Chordoma: Diagnosis by fine-needle aspiration biopsy with histologic, immunocytochemical, and ultrastructural confirmation

Chordoma: Diagnosis by Fine-Needle Aspiration Biopsy With Histologic, Immunocytochemical, and Ultrastructural Confirmation James L. Finley, M.D., Jan F. Silverman, M.D., David J. Dabbs, M.D., Robert L. West, M.D., Allen Dickens, M.D., Philip S. Feldman, M.D.,

~

and William J. Frable, M.D.

Five cases of chordoma, diagnosed by .fine-needle aspirution (FNA) biopsy, are presented. Four cases were histologically mnjrmed, and in one, immunocytochemical and ultrastructural studies were performed on both the aspirate and tissue specimen. Four cases presented as sacral masses, while in the ,fifth case, a destructive lesion qf the clivus extended into the sofi tissues of the lateral neck. A spectrum of cytomorphologic features was encountered including the presence of abundant microtissue fragments and cells in a dissociate pattern, often with abundant metachromatic extracellular matrix. Steilate and cuhoidal cells often contained intracytoplasmic vacuoles of varying sizes. Intranuclear inclusions, mitotic jigures, and anisonucleosis were prominent features of several cases. Immu- noperoxidase studies on a single case demonstrated cytoplasmic staining for low- and high-molecular-weight cytokeratins, vimentin, and epithelial membrane antigen, while glial fibrillary acidic protein and carcinoenibryonic antigen were negative. Ultrastructural features included the presence of mitochon- drial endoplasmic reticulum complexes, occasional desmosome-like junctions. and abundant extracellular matrix adherent to the tumor cells. We believe thr cytomorphologicjindings are characteristic and, when taken in concert with irnmunocyto- chemicut und ulfraslructurul studies, allow difltrentiation of rhordoma f rom other primary or metastatic neoplasms occur- ring in bone. As demonstrated in our series. chordoma is often un unsuspected diagnosis. Wr believe that FNA biopsy of these lesions can lead to a correct preoperative diagnosis and may also be utilized to document recurrence and thus facilitate the

. . .. . ~ . .

Received February 12, 1986. Accepted July 3. 1986. From the Department of Clinical Pathology and Diagnostic Medi-

cine, East Cadind University School of Medicine, Grcenville, NC; the Department of Pathology, Cytology Division, University of Virginia Medical Center, Charlottesville. VA; and the Department of Pathology, Mcdical College of Virginia, Richmond, VA.

Address reprint requests to James L. Finley, M.D., Department of Clinical Pathology and Diagnostic Medicine, East Carolina University School of Mcdicine, Grecnville, NC 27834.

evaluation and management o f patients with these lesions. Diagn Cytopathol 1986;2:330-337.

Key Words: Chordoma; Fine-needle aspiration cytology; Elec- tron microscopy; Immunocytochemistry

Chordoma is an uncommon tumor of the vertebral col- umn believed to arise from notochordal remnants. Most often located in the sacrococcygeal area, and somewhat less frequently in the sphcno-occipital region, this neoplasm is usually seen in the fourth to sixth decades of life and is slightly more common in males. Although the metastatic potential is variable, there is a high incidence of local recurrence.' Histologically, the differential diagnosis encompasses other primary clear-cell tumors, including chondrosarcoma, rnyxopapillary ependymoma, myxoid liposarcoma, and metastatic clear-cell carcinomas from many sites. This article describes the spectrum of cytologic features in five cases of chordoma. These findings, when taken in concert with immunoperoxidase, histochemical, and ultrastructural studies, allow differentiation of this tumor from other primary and metastatic neoplasms involving bone.

Materials and Methods The percutaneous fine-needle aspiration (FNA) biopsies were performed using a 22-gauge biopsy needle attached to a 20-cc syringe, usually under fluoroscopic or com- puted tomography (CT) guidance. The procedure and smear techniques have been previously described.233 The smears were immediately spray fixed in 9570 alcohol, while a few were air dried. The air-dried smears were stained by a modified Wright stain (Diff-Quick@), while

330 Diagnostic Cytopathology, V o l 2 , N o 4, December 1986 0 1986 IGAKU-SHOIN MEDlChL PUBLISHERS, INC.

CHORDOMA

the alcohol-fixed smears were stained by the standard Papanicolaou technique. A battery of histochemical stains was performed on direct smears and the cell block in case 1 , including periodic acid-Schiff (PAS) with and without diastase digestion and Alcian blue at pH 1.0 and 2.5 with and without prior hyaluronidase digestion. Muci- carmine stains were performed on the FNA smears of cases 1 and 2.

