We developed a rapid image sensor analytical system (RISAS) with a linear type charge-coupled device (CCD). We performed a discriminating standardization by Mahalanobis' generalized distance using a method of principal component analysis (multivariate analysis) on the basis of cells diagnosed by more than one cytopathologist as class III b (four cases, 54 cells) and class IV (three cases, 37 cells) in uterine cervical smears. We examined the system for quality control in cytodiagnosis by registering atypical cells (class III b, 14 cases, 81 cells; IV, 11 cases, 33 cells) diagnosed by cytoscreeners. Almost all cells diagnosed by the cytoscreeners were plotted into their respective class area made as a discriminated standard. It is suggested, therefore, that an image analytical system like ours is useful for quality control in cytodiagnosis.

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The purpose of this study is to demonstrate the cytological and histological localization of antigen CA 15-3 by using novel monoclonal antibodies, 115 D 8 and DF 3 in the proliferative epithelial cells of benign and malignant human breast lesions.
The preparations of a total of 12 cases which included the cytological and histological specimens consisted of 5 cases of benign lesions and 7 cases of breast cancers.
For a comparative study, tissue specimens of 19 cases from extramammary lesions (stomach, colon, liver, pancreas, lung, etc) and cytologic imprinted smears of 10 cases from extramammary lesions were added to this study.
Immunoperoxidase staining of cytological specimens and paraffin embedded tissue sections were stained with 115 D 8, DF 3, and respective antibodies of epithelial membrane antigen (EMA).
The results were as follows:
1) The antigens of 115 D 8, DF 3 (MAM-6, DF 3-antigen) and EMA were found in all cases of breast lesions, but these antigens were localized in the apical surfaces of the benign breast lesions. In malignant breast lesions, these three antigens were localized in the cytoplasm of the carcinoma cells. These antigens may serve as useful markers in the separation of each benign breast lesion and breast carcinomas because of the localization pattern in these proliferative epithelial cells. The same conclusion may be applied to the tissue specimens.
2) In extramammary lesions: MAM-6, DF 3-antigen, EMA were considered inappropriate as helpful diagnostic markers, because of low positive rate and the absence of clear difference in localization between benign and malignant lesions on both cytologic and tissue specimens.

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Quantitative microspectrophotometry was performed in female breast cancer cases by dual parameter analysis of cellular nuclear protein and nuclear DNA contents using the Feulgen-naphthol yellow staining method. Tumors were studied by means of cell suspensions made from paraffin embedded blocks in 24 cases and imprint smears in 41 cases. The DNA histograms were classified into 4 types: type 1 showed a 2 C pattern, type 2 showed 2 C·4 C patterns, type 3 showed a 2 C·4 C serial pattern and type 4 showed aneuploid pattern. The results were follows.
In the cases that died within 42 months, the proportion of types 3 and 4 was high and percentage of cells with over 2.0 nuclear protein to nuclear DNA ratio (P/C>2.0 cell percentage) was significantly higher (p<0.05). Even in long survivor many patients who showed a high P/C>2.0 cell percentage had tumor recurrence over 6 years after therapy.
In stage 3 cases the proportion of type 4 was high. In contrast, in type 1 cases the proportion of stage 1 cases was high.
There was no significant difference in the histological type. In estrogen receptor positive cases the proportion of type 1 and 2 was high (76.2%), and in negative cases the proportion of type 3 and 4 was high.(80.0%)
In premenopausal cases the proportion of types 1 and 3 was high, but in postmenopausal cases the proportion of type 4 was high. P/C>2.25 cell percentage was significantly higher in postmenopausal cases (p<0.05).
Investigation of the DNA histogram and P/C>2.25 (2.0) cell percentage allows estimation of the character and prognosis of breast cancer cases.

