Cardiac tamponade as a presentation of malignant thymoma

Morphologic, cytochemical and immunocytochemical studies of pericardial fluid from a 30-year-old man presenting with cardiac tamponade are described. Based on the results of the immunocytochemical studies and the histologic examination of excised pericardium, a diagnosis of malignant thymoma was made. This is the first documented case in which malignant cells were found in the pericardial effusion in a patient with invasive thymoma. The significance of using a multidisciplinary approach to the study of body fluids is discussed.     

Granulocytic sarcoma with pleural involvement. Identification of neoplastic cells with cytochemistry

Malignant cells were detected in the pleural effusion of a patient with three separate primary malignancies. These cells were judged by conventional morphologic studies to be poorly differentiated cells, but cytochemical studies showed them to be granulocytic precursor cells. The use of cytochemical or immunochemical techniques may be most practical for the cytodiagnosis of malignant cells in serous effusions.     

Blast transformation in chronic myeloid leukemia with synovial involvement

In a patient with chronic myeloid leukemia in blastic transformation, the blast cells in a knee joint effusion were identical morphologically to those in the marrow. Cytochemical and immunocytochemical studies showed that these blasts were primitive erythroblasts and not granulocytic or monocytic precursors. The combined use of cytologic, cytochemical and immunocytochemical techniques will help to accurately identify cells in joint effusions and to elucidate the mechanism of synovial involvement by hematologic disorders.     

Immunocytochemical diagnosis of lymphoma in serous effusions

An immunoalkaline phosphatase technique was used to examine the lymphoid cells in serous effusions from five patients with malignant lymphoma. The results were interpreted along with the morphologic studies and retrospective assessments of the clinical conditions of the patients. Two patients had no involvement of the serous cavities, and two had proven involvement. The fifth patient was studied while his lymphoma was evolving from inapparent to disseminated disease. In the two patients without involvement of the serous cavities, the effusion lymphocytes were predominantly monoclonal T cells, comparable to those in six patients with diseases other than lymphoma. In those with involvement of the serous cavities, the effusion lymphocytes were predominantly monoclonal B cells. In the patient with lymphoma in evolution, immunocytochemical studies accurately reflected the progression of disease. We conclude that immunocytochemical studies of the lymphocytes in serous effusions help not only to differentiate reactive from neoplastic lymphoproliferation but also to assess the status of lymphomatous involvement of the serous cavities. The immunocytochemical studies are most effective when correlated with clinical and cytologic studies.     

Cytochemical examination of acid phosphatase and beta-glucuronidase enzymes in low-grade B cell non-Hodgkin's lymphomas

The differential diagnostic significance of acid phosphatase and beta-glucuronidase were studied in 77 cases of low-grade B cell non-Hodgkin's lymphomas. In most cases the results of cytochemical enzyme studies performed on malignant cells of the bone marrow were evaluated. B cell chronic lymphocytic leukaemia, centrocytic and centroblastic/centrocytic lymphomas were characterized by a weak or a negative acid phosphatase and beta-glucuronidase activity. Stronger positivity was observed in immunocytoma and in Waldenstr?m's macroglobulinaemia, while the highest activity was found in multiple myeloma. Hairy cell leukaemia of B cell origin showed intensive tartrate-resistant acid phosphatase activity. The cytochemical examination of these lysosomal enzymes may be useful in the diagnosis of low-grade malignant lymphomas of B cell origin by completing other methods.     

Morphological, cytochemical and ultrastructural studies of a case of systemic mastocytosis

Morphological, cytochemical and ultrastructural studies of mast cells were carried out in a patient affected with systemic mastocytosis. Neoplastic mast cells showed morphological features between classic tissue mast cells and circulating basophils. They showed strong granule metachromasia after toluidine blue, faint positivity to Hotchkiss reaction, strong positivity to chloroacetate esterase, while they were negative to alkaline phosphatase. Ultrastructural observations showed heterogeneity of granules, most of which had homogenous fine dotted contents. The origin and linkage between mast cells and circulating basophils are discussed.     