Immunoperoxidase studies were performed on air- dried, acetone-fixed direct smears, FNA cell block, and the methyl carnoy's fixed surgical tissue using the avidin- biotin complex technique according to standard tech- n ique~ .~ In case 1, these included high (34BE12)- and low (35BHll)-moIecular-weight cytokeratins (Enzo Bio- chemical, New York), vimentin (Enzo Biochemical), carcinoembryonic antigen (CEA) (DAKO Patts, Santa Barbara, CA), epithelial membrane antigen (EMA) (DAKO Patts), glial fibrillary acidic protein (GFAP) (DAKO Patts), and 5-100 protein (DAKO Patts). All antibodies were monoclonal except for CEA (rabbit antihuman), GFAP (rabbit antihuman), and s-100 protein (rabbit antihuman), which were polyclonal.

In the cases with surgical confirmation (cases 1, 3, 4, and 5 ) , the specimens were fixed in 10% buffered formal- dehyde and processed in a routine fashion. Sections were stained with hematoxylin- eosin (H&E). Selected cases were also stained with PAS with and without diastase digestion, Alcian blue at pH 1 .O and 2.5 with and without prior h yaluronidase digestion (case 1 ), and mucicarmine (case 1). Oil red-0 staining was performed on air-dried direct smears from the FNA in case 1.

For electron microscopy on case 1, semisolid material from the fine-needle aspiration was fixed overnight at 4 "C in 4% glutaraldehyde and 0.1 mol/l sodium cacody- late buffer (pH 7.4). Dehydration, infiltration, and embedding in Epon 8 12 plastic were performed according to standard techniques. Semithin plastic sections stained with toluidine blue were selected for further study. Ultra- thin sections were mounted on copper grids, stained with

Table I. Clinicoaatholoaic Data

uranyl acetate and lead citrate, and examined with a Zeiss EM 109T electron microscope.

Results Clinical Data Table 1 lists the pertinent clinicopathologic data. All of the patients were elderly, with a male predominance, and sacral lesions were the most common tumors. The patients with sacral lesions presented with back and/or leg pain, while the female patient with a cervical lesion presented with neck pain and latter a mass in the left lateral neck region beneath the sternocleidomastoid mus- cle. The radiographs of the sacral lesions showed typical bony destruction, while the preoperative radiograph of the patient with the cervical tumor showed destruction of the clivus with extension of the mass into the soft tissue of the lateral neck. In case 3 , the FNA biopsy was used to document recurrence. In three of the five patients, chordorna was an unsuspected diagnosis.

Cytologic Findings The cytomorphologic findings showed considerable variability (Table 11). In four cases, microtissue fragments and extracellular matrix material were conspicuous. In the Diff-Quick-stained smears, the matrix was intensely metachromatic (Fig. C- 1 ), while in the Papanicolaou stain, the appearance was fibrillary and eosinophilic. Variable numbers of single dissociated cells and small epithelial clusters were present. Individual cells generally showed low nuclear to cytoplasmic ratios and considerable variability in size. Most cells were cuboidal to polygonal (Figs. (2-2 and C-3), although there were occasional stellate forms (Fig. C-4). Nucleoli were generally small, irregular, and mostly single, although case 1 deviated from this pattern, showing prominent large irregular forms (Fig. (2-5). Similarly, mitotic figures were not observed, except in case 1, where they were numerous (up to three per high-power field). Intranuclear inclusions, prominent in two cases, were accentuated by the

Case AKe number (yr) Sex Site of tumor Clinlcal presentation

1 1 5 M Sacrum Lower back pain

2 I5 M Sacrum Leg and back pain 3 55 F Sacral Back pain

.._"I . I _ . ~

4 66 M Sacrum Back pain and difficulty

5 63 F Clivus Pain and mass in lateral in urination

cervical region

*FNA used to document recurrent chordorna.