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In our first paper histochemical and scanning electron microscopic features of human mesothelial cells in vitro were discussed. This paper is concerned with the temporal change of lineage-directed antibodies, including those to keratins, vimentin, epithelial membrane antigen (EMA) and carcinoembryonic antigen (CEA) in cultured mesothelial cells.
1. In the early stages of culture the prominentlyproliferating cells stained for low (40, 45 kdaltons) molecular weight keratins said to be the main mesothelial proteins, but were negative for high (65 kdaltons) molecular weight keratin and EMA. After several weeks in reaching confluence, both types of keratin were expressed together with EMA by many cultured cells.
2. From the beginning of culture antisera to vimentin always reacted with the proliferating cells.
3. Throughout culture, none of the cultured cells showed immunoreactivity for CEA.
Our results show that cultured mesothelial cells expressed non-epithelial markers (vimentin) in the early stages of culture. On the other hand, in the course of culture, coexpression of non-epithelial and epithelial (EMA and high molecular weight keratins) markers was observed similar to the morphologically bi-phagic pattern, reported in the previous paper.

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Sixty-two cases of hepatocellular carcinoma examined cytologically (29 fine-needle aspirates, 28 touch preparations of biopsy or resected specimens, and 5 aspirates or touch preparations of autopsy specimens) were subclassified into four grades (GI-IV) using Edmondson's classification modified for cytodiagnosis and according to several specific types (clear cell, pleomorphic and spindle cell types). In this report cytomorphological characteristics of each grade and specific types of HCC were described.
In G-I HCC, tumor cells were more cohesive than in normal hepatocytes, and were present in sheet or trabecular patterns. They were large, polygonal and uniform with abundant granular eosinophilic cytoplasm. Nuclei with slightly enlarged nucleoli were round and centrally located, and the N/C ratio was slightly higher than that of normal hepatocytes. Cytologic specimens of G-II HCC contained a large number of tumor cells which were present singly or in large clusters. They had most typical features of HCC showing both evdience of hepatocytic origin and of distinct malignancy, that is, granular eosinophilic cytoplasm, enlarged nuclei with prominent nucleoli, and a high N/C ratio. In GIII HCC, evidence of hepatocytic origin was decreased, although malignancy was obvious. Tumor cells were pleomorphic, less cohesive and present singly or in small clusters. The most characteristic feature of this grade was the presence of mono or multinucleated giant cells with abundant cytoplasm and large irregularly shaped hyperchromatic nuclei containing a few prominent nucleolei. In G-IV HCC, evidence of hepatocytic origin was completely absent and tumor cells were cohesive, relatively small and uniform in size and shape. They had scanty cytoplasm and round hyperchromatic nuclei with indistinct nucleoli. The N/C ratio was the highest of the four grades.
Clear cells of specific type contained glycogen and/or lipid, and showed the same nuclear appearance as that of GI or G-II HCC. They resembled tumor cells of renal cell carcinoma in appearance. In the pleomorphic type, although bizarre giant cells were observed, they were much more numerous and larger than those of G-III HCC. Tumor cells of the spindle cell type were large, and present in small clusters. They had vesicular cytoplasm with distinct cell borders and nuclei with indistinct nucleoli, resembling sarcomatous cells.

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Cytological findings of urine in intravesical Bacillus-Calmette-Guerin (BCG) in the treatment of bladder tumors were studied. The cytological findings in eight cases of superficial bladder tumor, involving six men and two women, were as follows:
1. Before BCG treatment, there was no difference between cases in which BCG was later effective and those in which it was ineffective in morphological changes, cancer cell appearance and background.
2. In BCG treatment, it seemed difficult to presume the prognosis because there were no characteristic findings concerninig cancer cells.
3. The reference for background findings such as increasing numbers of neutrophils, multinuclear giant cells and epitheloid cells was useful in presuming the prognosis in BCG-effective cases.

