Differential diagnosis of lymph node aspirates by intermediate filament typing of tumor cells

Twenty-one lymph node aspirates for which a differential diagnosis was difficult or not possible by light microscopy alone were selected for further study by intermediate filament typing. The use of four monoclonal antibodies specific for different keratin subsets or for vimentin resulted in a definitive diagnosis in all instances. The results show that use of these antibodies can improve the accuracy of cytologic diagnosis and provide further evidence that intermediate filament typing can help differentiate (1) malignant melanoma from adenocarcinoma, (2) malignant lymphoma from small-cell anaplastic carcinoma and (3) squamous-cell carcinoma from adenocarcinoma. Intermediate filament typing can also be used to identify very small numbers of carcinoma cells in specimens that are apparently negative for tumor cells by light microscopy. The method is quick, relatively simple, reliable and unambiguous, provided that appropriate questions are asked and antibodies with well-defined specificities are used.     

Neuroendocrine (Merkel-cell) carcinoma of the skin. Cytology, intermediate filament typing and ultrastructure of tumor cells in fine needle aspirates

The light and transmission electron microscopic findings and the intermediate filament typing of tumor cells from fine needle aspirates of primary, recurrent and metastatic neuroendocrine (Merkel-cell) carcinoma of the skin are described. The tumor cells in the smears coexpressed keratins and neurofilaments and were characterized by "intermediate filament buttons," i.e., buttonlike fragments of cytoplasm of tumor cells, which could be observed either by staining with intermediate filament-specific antibodies using immunofluorescence or by their appearance in hematoxylin-and-eosin-stained smears. These features may help in the differential diagnosis of fine needle aspirates of neuroendocrine carcinomas of the skin.     

Intermediate filament typing of tumor cells in fine needle aspirates by means of monoclonal antibodies

Well-characterized monoclonal antibodies directed against different intermediate filament proteins were used in the typing of tumor cells in 30 fine needle aspiration (FNA) biopsy specimens received for routine cytologic examination to assess the value of intermediate filament typing in FNA cytology. Tumors from sites that included liver, large bowel, pancreas, breast, skin, thyroid, thigh and kidney were examined with monoclonal antibodies specific for either all keratins or keratin subsets, vimentin, desmin or specific neurofilament polypeptides. Intermediate filament typing helped to confirm, revise or refine the diagnoses made by light microscopy and provided information of value in the classification of tumors of uncertain origin.     

Immunochemical demonstration of keratin and vimentin in cytologic aspirates

Antibodies to intermediate filament proteins were used to characterize tumor cells present in peritoneal and pleural effusions and in thin needle aspirates from palpable lymph nodes. Metastatic adenocarcinoma cells (breast, ovary, endometrium, cervix, colon and stomach) as well as squamous-cell carcinomas and mesotheliomas stained specifically with antibodies to keratin while mesenchymally derived tumor cells (lymphomas, melanoma, fibrosarcoma and neurofibrosarcoma) were positive only for vimentin. Especially in cases of lymph node aspirates, keratin staining in cells was a direct indication of metastatic carcinoma. Antibodies to these different components of the cytoskeleton can thus be used in cytopathologic diagnosis when a definitive diagnosis cannot be made on the basis of conventional cytologic features.     

Endothelial cells help in the diagnosis of primary versus metastatic carcinoma of the liver in fine needle aspirates. An immunofluorescence study with vimentin and endothelial cell-specific antibodies

Fine needle aspirates of 5 primary hepatocellular carcinomas and 24 carcinomas metastatic to the liver were studied using vimentin and endothelial cell-specific monoclonal antibodies. Numerous endothelial cells dispersed and in bundles overlying clumps of tumor cells were positively stained by both antibodies in smears of primary hepatocellular carcinomas while such cells were rare or absent in metastatic carcinomas, with the exception of clear cell carcinoma of the kidney. It is concluded that endothelial cells, if present in large numbers in fine needle aspirates of a hepatic carcinoma and arranged in bundles that envelope the clumps of tumor cells, can (1) suggest the presence of a primary hepatocarcinoma and (2) narrow the differential diagnosis with the most common metastatic cancers to renal cell carcinoma.     