Clinical diagnosis

Chordoma versus chondro-

Metastatic prostate cancer Chordoma versus chondro-

sarcoma

sarcoma*

Metastatic prostate cancer

Salivary gland tumor versus malignant lymphoma

Treatment

Surgical excision

Radiation therapy Surgical excision with post-

operative radiation therapy

Surgical excision

Wide local excision with radiation therapy

Diagnostic Cytopathology, Vol2, No 4. December 1986 33 1

FlNLEY ET AL.

Table 11. Cytomorphologic Findings - Case I Case 2 Case 3 Case 4 Case 5

,_ , _ I . I ~ ~ ____~- “ , _- ~~ - ~ . ___-- - __ Microtissue fragments ++++ ++ +++ ++++

Dissociate pattern ++ + + t ++ + +tt+ Extracellular matrix fragments + + + + ++ + + + - t ++++ + Chondtoid cells - ++ Clear cells + .t- + + +* + / - + Physaliphorous cells + + + Steliate cells + +/ - ++ + + Cuboidal cells “t + / - + + + + + +++ Nuclear to Cytoplasmic ratio low low low low/hight low/ high? Anisonucleosis ++++. + + + + + + + + + +++ Nuclear eccentricity + + + + f + + + Binucleation ++ ++ + + ++ Multinucleation I t- - I” ++ + htranuclear inclusion* + + - + / - + + + + Nuclear indentation by + Mitotic figures + & $ lntracytoplasrnic vacuoles +++ ++ ++ ++ .t t

+ I - - - -

- - - +/- -

vacuoles - - - -

(large and small) (large and small)$ (finely vacuolated) (finely vacuolated) (finely vacuolated) Nucleoli Large and irregular Small and irregular Small and irregular Small and irregular Small and irregular

Features are scored from absent (-) to strongly present (+ i- + +); +/-- indicates that the feature is only very rarely found. *Accentuated by Diff-Quik stain. tLow nuclear to cytoplasmic ratio in small cells and higher ratio in large cells. $Up to three mitotic figures per high-power field in selected areas. $Occasional targetoid bodies within vacuoles.

Diff-Quik stain (Fig. C-6). The cells possessed ampho- philic to basophilic cytoplasm. The numerous vacuoles were generally uniform in size except for cases I and 2, where lesser numbers of cells contained large coarse vacuoles (physaliphorous cells), and a few cells had single large vacuoles, which, in general, tended not to indent the nucleus (Fig. (2-7). Cells with large perinuclear vacuoles, having a “chondroid” appearance, were numerous in cases 2 and 5 (Fig. C-8) . Fragments of nonneoplastic bone and cartilage were seen in the first case but were not prcscnt in the other cases.

Light Microscopy and Histochemistry In cases 1, 3, 4, and 5 , surgical tissue was available for microscopic studies. In these cases, the tumors showed fibrous septa dividing the neoplasm into lobules of varying sizes (Fig. C-9). The neoplastic cells and lobules were surrounded by a basophilic matrix of mucinous material. Individual cells were predominantly round to polygonal with occasional stellate forms. In some areas, they were arranged in solid sheets, while in other regions, cords and small clusters were present. The cells had pale eosinophilic cytoplasm, which was predominantly finely vacuolated, although scattered cells had large intracytoplasmic vacuoles (Fig. C- 10). The nuclei showed marked anisonucleosis with a granular dense chromatin pattern and predominantly single small round nucleoli. Intranuclear inclusions were a prominent feature in one case (case 4 ) and wcrc seen less frequently in two other cases (cases 1 and -5) (Fig. C-10).

Histochemical studies performed on direct smears and the surgical tissue in case 1 showed strong matrix staining with Alcian blue a t pH 2.5 and pH 1.0, which was almost completely digested with testicular hyaluronidase. Mod- erate peripheral cytoplasmic and faint intracytoplasmic staining were seen in the smears and tissue sections with the Alcian blue stain. A mucicarmine stain on a direct smear from case 2 was negative, although strong positivity was seen in the tissue specimen from case. 1.

A PAS stain showed strong staining of the extracellular matrix material and globular intracytoplasmic masses, which were removed by diastase digestion. An oil red-0 stain performed on the cytologic smears (case 1) showed positive staining within small vacuoles, although the larger vacuoles were negative.