Ancillary alpha-methylacyl-CoA racemase immunocytochemistry in the diagnosis of adenocarcinoma of the prostate in urinary cytology: a case report

BACKGROUND: Alpha-methylacyl-coA racemase (AMACR) was recently shown to be a sensitive immunohistochemical marker for substantiating a diagnosis of adenocarcinoma of the prostate. Its applicability to exfoliative urinary cytology has not been investigated before. CASE: The patient was a 77-year-old male with a history of persistent intermittent hematuria who was evaluated with urine cytology. Cytopathologic studies were interpreted to demonstrate "atypical urothelial cells." The patient was evaluated with computed tomography, which demonstrated new left hydroureter and hydronephrosis. He underwent cystoscopy, which showed an abnormal left hemitrigone with the left ureteral orifice obscured by an infiltrative mass. This area was biopsied, and histologic studies of the tissue chips demonstrated underlying prostatic adenocarcinoma directly invading the urothelium. We used AMACR immunoreactivity on a retrospectively studied, catheterized urine slide to confirm the diagnosis. CONCLUSION: This case suggests that combined cytomorphology and immunocytochemisty for AMACR may allow an accurate identification of cells of prostatic adenocarcinoma when cytomorphologic studies or the clinical history raises the differential diagnosis of prostate cancer presenting with exfoliation of malignant cells into the urine.     

Urinary cytology in renal transplant recipients with stable graft functions

The utility of routine urinary cytology in renal transplant recipients was investigated. Slides of 79 urine samples obtained from 59 renal transplant patients shortly after transplantation and of 275 urine sediments from 126 patients who had received a transplant before 1978 were screened for abnormal urothelial cells. None of the samples taken within one year of transplantation contained malignant cells. For five patients transplanted before 1978, repeated cytologic examinations showed malignant cells, but neither urologic examination nor clinical nor postmortem follow-up studies revealed a tumor. In all five cases, the abnormal cells disappeared from repeat samples within two to three months. None of the other 121 patients, with repeatedly normal urinary cytologies, exhibited a urinary tract carcinoma in the 24-month follow-up period. It would appear that the cytologic findings in the urine of renal transplant patients who have received long-term immunosuppressive medication may be transiently abnormal.     

Immunophenotyping of acute non-lymphoblastic leukaemias

Using monoclonal antibodies specific for myelomonocytic cells, 40 non-lymphoblastic leukaemias were analysed applying immunostaining to cytospin preparations. Based on the reactivity patterns six groups of acute non-lymphoblastic leukaemias could be determined, mirroring the bimodal differentiation pathway of myelomonocytic cells. Comparative enzyme cytochemical analysis did not render a clear cut correlation and discrimination of the immunocytochemically defined groups. It is concluded that only the application of a broad panel of immunocytochemical and enzyme cytochemical methods allow a sound subdivision and diagnosis of acute non-lymphoblastic leukaemias.     

Immunocytochemical localization of polyamines in normal and neoplastic cells. Comparisons to the formaldehyde-fluorescamine and o-phthalaldehyde methods

Summary Polyamines are low molecular weight organic cations, necessary for cell proliferation and implicated in numerous biochemical events. Their light microscopical distribution has previously been studied by the use of two fluorescence cytochemical methods. With the aid of an antibody recognizing the two main polyamines, spermidine and spermine, we now report on their immunocytochemical localization in animal tissues. Polyamine immunocytochemistry was found to require very well controlled conditions of fixation in order to prevent diffusion, loss and redistribution of endogenous polyamines. Moreover, in certain cellular compartments, polyamine immunoreactivity was masked by proteins, necessitating proteolytic pretreatment of sections prior to staining. The fluorescence cytochemical methods, employing low molecular weight reagents, did not require such unmasking. The results of the optimized immunocytochemical procedure were in complete agreement with the results obtained by the fluorescence cytochemical methods. Although fluorescence cytochemistry, is simpler and quicker to perform than immunocytochemistry, the latter technique may be extended to studies of polyamines at the ultrastructural level.     