Malignant melanoma in fine needle aspirates and effusions. An immunocytochemical study using monoclonal antibodies

The value of monoclonal antibodies (MAbs) for the immunodetection of keratin, vimentin and two melanoma-associated antigens recognized by NKI/C3 and NKI/Bteb for the diagnosis of malignant melanoma has been previously established on histologic preparations. In the present study, cytologic preparations from 20 fine needle aspirates and effusions from patients with malignant melanoma were evaluated using these antibodies. Twenty of 20 smears were negative for keratin, and 20 of 20 smears were positive for vimentin. Positivity for NKI/C3 was seen in 12 of 12 cases studied, and for NKI/Bteb in 12 of 13 cases. These results indicate that a panel of MAbs consisting of anti-keratin, anti-vimentin, NKI/C3 and NKI/Bteb is useful for a more accurate diagnosis of malignant melanomas on cytologic preparations. The expression of these antigens in melanoma cells in cytologic smears can be a valuable aid in the detection of primary (noncutaneous) and metastatic melanomas by fine needle aspiration.     

Parotid metastasis of Merkel cell carcinoma in a young patient with ectodermal dysplasia. Diagnosis by fine needle aspiration cytology and immunocytochemistry.

Fine needle aspiration (FNA) biopsy was performed on an intraparotid lymph node metastasis of a Merkel cell carcinoma of the eyelid in a 15-year-old girl with antecedent ectodermal dysplasia syndrome. The cytologic appearance of the aspirate and the results of immunocytochemical typing of intermediate filaments on the FNA smears provided a definitive diagnosis. The Romanowsky stain provided an excellent delineation of paranuclear intracytoplasmic "buttons," which appeared to contain both cytokeratin and neurofilaments by immunocytochemical studies. These findings confirm previous data emphasizing the role of light microscopic observations, supplemented by proper immunocytochemical investigations, in the differential diagnosis of metastatic Merkel cell carcinoma in fine needle aspirates.     

Cytoskeletal proteins as tissue-specific markers in cytopathology

Antibodies to intermediate filament proteins react in a tissue-specific manner and can be used to characterize tumor cells present in thin-needle aspirates from solid tumors, from palpable lymph nodes and cells present in samples from peritoneal and pleural effusions. From our studies so far the following conclusions can be drawn: Polyclonal antisera to cytokeratins can identify carcinoma metastases in thin-needle aspirates from palpable lymph nodes and distinguish them from malignant lymphomas and nonmalignant lesions such as chronic lymphadenitis, which show only vimentin-positive cells. Monoclonal antibodies to specific cytokeratin polypeptides are able to distinguish between different types of epithelial tumor metastases, i.e. metastases from adenocarcinomas and metastases from squamous cell carcinomas. Cells present in peritoneal and pleural effusions can be partly characterized using intermediate filament antisera. We have found that metastatic adenocarcinoma cells from breast, ovary, endometrium, cervix, colon and stomach, as well as squamous cell carcinomas and malignant mesothelioma stain specifically with antibodies to cytokeratin while mesenchymally derived tumors such as malignant lymphomas, malignant melanomas, and fibrosarcomas, are positive for vimentin only. Metastatic tumor cells of epithelial origin present in aspirates from human serous body cavity fluids may coexpress vimentin next to their original cytokeratin intermediate filaments. Benign mesothelial cells present in body cavity fluids frequently coexpress cytokeratins and vimentin. Tumor cells present in thin-needle aspirates from solid tumors such as pleomorphic adenomas of the parotid gland can be identified as such because of their typical patterns of intermediate filament (co-)expression.(ABSTRACT TRUNCATED AT 250 WORDS)     

Leukocyte-common antigen and vimentin are reliable adjuncts in the diagnosis of non-Hodgkin's lymphoma in fine needle aspirates

Alcohol-fixed fine needle aspirates of 82 non-Hodgkin's malignant lymphomas (NHLs) were tested for the presence of vimentin and leukocyte-common antigen (LCA) by means of monoclonal antibodies (MAbs) and indirect immunofluorescence. All NHLs stained positively for vimentin; the staining was strong in all except three cases. Of the 69 NHLs tested for LCA, 1 (a large cell T-cell lymphoma) was negative while the staining was weak in 6. Thus, vimentin and LCA MAbs are sensitive, specific and reliable complementary diagnostic adjuncts that are useful in the definitive diagnosis of NHLs in alcohol-fixed fine needle aspirates. Their presence in the aspirate confirmed a cytologic diagnosis of NHL in 47 cases, helped to diagnose NHL in 31 cases in which a cytologic differential diagnosis with small cell anaplastic carcinoma could not be made with confidence and helped to change the initial cytologic diagnosis of anaplastic carcinoma to NHL in 4 cases.     