Immunoperoxidase Findings

Tmmunoperoxidase studies performed in the first case both on the material obtained by fine-needle aspiration biopsy (direct smears and cell block) and on the surgical tissue showed similar results. An antibody directed against low-molecular-weight (K35) cytokeratin demonstrated strong cytoplasmic staining of the tumor cells (Fig. C- 1 l ) , while high-molecular-weight (K34) cytokeratin showed weak staining in the majority of the tumor cells. Antibodies against vimentin and EMA showed strong cytoplasmic staining, which appeared accentuated around the cytoplasmic vacuoles (Fig. C-12). The CEA and GFAP markers were negative with satisfactory con-

332 Diagnostic Qtopathology. Vol2. N o 4 , December I986

Fig, c-1 Fig. C-2

Fig. C-3

Fig. C-5

Fig. C-4

Fig. C-6

Fig. C-1. Matrix fragment showing intense metachromatic staining (Diff-Quik stain, original magnification, x 250). fig. C-2. Round to cuboidal cells in small clusters generally showing low nuclear to cytoplasmic ratios (Papanicolaou stain, original magnification, x 100). Fig. C-3. Loose clusters of polygonal-shaped cells with low nuclear to cytoplasmic ratio and surrounding pale cytoplasm (Diff-Quik, original magnification, x 400). Fig, C-4. Occasional spindle-shaped cells are present within small epithelial clusters. Note occasional finely vacuolated cells (Papanicolaou stain, original magnification, x 400). Fig. C-5. Cells demonstrating nuclear pleomorphism with large irregular nucleoli and surrounding metachromatic stroma (Papanicolaou stain, original magnification, x 400). Fig. C-6. Large intranuclear inclusion is accentuated by the Diff-Quik stain (Diff-Quik stain, original magnification, x 400).

Fig. C-1 Fig. C-8

Fig. C-9 Fig. C-10

Fig. C-11 Fig. C-12

Fig. C-7. Occasional physaliphorous cells demonstrating large coarse vacuoles often obscuring nuclear detail (Papanicolaou stain, original magnification, x 400). Fig. C-8. Cells with large perinuclear vacuoles (center) display a “chondroid” appearance (Papanimlaou stain, original magnification, x 250). Fig. C-9. Tissue section ofchordoma showing fibrous septae dividing the tumor into lobules of varying sizes (H&E, original magnification, x 250). Fig. C-10. Tissue section of chordoma showing vacuolated nature of cells and intranuclear inclusions (H&E, original magnification, x 400). Fig. C-11. Immunostaining of tumor cells obtained by FNA biopsy using an antibody against low-molecular-weight cytokeratin. Diffuse cytoplasmic staining is seen within the tumor cells (immunoperoxidase stain, original magnification, x 400). Fig, C-12. Irnmunostaining of tumor cells obtained by FNA biopsy using an antibody against virnentin. The cytoplasmic staining is accentuated around the large vacuoles (immunoperoxidase stain, original magnification, x 400).

CHORDOMA

trols. Occasional tumor cells were weakly positive for 23-100 protein.

Ultrastructural Studies Studies performed on material obtained by fine-needle aspiration biopsy and on surgical tissue demonstrated similar features. The tumor cells formed tightly cohesive clusters with close approximation and interdigitation of cytoplasmic membranes. The extracellular space contained a granular amorphous fiocculant material closely adherent to the plasma membranes. Occasional cells showed well-developed desmosome-like junctions. Within the cell cytoplasm, abundant organelles and intermediate filaments were seen. Stacks of rough endoplasmic reticulum, which tended to encircle mitochondria1 complexes (MER-complexes), were particularly prominent (Fig. 13). Small droplets of material with a medium density suggestive of lipid material were also seen. Some cells contained diffuse arrays of intermediate filaments to the exclusion of most other organelles. The cell nuclei were

Fig. 13. Electron microscopy performed on material obtained by FNA biopsy. Stacks of rough endoplasmic reticulum encircling mitochondria (MER-complexes) are characterically seen in chordoma ( x 12,100).

round to oval and contained dispersed chromatin with small nucleoli.

Discussion The diagnosis of chordoma by FNA biopsy has been infrequently made, as reflected by review of large series of FNA biopsies of b ~ n e . ~ - ~ In one series of 1,000 bone FNAs, chordoma was not en~ountered.~ In another series of 430 biopsies, five cases of chordomas were diagnosed (1.2%).* In two studies from Memorial Hospital, repre- senting approximately 550 cases, only six cases of chordoma were enc~untered.~ '~ There have been only four prior reports describing the FNA cytologic features of c h o r d ~ m a s ' ~ ' ~ and three cases diagnosed by intraopera- tive touch-prep/squash-smear technique.'"I6 Elliott et a1." diagnosed a metastatic chordoma to the skin by the FNA technique, while O'Dowd and Shumannl' were the first to report a preoperative diagnosis of coccygeal chordoma by FNA cytology. We report five additional cases of chordoma diagnosed largely preoperatively by

Diagnostic Cytopathology, Vol2, N o 4. December 1986 335

HNLEY ET AL.

this technique and detail the spectrum of cytologic features of this primary bone neoplasm.

