Hritish loiirnat of Dfrnuilnlocjii 119921 126. ^7-59.
A simple device for optimal tissue preparation for Mohs micrographic surgery R.|.MOTLEY AND P.J.A.HOLT Department of Dermutohnfii. Vmwrsiln Hospital of Wales. Hcaih Park. Ciiniiff Cl'4 4XW. U.K. Accepted for p u b l i c a t i o n 2 9 A u g u s t i 9 9 l
Summary
A novel device is described which allows the rapid preparation of perfectly orientated tissue specimens for Mohs micrographic surgery.
Mohs micrographic surgery is the most effective means for ensuring complete removal of cutaneous tumours.' Its principle is to examine microscopically the entire peripheral margin of an excised tissue specimen to precisely localize the site of any residual tumour and enable its subsequent removal. Tumour diagnosis is established by histological examination of formalin-fixed tissue prior to the procedure, firstly because the quality of frozen sections is inferior for diagnosis and secondly because there may be no visible (i.e. residual) tumour in the niicrographic sections. The technique is mainly employed for basal cell carcinomas although theoretically it can be extended to almost any neoplasm within soft tissue where local recurrence is a major concern." ^ Slide interpretation is generally performed by the micrographic surgeon whose experience in the technique within a limited range oftumour systems provides a high degree of diagnostic accuracy equal to that of the pathologist." "* Although Mohs originally used in-vivo tixation of the skin, his technique was adapted for fresh-frozen tissue sections, and these are now used.'' ^ In order to display the entire periphery of the tissue specimen it is necessary to manipulate its lateral and deep margins into a single flat plane before freezing. The tissue must then be mounted with this plane parallel to the cutting stroke of the microtome knife to avoid excessive tissue loss in 'facing' the block and to reduce the number of sections necessary to evaluate the complete margin. Several methods of tissue preparation have been previously reported.*^"" We have developed a new device which allows the preparation of ct)nsistently well-orientated specimens for micrographic surgery.
Correspondence: i>r K.|.Mi)tiey.
Methods The apparatus
The equipment consists of a central cylinder of high density plastic into which the cryostat chuck can be mounted. A small pipe in the centre of the cylinder delivers liquid nitrogen from a cryospray ("Cry-Ac' Brymill Corporation, U.S.A.) titted with a foot-pedal operated valve. The cylinder is surrounded by a closefitting metal sleeve IFIg. 1). Method
The bulk of the tumour is removed by curettagc and a layer of tissue, encompassing the entire wound bed. is excised from around and tmderneath the wound. The Incision through the skin is made at an angle of 30-45° thus creating a gradual transition between the lateral and deep margins. This disc of tissue is divided into quadrants and the divisions marked with tissue dyes for
Figure I. The appar.ilus is shown with Ih*? specimen ready for frc on a glass microscope slide. Note the two concentric cylinders and the central pipe to deliver liquid nitrogen. The specimen is frozen with its edges held (lat against the slide.
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R.J.MOTLEY AND P.J.A.HOLT
Figure 2. A cross-sectional view of the apparatus. A cryostat chuck is placed in the centre of the apparatus and OCT compound applied to its surface. The external sleeve of the apparatus is then elevated and the slide, inverted with the frozen specimen underneath, placed on top. tJsing the sleeve as a support the specimen is lowered inlu tbe OCT and then frozen onto the cryostal chuck. Vent holes in the bottom of the apparatus allow nitrogen to escape.
later identification. The tissue pieces are tben prepared for examination using the apparatus described. Tbe specimen is placed witb its deep surgical margin against the face of a glass microscope slide laid on top of the apparatus (Fig. 1). Tbe edges of the specimen, particularly the epidermal edge, are held gently against the glass slide wbilst liquid nitrogen is sprayed on its undersurface. freezing tbe tissue. This takes only a few seconds, although specimens which are difficult to flatten may be frozen more slowly if desired. Tbe tissue can be examined after freezing to ensure its entire surface is fiat before embedding in OCT. The cryostat chuck is now placed in the centre of the apparatus and OCT compound liberally applied to it. The external sleeve is elevated and the slide, now inverted with the specimen underneath, placed across it. The sleeve is tben lowered until tbe specimen sits within the OCT compound and liquid nitrogen is sprayed on the undersurface of the cryostat chuck, freezing the OCT and the specimen in situ (Fig. 2). Finally, the glass slide is detached by gently warming its surface, leaving the specimen perfectly flat and embedded on the cryostat chuck ready for sectioning.