Fine needle aspiration cytology of neuroendocrine tumors of the pancreas. A cytologic, immunocytochemical and electron microscopic study.

We report the fine needle aspiration cytology findings in six cases of neuroendocrine tumor of the pancreas. Three cases were from the pancreas, two from hepatic metastases and one from a peripancreatic lymph node metastasis. The cytologic features that permitted a preoperative diagnosis of pancreatic neuroendocrine tumor were: a cellular aspirate; numerous isolated cells and irregular, loose, dyshesive cellular aggregates; minimal nuclear pleomorphism; infrequent mitoses; fine, evenly dispersed nuclear chromatin with occasional inconspicuous nucleoli; a scant-moderate amount of granular, amphophilic, well-defined cytoplasm; clustering of tumor cells around segments of capillaries; and rosette formation. The differential diagnosis includes cells derived from normal pancreatic acini, islet cell hyperplasia, acinic cell carcinoma, well-differentiated pancreatic adenocarcinoma, metastatic small cell undifferentiated carcinoma of the lung, pancreatic small cell anaplastic carcinoma and malignant lymphoma. The application of immunocytochemistry to cytologic smears can be easily and reliably performed to confirm the neuroendocrine nature of the tumor and identify the specific type of polypeptide hormone or hormones produced by these tumors. Four aspirates showed immunoreactivity for chromogranin, and one was positive for gastrin. Cells of a lipid-rich neuroendocrine tumor were negative for chromogranin; however, the tissue section contained neuron specific enolase, and neurosecretory granules were demonstrated by electron microscopy.     

Leiomyosarcoma of the breast: report of a case with fine needle aspiration cytologic, histologic and immunohistochemical features

BACKGROUND: Leiomyosarcoma of the breast is a rare neoplasm. We present a case of primary leiomyosarcoma of the breast in a middle-aged female in whom fine needle aspiration cytologic features suggested sarcoma. CASE: A 55-year-old female presented with a rapidly growing breast lump of 1 month's duration. On examination, an ulcerating, 12 x 10 cm tumor was seen involving the lower medial and lateral quadrants of the right breast. Fine needle aspiration cytology showed variably sized, dissociated and loosely clustered polygonal, plump and spindle cells with pale blue cytoplasm and vesicular nuclei that were round, oval or irregular. Occasional giant forms and nucleolated and mitotic cells were present. A single cluster of benign ductal cells was seen. The tumor cells did not express immunocytologic reactivity to estrogen receptor protein. A cytologic diagnosis of sarcoma was given with differential diagnoses of metaplastic carcinoma and malignant phyllodes tumor. Histologic study established the diagnosis of leiomyosarcoma. Leiomyosarcoma of the breast shows fine needle aspiration cytologic features of sarcoma, but specific tumor typing may not be possible, especially when the cytologic material is inadequate for ancillary staining required to distinguish leiomyosarcoma from metaplastic carcinoma and malignant phyllodes tumor.     

Cytologic findings in primary malignant carcinoid tumor of the cervix. Including immunohistochemistry and electron microscopy performed on cervical smears

The cytologic findings in a primary malignant carcinoid tumor of the cervix are presented. In addition to the presence in the smears of cells suggestive of squamous carcinoma and adenocarcinoma, which led to an initial diagnosis of adenosquamous carcinoma, there were multinucleated giant cells with prominent, reddish nucleoli, finely granular chromatin and grayish-blue to eosinophilic cytoplasm, as well as smaller pleomorphic cells, against a tumor-diathesis type of background. Immunocytochemistry performed on the cervical smears showed the presence of serotonin, and ultrastructural analysis revealed abundant intracytoplasmic, membrane-bound granules in malignant cells, thus confirming the diagnosis of a carcinoid tumor.     

Immunocytologic diagnosis of small-cell lung cancer in imprint smears.