Distribution patterns of ornithine decarboxylase in cells and tissues: facts, problems, and postulates.

Ornithine decarboxylase (ODC) is a key enzyme in polyamine biosynthesis. Increased polyamine levels are required for growth, differentiation, and transformation of cells. In situ detection of ODC in cells and tissues has been performed with biochemical, enzyme cytochemical, immunocytochemical, and in situ hybridization techniques. Different localization patterns at the cellular level have been described, depending on the type of cells or tissues studied. These patterns varied from exclusively cytoplasmic to both cytoplasmic and nuclear. These discrepancies can be partially explained by the (lack of) sensitivity and/or specificity of the methods used, but it is more likely that (sub)cellular localization of ODC is cell type-specific and/or depends on the physiological status (growth, differentiation, malignant transformation, apoptosis) of cells. Intracellular translocation of ODC may be a prerequisite for its regulation and function.     

Cytochemical characterization of pituitary target cells for biotinylated gonadotropin releasing hormone

These studies describe the application of new cytochemical stains that co-localize a biotin-labeled gonadotropin releasing hormone (GnRH) analog and FSH or LH in the same field or cell. Pituitary monolayer cells were stimulated with the [D-Lys6] GnRH analog or the same analog labeled with biotin. Biotinylated [D-Lys6] GnRH exhibited a higher affinity and was 7-10 X more potent than unlabeled [D-Lys6] GnRH. The avidin-biotin peroxidase complex technique (ABC) was applied to localize the biotinylated GnRH on the cells with the use of a dense black peroxidase substrate. Specificity tests showed that the stain could be eliminated by competition with unlabeled [D-Lys6] GnRH. The GnRH stain was followed by immunocytochemical stains for LH beta, FSH beta or 25-39ACTH with a different peroxidase substrate (amber or orange-red). Stain for GnRH was found on the surfaces of 16% of the cells and 60-90% of the GnRH stained cells also stained for one of the gonadotropins. Most (90-100%) of the gonadotropes showed stain for GnRH. Our studies demonstrate that a potent biotinylated GnRH analog binds cells that can be identified specifically as gonadotropes.     

Cell surface distribution and intracellular fate of human beta-very low density lipoprotein in cultured peritoneal mouse macrophages: a cytochemical and immunocytochemical study

The receptor-mediated endocytosis pathway of colloidal gold labeled beta-very low density lipoprotein (beta-VLDL-Au) derived from patients with familial dysbetalipoproteinemia was analyzed at the ultrastructural level in macrophages. The results showed that beta-VLDL-Au complexes were specifically recognized by a cell surface receptor of the macrophages. beta-VLDL-Au particles once bound to the randomly distributed cell surface receptors clustered in coated pits and were taken up by coated vesicles. Subsequently, the beta-VLDL-Au particles passed through tubular structures and small endosomes before deposited into large electron lucent smooth surfaced endosomes. As revealed by ruthenium red and enzyme cytochemistry the endosomes appeared to be separated from the extracellular space and did not contain acid phosphatase. There were no clear signs of passage of beta-VLDL through the Golgi complex. The accumulation of many flocculated gold particles within Ac-Pase positive vesicles suggests that beta-VLDL once internalized by the macrophages is diverted into a degradative pathway. Incubation of beta-VLDL-loaded macrophages with the hydrophobic fluorescent dye nile red revealed numerous large fluorescent bodies within the cells indicating that the macrophages accumulate large amounts of lipid droplets with time. Additional studies large amounts of lipid droplets with time. Additional studies with native beta-VLDL in conjunction with postembedding immunocytochemical techniques were used to delineate further the intracellular pathway. Immunolabeling was carried out on thin sections of LR White embedded cells using affinity-purified polyclonal rabbit antibodies against apolipoprotein B with the protein A-gold or goat anti-rabbit IgG-gold technique. Indirect visualization of beta-VLDL by these immunocytochemical studies yielded results comparable to those with gold-labeled beta-VLDL. On the basis of both indirect immunocytochemical and direct cytochemical localization of beta-VLDL it is concluded that although colloidal gold labeling of beta-VLDL molecules unquestionably modifies their morphology, their function appears to be unaltered, at least with respect to the process of receptor-mediated endocytosis.     