Discussion This technique offers several advantages over previously described metbods of preparing fresb tissue for frozensection micrograpbic analysis. Specimen preparation is not dependent upon the use of the cryostat. eliminating the need to work within tbe confines of the cryostat and preventing undesirable warming of tbe chamber. Freezing with liquid nitrogen is considerably faster than using
traditional aerosol freezing sprays or the cryostatmcnmted 'heat sink". Botb fiattening of tbe specimen and its freezing are entirely controllable by tbe operator, and the quality of tissue preparation can be visually assessed before embedding the tissue. The use of a close-fitting external sleeve to support tbe inverted slide ensures tbat the fiattened surface of the frozen specimen Is embedded parallel to the face of the cryostat chuck. There is complete freedom to rotate tbe specimen relative to the cryostat chuck prior to embedding, thus ensuring optimal orientation with the epidermal edge perpendicular to tbe knife wben tbe preparation is subsequently mounted in tbe microtome. (This is of importance only when square non-rotatable chucks are used.) OCT compound adberes to tbe face of tbe cbuck more consistently when applied at room temperature and subsequently frozen tban wben applied to a pre-cooled chuck—a common method witb some techniques. This reduces the chance of the specimen being dislodged during sectioning. The OCT is sandwiched between tbe face of the chuck and the glass slide during freezing, eliminating irregularities in the final surface of tbe preparation and facilitating smooth passage of the microtome knife through the block. Because the specimen is simply lowered into OCT and then frozen, without any need for compression, it is not necessary for the surgeon to cut the traditional thin slices of tissue used in micrographic surgery: irregular lumps of tissue can be prepared for micrographic examination just as easily. In conclusion, our technique is quickly learnt, simple to perform and allows rapid production of perfectly orientated specimens for Mohs micrographic surgery. This technique is now routinely used in Cardiff.
Acknowledgment We are grateful to l.L. and A.D.Motley for their assistance in constructing this device.
References 1 Swanson N. Mohs surgery: technique, indications, applications and the future. Arch Dermatol 198i: 119: 7h\-7i. 2 Headington JT. A dermatopatbologist looks at Mohs micrographic surgery. Arch DermatoJ 1990; 126: 9SO-1. I Bennett RG. Mohs surgery. New concepts and applications. Dermatolofjic Clinics 1 9X7: 5: 409-28. 4 Zitelli )A. Moy KL. Abell E. The reliabilily of frozen sections in the evaluation of surgical margins for melanoma. / Am Acad Dermatol 1991: 24: 102-f>. 5 Grabski Wj, Salasche SI. McCollough ML cl al. Interpretation of Mohs micrographic frozen sections: A peer review comparison study. / Am Acad Dermato! 1989: 20: 670-4. ft Tromovitch TA. Stegman S]. Microscopic-controlled excision of
OPTIMAL TISSUE PREPARATION IN MOHS SURGERY
7 S 9 11)
cutaneous tumours. Chemosurgery, fresh tissue technique. Cancer 1978; 41: 653-8. Cottel Wl, Proper S. Moh's surgery, fresh tissue technique. Our tecbnique with a review. I Dermtito! Simi Onco! 1982; 8: 576-87. Picoto AM. Picoto A. Technical procedures for Mohs fresh tissue surgery. I Dermatol Surti Oncol 1986; 12: 134-8. Hanke CW, Ix^e MW. Cryostat use and tissue processing in Mobs micrographic surgery. / Dermatol Siirei Oncol 1989: 15: 29-32. Honda NS. Friedman DH. A simple method of tissue embedding for Mobs microgrtiphic surgery. / Dermutol Surg Oncol 1989: 1 5: 502-4.
59
1 1 Carter VH. A new method for preparing tissue blocks for cryostat sectioning. / Dfmr.il.i; Surii Oncol 1985: I I : 687-9. 12 Gormley DE. F.valuation of a method for controlled tissue embedding for bistologic evaluation of tumor margins. Am / Dirmalo/wt/ioJ 1987: 9: 308-15. 13 Lesbin B. Cook SR. Frye DW. Cryomold: a device for tissue embedding in Mobs micrographic surgery, j I'h-rmatol Surfi Oncol 1991; 17: 234-6.