Imprints of histologic or autopsy specimens from 12 small-cell lung cancers (SCLCs), 82 non-SCLCs (50 adenocarcinomas, 25 squamous-cell carcinomas, 1 adenosquamous carcinoma and 6 large-cell carcinomas), 2 carcinoid tumors, 1 malignant lymphoma and 8 metastatic carcinomas were examined immunocytologically for the presence of cluster 1 SCLC antigen (neural-cell adhesion molecule: N-CAM), chromogranin A, Leu-7, neuron-specific enolase (NSE) and gastrin-releasing peptide (GRP). The monoclonal antibodies NCC-LU-243 and NCC-LU-246, which are reactive with cluster 1 SCLC antigen/N-CAM, diffusely stained the cell membranes of all SCLCs and carcinoid tumors (100%) and diffusely and focally stained those of two of the large-cell carcinomas, two of the adenocarcinomas, two of the squamous-cell carcinomas and the one adenosquamous carcinoma. Malignant lymphoma and metastatic carcinoma were negative for this antigen. A few cases of large-cell carcinoma, adenocarcinoma, squamous-cell carcinoma and adenosquamous carcinoma were also stained with these antibodies, which may indicate a neuroendocrine differentiation. However, these tumors were different from SCLCs in that their positive tumor cell population was definitely smaller than that in SCLC, in which almost all tumor cells were positive. This confirmed the usefulness of antibodies against cluster 1 SCLC antigen for the immunocytologic diagnosis of SCLC and carcinoid tumor in imprint smears. Chromogranin A, GRP, NSE and Leu-7 were not useful in immunocytologically differentiating the imprints from these cases since only a few tumor cells were reactive with these antibodies. The antibodies against cluster 1 SCLC antigen/N-CAM can also be applied to cytologic preparations of sputum, pleural fluid and fine needle aspirates stained routinely by the Papanicolaou method since the antigen is preserved in such alcohol-fixed smears.     

Immunocytochemistry in head and neck aspirates. Diagnostic application on direct smears in 16 problematic cases.

This report describes our experience with immunocytochemical staining of routinely processed smears in the fine needle aspiration (FNA) biopsy diagnosis of 16 tumors of the head and neck. Immunocytochemistry (ICC) was performed on alcohol-fixed or air-dried smears using commercially available monoclonal antibodies followed by a streptavidin-biotinylated peroxidase labeling method. In 12 aspirates with cytologically unclassifiable and undifferentiated cells, immunostaining for cytokeratin, leukocyte common antigen, S-100 protein and vimentin provided conclusive evidence of cell lineage. ICC permitted the correct identification and differential diagnosis of four additional tumors: a positive immunoreaction for thyroglobulin identified a metastatic Hürthle cell carcinoma of the thyroid; a coexpression of two distinct classes of intermediate filaments helped support the FNA diagnoses of a parathyroid adenoma and of a synovial sarcoma; and the double immunoreaction for CD15 and CD30 antigens helped identify Reed-Sternberg cells within an unusually suppurative harvest. Two immunostains were required for proper diagnosis in 13 cases and four in the remaining 3. In all cases but one only unstained slides were used. These data demonstrate that immunostaining can conveniently and advantageously be performed on direct smears of aspirated samples of head and neck lesions, but cases should be carefully selected for this procedure.     

Carcinoembryonic antigen content in fine needle aspirates of the lung. A diagnostic adjunct to cytology.

Carcinoembryonic Antigen (CEA) was measured in 59 consecutive fine needle aspirates (FNAs) of the lung from 58 patients to determine if the CEA content would enhance the sensitivity of the cytologic diagnosis. Twenty-eight males and 30 females with tumors 1-40 cm in diameter were studied. Final diagnoses were correlated with the clinical history, radiologic studies, tissue (when available) and follow-up. Image-guided FNAs were performed by radiologists using a 22-gauge Chiba needle and 20-mL syringe with one to four passes per specimen. Cytologic examination included rapid assessment in the radiology suite and a final diagnosis in 24 hours. CEA was measured by enzyme immunoassay using monoclonal antibody. Nine benign aspirates and 50 malignant aspirates were diagnosed. The sensitivity of cytology was 86% and specificity, 100%. Using 5 ng/mL as the cutoff, the sensitivity of CEA for malignant aspirates was 50% and specificity, 90%. The combined sensitivity of CEA and cytology was 95%. The mean CEA in malignant aspirates was 131 ng/mL and in benign aspirates, 2.41. The highest mean CEA was seen in adenocarcinoma, 402.6 ng/mL. Lower CEA content was seen in epidermoid carcinoma (58.6 ng/mL), large cell carcinoma (8.09), oat cell carcinoma, metastatic carcinoma of the kidney and breast, thymoma and lymphoma (each less than 1 ng/mL). Elevated CEA alone was diagnostic in two aspirates of bronchioloalveolar carcinoma; carcinoma with an unknown primary source, three; and large cell carcinoma, one. The adjunctive use of CEA in FNAs of the lung enhances the sensitivity of the cytologic diagnosis.     