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We developed a modification of papanicolaou's staining which permits staining more quickly. Proceding:
1) Nucleus staining solution was prepared by Gill's method.
2) 0.5% HCl and 0.9% NaCl were used for differential staining.
3) A staining carrier tray set was devised so as to omit washing on running water.
4) One solution was used, instead of two solution, to stain cytoplasma by the modification 267 of Papanicolaou staining.
Result:
The results of staining by the present modified method was conparable to actual Papanicolaou staining in terms of fixation, clearness and unfadingness.

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Optimum times of fixation using Buan's solvent and sequential stock into 70% ethanol for lectin immuno-staining were studied on monolayer cultured cells. HRA cells derived from human ovarian serous cyst adenocarcinoma were used in the present study. HRA cells were incubated as monolayer culture into the Lab-Tek chambers. Lectin immuno-staining was performed using biotinized ConA, SBA and PNA lectin by Avidin-Biotin peroxidase complex method. In comparison with 0.5, 1, 3, 6, 24, 48 hours, 7 and 14 days as fixation time of Buan's solvent, 24 hours treatment produced the finest stainings. Next, sequential treatment of 70% ethanol after Buan's fixation for 24 hours were tested at the periods of 2, 48, 72 hours, 7 and 14 days. Fourty eight hours produced the most apparent stainings for identifying the location binded by lectin. In addition, stock into 70% ethanol over 7 days feagibled stainings. In conclusion, the optimum times of Buan's fixation and sequential stock into 70% ethanol were 24 hours and 2 to 7 days, respectively.

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We report a rare case of the cervical carcinoid tumor with squamous cell carcinoma in situ developed in 34 year old female. Routine examination of vaginal smear and cervical biopsy revealed monotonous small neoplastic cell growth in which N/C ratio was considerably high. Thus, the lesion was originally diagnosed as squamous cell carcinoma of non-keratinizing small cell type. Then, she had admitted to the hospital of Shiga University of Medical Sciense for the surgery. Post operative histological examination revealed cords or ribons of small polygonal neoplastic cells in the subepithelial layer. Nuclei were relatively large, but not so pleomorphic. The tumor was identified as carcinoid since the neoplastic cells were argyrophilic and neuro-secretory granule were demonstrated by EM observation. In the vicinity, squamous cell carcinoma in situ was also detected and final diagnosis of double cancer was done. Thereafter, review of the preoperative cytological smear was carried out and most of the neoplastic cells were identified as carcinoid cells, since their chromatin textures were not so coarse as in squamous cell carcinoma and nuclei were not so large. From our experience, it is concluded that precise observation of intranuclear structure seems to be important for the differential diagnosis of carcinoid and small cell carcinoma.
For the confirmation of the diagnosis of carcinoid, immunohistochemistry study of neuroendocrine markers such as NSE and Leu 7 were quite useful.

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A relatively rare case of papillary hidradenoma of the vulva is reported mainly with regards to cytological and ultrastructual findings.
Touch smears of the tumor were composed of two cell types. One type showed clusters arranged in a sheet-like pattern and the other showed slight aggregation of cells with a glandular structure. The individual cells showed slight atypia with mild anisokaryosis and small nucleoli, but did not show any findings of large nuclei, prominent nucleoli or nuclear overlapping. Therefore, we suggest that these findings are potentially useful for the differential diagnosis between papillary hidradenoma and adenocarcinoma.
Ultrastructural findings showed segregation into the lumen by decapitation and myoepithelial cells were also contained. It is suggested that these findings might indicate a differentiating process of the apocrine glands.

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The cytological findings of adenoma of the nipple is described.
An 11-year-old girl was admitted because of a tumor of the areolar region of the right breast. There was no nipple discharge or pain. An oval mass was palpable, and the skin surface was intact.
A sample aspirated from the tumor contained a large number of epithelial cells. These cells were mainly arranged in tight clusters or sheets, and papillary structures. The epithelial cells had round or oval nuclei of equal sizes, with a regular and finely granular chromatin and small inconspicuous nucleoli. A small number of dissociated cells had a myoepithelial appearance, with naked nuclei. Moderate amounts of secretory material and few histiocytes were also present. The cellular aggregates with a papillary structure consisted of small cells with hyperchromatic and irregular shaped nuclei.
The tumor was removed. It was solid and diagnosed an adenoma of the nipple, showing florid proliferation of ductal epithelial cells. Histologically, no sign of malignancy was found in the resected tumor.

