or additional treatments. Please see StatPearls' companion resource, "Mohs Micrographic Surgery Mapping Techniques," for more information.

Releasing, marking, and orienting techniques are crucial to the success of MMS. They improve coordination and communication within the healthcare team, ensuring that each member has a precise understanding of the tumor's margins and the location of residual cancer cells. By enhancing the accuracy of tissue removal and minimizing the loss of healthy tissue, these techniques contribute to better surgical outcomes, higher cure rates, and increased patient satisfaction.