The use of antibodies to intermediate filament proteins in the differential diagnosis of lymphoma versus metastatic carcinoma

Summary Forty-nine cases encompassing 16 different types of malignant lymphoma were examined for their intermediate filament protein (IFP) type by indirect immunofluorescence microscopy of cryostat sections. In all cases, vimentin was shown to be the only IFP type detectable in these tumours. Lymphomas are negative for keratin and desmin, which are characteristic for benign and malignant epithelial or muscular tissues respectively. In addition, eighteen cases are described in which antibodies to intermediate filament proteins were used successfully to distinguish between lymphoma and metastatic carcinoma where differential diagnosis was difficult or impossible on the basis of routine histology.     

A new epithelial membrane antigen (Calam 27) as a marker of carcinoma in serous effusions

A new monoclonal antibody (Calam 27) that reacts with a membrane antigen present on cells of epithelial origin, but not on cells of mesothelial origin, was investigated as a means of distinguishing between mesothelial cells and malignant cells in cytologic smears of serous effusions from patients with carcinoma. Immunofluorescence staining of cells in 151 effusions from 109 patients with different diseases showed a good correlation between the cytologic diagnosis on routine preparations and the staining with Calam 27. Calam 27 was also used to study the ploidy and cell cycle kinetics of carcinoma cells versus reactive mesothelial cells and normal cells by flow cytometry; these experiments confirmed that Calam 27 is not reactive with mesothelial cells. In conclusion, Calam 27 staining can help to confirm the cytodiagnoses in cases with carcinomatous effusions.     

Development of monoclonal antibodies that recognize antigens associated with human cervical carcinoma

Six monoclonal antibodies, generated by immunization of mice with human cervical carcinoma cells maintained in tissue culture or with cells from fresh tumor tissue, reacted specifically with the malignant cells in 71% to 90% of the tumor tissue imprints and cervical smears containing neoplastic cells but not with normal cervical epithelial cells in smears from 21 to 23 healthy donors. Antibody CE 402 bound to epithelial cells associated with regeneration in 2 of the 23 normal smears tested. Considerable heterogeneity of antibody binding by malignant cells was observed. Antibody CE 400 was the most reactive, binding to more than 50% of the tumor cells in all reactive specimens. Five of these monoclonal antibodies detected protein antigens in the 80 K to 110 K molecular weight range. Our studies demonstrate the feasibility of producing monoclonal antibodies with selected specificity for cervical carcinoma. These antibodies may be of considerable diagnostic value.     

Nonaspiration-needle smear preparations of pulmonary lesions. A comparison of cytology and histology

Material for cytologic smears was obtained from pulmonary lesions in 146 patients at the Ohio State University between 1979 and 1984 using Rotex or Lee screw needles. Corresponding histologic specimens were available for comparison in 77 of these cases. Diagnoses of malignant neoplasms made by cytologic evaluation (55 cases) were confirmed by the corresponding histologic specimens in 93% of those cases. Possible explanations for the cytologic false-positive diagnoses of malignancy are presented. Correlations between the cytologic and histologic diagnoses of the morphologic type of tumor were 100% for adenocarcinoma, 75% for squamous-cell carcinoma and 20% for large-cell undifferentiated carcinoma. The correlation was 100% for small-cell carcinoma when the histology specimen represented the tumor. Nonneoplastic benign lesions diagnosed cytologically had corresponding benign histologic diagnoses in 94% of the cases. These results compare favorably with those reported for other fine needle aspiration studies of pulmonary lesions. The advantages of using Rotex needles as compared to fine needle aspiration are discussed.