A simple device for optimal tissue preparation for Mohs micrographic surgery R.|.MOTLEY AND P.J.A.HOLT Department of Dermutohnfii. Vmwrsiln Hospital of Wales. Hcaih Park. Ciiniiff Cl'4 4XW. U.K. Accepted for p u b l i c a t i o n 2 9 A u g u s t i 9 9 l
Summary
A novel device is described which allows the rapid preparation of perfectly orientated tissue specimens for Mohs micrographic surgery.
Mohs micrographic surgery is the most effective means for ensuring complete removal of cutaneous tumours.' Its principle is to examine microscopically the entire peripheral margin of an excised tissue specimen to precisely localize the site of any residual tumour and enable its subsequent removal. Tumour diagnosis is established by histological examination of formalin-fixed tissue prior to the procedure, firstly because the quality of frozen sections is inferior for diagnosis and secondly because there may be no visible (i.e. residual) tumour in the niicrographic sections. The technique is mainly employed for basal cell carcinomas although theoretically it can be extended to almost any neoplasm within soft tissue where local recurrence is a major concern." ^ Slide interpretation is generally performed by the micrographic surgeon whose experience in the technique within a limited range oftumour systems provides a high degree of diagnostic accuracy equal to that of the pathologist." "* Although Mohs originally used in-vivo tixation of the skin, his technique was adapted for fresh-frozen tissue sections, and these are now used.'' ^ In order to display the entire periphery of the tissue specimen it is necessary to manipulate its lateral and deep margins into a single flat plane before freezing. The tissue must then be mounted with this plane parallel to the cutting stroke of the microtome knife to avoid excessive tissue loss in 'facing' the block and to reduce the number of sections necessary to evaluate the complete margin. Several methods of tissue preparation have been previously reported.*^"" We have developed a new device which allows the preparation of ct)nsistently well-orientated specimens for micrographic surgery.
Correspondence: i>r K.|.Mi)tiey.
Methods The apparatus
The equipment consists of a central cylinder of high density plastic into which the cryostat chuck can be mounted. A small pipe in the centre of the cylinder delivers liquid nitrogen from a cryospray ("Cry-Ac' Brymill Corporation, U.S.A.) titted with a foot-pedal operated valve. The cylinder is surrounded by a closefitting metal sleeve IFIg. 1). Method
The bulk of the tumour is removed by curettagc and a layer of tissue, encompassing the entire wound bed. is excised from around and tmderneath the wound. The Incision through the skin is made at an angle of 30-45° thus creating a gradual transition between the lateral and deep margins. This disc of tissue is divided into quadrants and the divisions marked with tissue dyes for
Figure I. The appar.ilus is shown with Ih*? specimen ready for frc on a glass microscope slide. Note the two concentric cylinders and the central pipe to deliver liquid nitrogen. The specimen is frozen with its edges held (lat against the slide.
57
58
R.J.MOTLEY AND P.J.A.HOLT
Figure 2. A cross-sectional view of the apparatus. A cryostat chuck is placed in the centre of the apparatus and OCT compound applied to its surface. The external sleeve of the apparatus is then elevated and the slide, inverted with the frozen specimen underneath, placed on top. tJsing the sleeve as a support the specimen is lowered inlu tbe OCT and then frozen onto the cryostal chuck. Vent holes in the bottom of the apparatus allow nitrogen to escape.
later identification. The tissue pieces are tben prepared for examination using the apparatus described. Tbe specimen is placed witb its deep surgical margin against the face of a glass microscope slide laid on top of the apparatus (Fig. 1). Tbe edges of the specimen, particularly the epidermal edge, are held gently against the glass slide wbilst liquid nitrogen is sprayed on its undersurface. freezing tbe tissue. This takes only a few seconds, although specimens which are difficult to flatten may be frozen more slowly if desired. Tbe tissue can be examined after freezing to ensure its entire surface is fiat before embedding in OCT. The cryostat chuck is now placed in the centre of the apparatus and OCT compound liberally applied to it. The external sleeve is elevated and the slide, now inverted with the specimen underneath, placed across it. The sleeve is tben lowered until tbe specimen sits within the OCT compound and liquid nitrogen is sprayed on the undersurface of the cryostat chuck, freezing the OCT and the specimen in situ (Fig. 2). Finally, the glass slide is detached by gently warming its surface, leaving the specimen perfectly flat and embedded on the cryostat chuck ready for sectioning.