Preparatory methods for DNA hydrolysis, cytochemistry, immunocytochemistry and ploidy analysis. Their application to automated and routine diagnostic cytopathology

A review is presented of some methods used to prepare cytologic specimens for analytical and/or automated studies, with the steps of the procedures detailed in appendices. The preparation of the cell monolayers required for optimal automated cell image analysis and classification, e.g., by the Cytoscan 110, is discussed, as is the preparation of poly-L-lysine-coated slides used in the production of monolayered specimens. These monolayers, which can be prepared from a variety of specimens, are also useful for cytochemical and immunocytochemical studies and DNA ploidy analysis. For DNA analysis, a modified gallocyanin chrome alum staining procedure is described as a stoichiometric alternative to the time-consuming Feulgen reaction. The hydrolysis technique required by the latter method is also detailed. The freeze-fracturing technique for the enhancement of monoclonal antibody immunocytochemical staining of detectable antigens is described, along with an indirect immunoalkaline phosphatase staining method. The use of enzyme cytochemical reactions for glucose 6 phosphate dehydrogenase and lysosomal naphthylamidase is also presented.     

Fine needle aspiration cytology of malignant melanoma of soft parts. Report of two cases

Two cases of malignant melanoma of soft parts were diagnosed by fine needle aspiration (FNA) biopsy. One case involved the right thumb of a 22-year-old woman; the other involved a submental mass in a 36-year-old man who had been previously diagnosed as having malignant melanoma of soft parts. The FNA smears showed discohesive polygonal or oval and fusiform cells with eccentric nuclei, irregular nuclear chromatin and prominent nucleoli. FNA cell block sections contained polygonal or fusiform cells with clear cytoplasm, eccentrically placed nuclei and prominent nucleoli. Ultrastructural and immunocytochemical studies on aspirated material supported the diagnosis of malignant melanoma.     

Microphotometric quantitation of the reaction product of several indirect immunoperoxidase methods demonstrating monoclonal antibody binding to antigens immobilized on nitrocellulose

The nitrocellulose model and microphotometry were used to investigate whether in immunoperoxidase cytochemical methods the amount of final reaction product reflects the amount of cell surface antigen. The results obtained with four cytochemical peroxidase methods, i.e., those using diaminobenzidine/H2O2 (DAB/H2O)2, DAB/H2O2/COCl2, DAB/H2O2/imidazole, and silver intensification of the DAB end product, were compared first. The quantitative DAB/H2O2/imidazole method proved to be the most sensitive and was selected for further studies. Cell surface antigens prepared by solubilization of peritoneal macrophages with octyl-beta-D-glucopyranoside were immobilized on nitrocellulose. Monoclonal antibody binding to these cell antigens was detected by peroxidase immunocytochemistry. Comparison of the sensitivity of the indirect immunoperoxidase and the biotin-(strept)avidin immunoperoxidase methods on the basis of the highest detectable dilution of a cell lysate showed that these methods were equally sensitive. A linear relationship between the absorbance of the peroxidase reaction product and the amount of cell lysate immobilized on nitrocellulose was found for all three indirect immunoperoxidase methods. This proves that the amount of final immunocytochemical peroxidase reaction product is proportional to the amount of antigen in cell lysates. However, the relative expression of antigens in intact cells differs from that in cell lysates. Therefore, the present method to solubilize cells and immobilize cell antigens cannot be used to quantitate the antigen content of cells.     