A number of different cell types and patterns have been noted in the prior FNA and touch-prep cytologic exami- nations of chordoma. Cell types include physaliphorous cells with large vacuoles,' i~14~16*17 tumor giant-cells (mono- nuclear or m~ltinucleated), '~"~ and smaller nonvacuo- lated ~ e l l s . ' ~ ~ ' ' ~ ~ ~ Some authors state that the nuclei of chordoma cells have a bland uniform hypochromatic appearance,'' while others describe cells having hyper- chromatic nuclei with nucleoli and irregular chromatin clumping.1n*'6 The assessment of mitotic activity varies from infreq~ent '~J ' to occasional mitotic figure^.'^ O'Dowd and Schumann" reported in their case of coccygeal chordoma that the specimen was hypocellular. Large (30--50 p), round, highly vacuolated clear cells having a low nuclear cytoplasmic ratio were either singly arranged or present in two-dimensional cords and sheets with occasional three-dimensional clusters. The finding of three-dimensional clusters might reflect their preparatory technique, since the finc-needle aspirate consisted of 2 cc of bloody fluid suspended in 10 cc of saline, from which cytocentrifuge smears were prepared and stained by the Papanicolaou method. Our experience and other reports emphasize the moderate to hypercellular nature of the aspirate when utilizing direct smears." Additional findings noted in chordomas include perinuclear halos" and cannibalism by the cells.14 We believe the two most important diagnostic features of chordoma are the presence of vacuolated cells and background myxoid stroma. Classic physaliphorous cells, although not uniformly seen, are extremely helpful for diagnosis when present,

The FNA biopsy differential diagnosis of chardoma includes other primary and metastatic clear-cell tumors. Entities to be considered include chondrosarcoma, myxo- papillary ependymoma, alvcolar soft-part sarcoma, myxoid liposarcoma, and metastatic clear-cell carcinomas from a number of primary sites including kidney, adrenal, lung, and liver. Other considerations include metastatic amelanotic melanoma and leiomyoblastoma." The mucin positivity of chordoma should differentiate them from metastatic acinic cell, Clara-cell, renal and adrenal cell, hepatoma, pheochromocytoma, alveolar soft-part sarcoma, clear-cell leiomyosarcoma, and amelanotic melanoma." When differentiating chordoma from either primary or metastatic mucin-producing tumors, one must rely on both the cytologic features of the neoplastic cells and the types of intra- and intercellular mucin produced by the neoplasm. Features favoring a mucin-producing epithelial malignancy include either the presence of co- lumnar cells arranged in acinar or glandular configura- tion and/or the presence of neutral rather than acidic mucin. ' '

Probably the major differential diagnostic problem is

the separation of chordoma from chondrosarcorna.'s-2' Features of chondrasarcoma include the presence of single cells located in lacunar spaces, metachromatic chondroid background substance staining with variable intensity, and tumor cells having varying appear- a n c e ~ . ~ ~ * ~ ~ - ~ ~ ~ ~ ~ Chondrosarcoma can be composed of iso- lated cells as well as cells arranged in sheets and loose aggregates with distinct cytoplasmic borders." The cytoplasm varies from foamy to vacuolated. Large vacuoles can displace the nucleus, forming signet ring cell^.^^'^^ Occasional binucleated to multinucleated tumor cells can be present.lg Bizarre tumor cells having enlarged nuclei with nucleoli have been n ~ t e d , ' ~ along with cells having optically clear nucleL2" Occasional chondrosarcomas can have fibroblastic-like spindle cells, which would not be appreciated in chord~mas. '~ Perhaps the most important diagnostic feature of chondrosarcoma is the presence of fragments of chondroid material in the ~ m e a r s . ' ~ ' ~ ~ In addition, chordoma cells differ from chondrosarcoma cells since the latter do not show the extensively vacuolated or bubbly cytoplasm seen in the diagnostic physaliphorous cells.'4 I n contrast, the vacuolated cells of chondrosarcoma often demonstrate perinuclear halos and/or large vacuoles displacing the nucleus. The metachromatic staining of the cytoplasm is said to be different in chondrosarcoma than in chordoma, with the former demonstrating staining at the periphery of the cell where it merges with the matrix or stroma, while the latter stains unif0rm1y.l~