Discussion This technique offers several advantages over previously described metbods of preparing fresb tissue for frozensection micrograpbic analysis. Specimen preparation is not dependent upon the use of the cryostat. eliminating the need to work within tbe confines of the cryostat and preventing undesirable warming of tbe chamber. Freezing with liquid nitrogen is considerably faster than using
traditional aerosol freezing sprays or the cryostatmcnmted 'heat sink". Botb fiattening of tbe specimen and its freezing are entirely controllable by tbe operator, and the quality of tissue preparation can be visually assessed before embedding the tissue. The use of a close-fitting external sleeve to support tbe inverted slide ensures tbat the fiattened surface of the frozen specimen Is embedded parallel to the face of the cryostat chuck. There is complete freedom to rotate tbe specimen relative to the cryostat chuck prior to embedding, thus ensuring optimal orientation with the epidermal edge perpendicular to tbe knife wben tbe preparation is subsequently mounted in tbe microtome. (This is of importance only when square non-rotatable chucks are used.) OCT compound adberes to tbe face of tbe cbuck more consistently when applied at room temperature and subsequently frozen tban wben applied to a pre-cooled chuck—a common method witb some techniques. This reduces the chance of the specimen being dislodged during sectioning. The OCT is sandwiched between tbe face of the chuck and the glass slide during freezing, eliminating irregularities in the final surface of tbe preparation and facilitating smooth passage of the microtome knife through the block. Because the specimen is simply lowered into OCT and then frozen, without any need for compression, it is not necessary for the surgeon to cut the traditional thin slices of tissue used in micrographic surgery: irregular lumps of tissue can be prepared for micrographic examination just as easily. In conclusion, our technique is quickly learnt, simple to perform and allows rapid production of perfectly orientated specimens for Mohs micrographic surgery. This technique is now routinely used in Cardiff.
Acknowledgment We are grateful to l.L. and A.D.Motley for their assistance in constructing this device.
References 1 Swanson N. Mohs surgery: technique, indications, applications and the future. Arch Dermatol 198i: 119: 7h\-7i. 2 Headington JT. A dermatopatbologist looks at Mohs micrographic surgery. Arch DermatoJ 1990; 126: 9SO-1. I Bennett RG. Mohs surgery. New concepts and applications. Dermatolofjic Clinics 1 9X7: 5: 409-28. 4 Zitelli )A. Moy KL. Abell E. The reliabilily of frozen sections in the evaluation of surgical margins for melanoma. / Am Acad Dermatol 1991: 24: 102-f>. 5 Grabski Wj, Salasche SI. McCollough ML cl al. Interpretation of Mohs micrographic frozen sections: A peer review comparison study. / Am Acad Dermato! 1989: 20: 670-4. ft Tromovitch TA. Stegman S]. Microscopic-controlled excision of
OPTIMAL TISSUE PREPARATION IN MOHS SURGERY
7 S 9 11)
cutaneous tumours. Chemosurgery, fresh tissue technique. Cancer 1978; 41: 653-8. Cottel Wl, Proper S. Moh's surgery, fresh tissue technique. Our tecbnique with a review. I Dermtito! Simi Onco! 1982; 8: 576-87. Picoto AM. Picoto A. Technical procedures for Mohs fresh tissue surgery. I Dermatol Surti Oncol 1986; 12: 134-8. Hanke CW, Ix^e MW. Cryostat use and tissue processing in Mobs micrographic surgery. / Dermatol Siirei Oncol 1989: 15: 29-32. Honda NS. Friedman DH. A simple method of tissue embedding for Mobs microgrtiphic surgery. / Dermutol Surg Oncol 1989: 1 5: 502-4.
59
1 1 Carter VH. A new method for preparing tissue blocks for cryostat sectioning. / Dfmr.il.i; Surii Oncol 1985: I I : 687-9. 12 Gormley DE. F.valuation of a method for controlled tissue embedding for bistologic evaluation of tumor margins. Am / Dirmalo/wt/ioJ 1987: 9: 308-15. 13 Lesbin B. Cook SR. Frye DW. Cryomold: a device for tissue embedding in Mobs micrographic surgery, j I'h-rmatol Surfi Oncol 1991; 17: 234-6.