Cytology of angiosarcoma in effusions

The cytologic and immunocytochemical findings in pleural effusions from three cases of angiosarcoma are presented. In two of the cases, the primary lesion was on the scalp; in the third case, an angiosarcoma of the small intestine developed after radiotherapy for Hodgkin's disease. Single malignant cells and small clusters of cells were seen in cytologic preparations from two cases while only single cells were seen in preparations from one case. The malignant cells had delicate, finely vacuolated cytoplasm with distinct borders. No specific morphologic features were noted. Immunoperoxidase studies revealed binding of Ulex europaeus and reactivity for vimentin in all three cases and expression of Factor VIII-related protein in two of the cases but no expression of epithelial markers. The clinical history and immunoperoxidase studies are necessary to distinguish angiosarcoma from metastatic adenocarcinoma and other malignancies in effusions.     

Demonstration of acetylcholinesterase molecular forms in a continuous tubular lysosomal system of rat pancreatic acinar cells.

By applying the highly sensitive cytochemical Gautron's technique, we were able to reveal AChE activity in rat pancreatic acinar cells, particularly at the level of a complex membrane-bound network formed by tubules with varicosities located around the nuclei and close to the basolateral membrane. The Golgi apparatus was devoid of cytochemical reaction beside the trans-Golgi network cisternae, which showed a positive reaction. The RER of some acinar cells also presented a signal, demonstrating their capability of synthesizing AChE. Immunogold using a specific anti-AChE antibody yielded similar results. Double-labeling experiments corroborated the presence of enzyme cytochemical and immunocytochemical signals in the same lysosomal tubular network. Biochemical sedimentation assays confirmed the presence of AChE in acinar cells, which exists as two globular molecular forms, G(1) and G(4). These results were obtained with pancreatic tissue in situ as well as with isolated acinar cells maintained in culture and devoid of neural elements. The existence of a continuous tubular lysosomal network containing AChE is in agreement with previous reports on acinar and other cell types, and supports a more general hypothesis on dynamic continuities among cell structures. Whether AChE is being secreted by the acinar cells or internalized through this endo-lysosomal system was not defined. However, the capability of the acinar cells to synthesize AChE and to channel it through a tubular system is a good indication that the cells can modulate their cholinergic stimulation for optimal secretion of digestive enzymes.     

粒单系造血祖细胞(CFU-GM)琼脂半固体培养的原位固定及鉴定染色

用粒单系造血祖细胞（CFU－GM）体外琼脂半固体培养法,琼脂块原位固定,Wright染色与细胞化学和免疫细胞化学技术,对培养人的骨髓CFU－GM进行集落性质鉴定,结果表明：Wright染色能区别集落的性质及细胞的分化程度：CFU－GM的细胞中PAS、POX呈阳性反应;CD14、CD15分别在单核细胞和粒细胞的胞浆或胞膜上表达。该方法对鉴定细胞集落类型、集落中细胞的分化与成熟程度是简便可靠的手段。     

Urine cytology of localized primary amyloidosis of the ureter: a case report

BACKGROUND: Morphologic findings of amyloid in urine cytology material have rarely been reported because amyloidosis of the urinary tract is a relatively uncommon disorder. We present a case of primary amyloidosis of the ureter, including catheterized urine cytologicfindings. CASE: A 78-year-old man had pollakiuria and dysuria for 5 years before admission after transurethral resection of the prostate. Clinical examination revealed left hydronephrosis and stricture of the lower part of the left ureter, and a malignant ureteral tumor was suspected clinically. In catheterized urine cytology, many clusters of epithelial cells, inflammatory cells and abundant, amorphous, waxy material were observed. The amorphous material stained light green by the Papanicolaou method and positive with direct fast scarlet (DFS), showing yellow-green birefringence under polarized light. Positivity with DFS staining was not affected by treatment with potassium permanganate. Immunocytochemically the material was AL-type amyloid protein. Atypia were absent from epithelial cells. The patient had no history of diseases that could cause secondary amyloidosis. The present case was considered to be primary amyloidosis localized to the left ureter because no particular morphologic change in the epithelial cells of the urinary tract was observed. CONCLUSION: Amyloid can be present in urine and should not be overlooked or confused with tumor diathesis when a malignant tumor is suspected clinically.