Myxoid liposarcoma can be confused with chordoma, but a helpful feature is the presence of malignant spindle- shaped cells set in a myxoid stroma." Physaliphorous cells can be distinguished from lipoblasts, since the latter will generally show nuclear impressions or indentation by vacuoles and lack a bubbly ~ y t o p l a s m . ~ ~ . ~ ~

Utilization of histochemical staining may also aid in the differentiation of chordomas from other sarcomas, including chondrosarcoma and liposarcoma. Alcian blue staining at varying pH can characterize the glycosamino- glycan content of myxoid tumors and thereby help in this distinction.2' Liposarcoma will stain with Alcian blue at pH 2.5, but not at pH 1, since chondroitin-sulfate and kerato-sulfate are found in chordomas and not liposarcomas.23 Prior testicular hyaluronidase digestion has been reported to eliminate the Alcian blue staining of liposarcomas, indicating the presence of hyaluronic acid, but will not remove the sulfated glycosaminoglycans of chondrosarcomas and c h ~ r d o m a s . ~ ~ This was not confirmed in the histochemical studies performed on our first case since the hyaluronidase digestion eliminated thc positive staining of the chordoma cells. Chordomas are reportedly mucicarmine positive in contrast to chandrosarcomas and liposarcomas."

In conjunction with the cytologic features, immunope-

336 Diagnostic Cytopathology, Vol2, No 4, December 1986

CHORDOMA

roxidase studies and electron microscopy can considera- bly narrow the differential diagnoses. The finding of both vimentin and epithelial (EMA and cytokeratin) markers within tumor cells has been reported in a limited number of epithelial neoplasms, malignant mesothelioma, and epithelial and synovial sarcoma.24 Myxoid liposarcoma and chondrosarcorna should not stain positively for epithelial membrane antigen or low-molecular-weight cytokeratin. Alternatively, metastatic prostate carcinoma, a common clinical diagnosis in this patient population, generally does not show vimentin positivity and will usually be positive for prostate-specific antigen. Myxo- papillary ependymoma, another uncommon tumor of this location, is GFAP positive while chordoma and chondrosarcoma are GFAP negatives2' Our findings are in accord with those reported by Salisbury and Isaacson,26 who studied adult and fetal cartilage, chondrosarcoma, chordoma, and fetal notochordal vestiges and demonstrated positive staining in surgical tissue with antibodies directed against EMA and cytokeratin in chordomas. Kontozoglou et al. described positive staining for neuron- specific enolase (NSE), s-100 protein, EMA, and keratin in a small series of FNA biopsies of chordoma.13

Vimentin positivity of chordoma has not been previously described. We believe this to be the first report detailing the application of an immunoperoxidase panel showing cytokeratin, vimentin, and EMA positivity with negative staining for GFAP and CEA on cytologic material in the diagnosis of chordoma.

Ultrastructural features can confirm and complement the histologic and cytologic findings. The MER-complex, first described by Erlandson et al.,27 consists of parallel arrays of alternating endoplasmic reticulum and dilated mitochondria and are characteristically seen only in chordoma and chondroid chordoma,28 The ultrastructural examination performed on the FNA material in case 1 demonstrated these characteristic complexes.

In summary, a spectrum of cytologic changes are described in five chordomas diagnosed by the fine-needle aspiration biopsy technique. While the cytologic features are relatively specific, application of histochemistry, immunocytochemistry, and electron microscopy on the FNA material can further aid in establishing a specific primary diagnosis or in documenting tumor recurrence.

References 1. Eriksson 9, Gunterberg B, Kindblom L. Chordoma. A clinicopathologic and prognostic study. Acta Orthop Scand 1981;52:49-58.

2. Bonfiglio TA. Cytopathologic interpretation of transthoracic fine needle biopsies. New York Mason, 1983:7-12.

3. Sinner WN. Pulmonary neoplasms diagnosed with transthoracic needle biopsy. Cancer 1979;43: 1533-40. 4. Hsu SM, Raine L, Fanger H. Use of avidin-biotin peroxidase complex in immunoperoxidase technique: a comparison of ABC and an unlabeled antibody (PAP) procedure. J Histochem 1981;29:57740. 5. Schajowicz F. Aspiration biopsy in bone lesions. Cytological and histological techniques. J Bone Joint Surg 1955;37:465-71. 6. Hajdu SI, Melamed MR. Needle biopsy of primary malignant bone tumors. Surg Gynecol Obstet 1971;133:829-32. 7. Snyder RE, Coley BL. Further studies on the diagnosis of bone tumors by aspiration biopsy. Surg Gynecol Obstet 1954805 17-22. 8. Xiaoling P, Xiangcheng Y. Cytodiagnosis of bone tumors by fine needle aspiration. Acta Cytol 1985;29:570-5. 9. Stormby N, Akerman M. Cytodiagnosis of bone lesions by fine needle aspiration. Acta Cytol 1973;17:166-72. 10. Elliott EC, McKinney S, Banks H, Fulks RM. Aspiration cytology of metastatic chordoma. A case report. Acta Cytol 1983;6:658-62. 11. O'Dowd GJ, Schumann GB. Aspiration cytology and cytohisto- chemistry of coccygeal chordoma: A case report and review of the literature. Acta Cytol 1983;27:178-83. 12. Carvalho G, Coelho LH. Chordoma of rhinopharynx. Report of a case. Acta Cytol 1974;18:425-8. 13. Kontozoglou T, Qizilbash AH, Sianos J, Stead R. Chordoma: cytologic and immunocytochemical study of four cases. Diagn Cyto- pathol 1986255-61. 14. Lefer L, Rosier PR. The cytology of chordoma. Acta Cytol

15. Gandolf A. Vertebral chordoma. lntraoperative cytologic diagnosis. Acta Neurol 1979;34:156-60. 16. Nguyen GK, Johnson ES, Mielke BW. Chondroid chordoma of the skull base diagnosed by squash smear technique. Diagn Cytopathol 1985;1:161-3. 17. Sanerkin NG, Jeffree GM. Cytology of bone tumors. Philadelphia: JB Lippincott, 198051-73. 18. Linsk JA, Franzen S. Clinical aspiration cytology. Philadelphia: JB Lippincott, 1983:349-59. 19. Koss LG, Woyke S, Olszewsk W. Aspiration biopsy. Cytologic interpretation and histologic bases. New York: Igaku-Shoin, 1984:431- 32. 20. Giszewski W, Woyke S , Musiatowicz B. Fine needle aspiration biopsy cytology of chondrosarcoma. Acta Cytol 1983;27:345-9. 21. Tsang S, Nguyen GK. Aspiration biopsy cytology of metastatic extraskeletal myxoid chondrosarcoma and its cytologic differential diagnosis, Acta Cytol 1985;29:566-9. 22. Frable WJ. Thin needle aspiration biopsy. Philadelphia, WB Saun- ders, 1983:286324. (Major problems in pathology, vol 14). 23. Walaas L, Kindbloom LG. Lipomatous tumors: a correlative cytologic and histologic study of 27 tumors examined by fine needle aspiration cytology. Hum Pathol 1985;16:6-18. 24. Gouin A, Vogel A. Monoclonal antibodies to human intermediate filament proteins. Am J Clin Pathol 1985;84:413-24. 25. Miettinen M, Lehto VP, Dahl D, Virtanen I. Differential diagnosis of chordoma, chondroid and ependymal tumor as aided by anti- intermediate filament antibodies. Am J Pathol 1983;112: 160-9. 26. Salisbury J, Isaacson P. Demonstration of cytokeratins and an epithelial membrane antigen in chordomas and human fetal notochord. Am J Surg Pathol 1985;9(11):791-7. 27. Erlandson RA, Tandler B, Lieberman P, Higinbotham N. Ultra- structure of human chordoma. Cancer Res 1968;28:2115-25. 28. Valderrama E, Kahn L, Lipper S, Mare J. Chondroid chordoma: electron microscopic study of two cases. Am J Surg Pathol 1983;7:625- 32.

1978;22:5 1-3.

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Chordoma: Diagnosis by fine-needle aspiration biopsy with histologic, immunocytochemical, and ultrastructural confirmation