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A case of malignant lymphoma of the breast diagnosed by an immunocytochemical study of a specimen for aspiration cytology is presented here, and its cytomorphology, immunocytochemical findings, gross and histopathologic appearances of resected tumor, immunohistochemical and ultrastructural features are also described, and the differences of immunohistochemical features between lymphoma and carcinomas of the breast are discussed. The patient was a 44 year-old woman, and she was admitted to the hospital for a breast tumor. Since there were many lymphoid cells of nuclear abnormalities in the aspiration smear, malignant lymphoma was strongly suspected. Then, immunocytochemistry using antibody against leukocyte common antigen (LCA) was applied on the smear slides after decoloration of Papanicolaou stain with 0.5% HCl-alcohol, and the diagnosis of malignant lymphoma was made because LCA was detected on the cell surface of aspirated cells. Histopathologic diagnosis of the resected tumor was non-Hodgkin's lymphoma, diffuse, large cell (LSG classification). The presented tumor was clearly differentiated from carcinomas on immunohistochemical findings such as positive stain with antibodies against lymphocyte related antigens (LCA, MB-1, MT-1) and no expression of cytokeratin or epithelial membrane antigen (EMA). The immunocytochemistry is thought to be a useful method for diagnosisof malignant lymphoma of the breast.

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We report a case of acute myelomonocytic leukemia (AMMoL) with infiltration of leukemia cells in uterine cervix diagnosed by smear examination.
The patient is a 72 year old woman who was diagnosed as acute myelomonocytic leukemia by hematological and chromosomal examination. She accepted chemotherapy, and had complete remission after one and a half year treatment.
During in remission, she had genital bleeding. Cytological examination of uterine cervix and endometrium was performed. Numerous isolated mononucleated cells which had increased N/C ratio, irregular shaped nuclei and enlarged nucleolei were found.
Abnormal cytological findings of the smear revealed that they were leukemia cells. The diagnosis of myelomonocytic leukemia was confirmed by esterase double staining (α-naphthyl butylate and naphthol AS-D chloroacetate) that demonstrated activity of esterase in the cytoplasm of the neoplastic cells.
Infiltration of leukemia cells in gynecological organs israrely reported. Routine cytological examination of vaginalsmear in female patients with leukemia should be recommended.

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There have been few reports referring to cytologic characteristics of clear cell sarcoma (CCSa) because of its rarity. Moreover, the exact cytogenesis of CCSa is still controversial, although a melanocytic origin seems to be widelyaccepted at present. In the present study, the cytologic and immunocytologic findings of tumor cells detected in sputum and pleural effusion in a case of CCSa were described, together with histological and immunohistological observations of the original tumor tissue.
A 39-year-old male noted a small finger tip-sized nodule in the right popliteal fossa, and a tumor was extirpated eleven months later. The elastic firm tumor, approximately three centimeters in diameter, arose in the aponeurosis of thelateral head of the right gastrocnemius muscle. In the cutsurface, the tumor was gray-white in color and solid, showing a lobulated pattern with a fairly distinct border. No necrotic or hemorrhagic foci were found. Histologically, a diagnosis of CCSa was made. Four years later, multiple pulmonary metastatic foci developed, and a number of large round epithelioid neoplastic cells were detected in sputum and pleural effusion, either sparcely or in small clusters. The cells possessed abundant, rather clear cytoplasm without obvious melanin granules. Characteristically, the vesicular nuclei contained one or two very conspicuous nucleolei located centrally. The tumor cells from pleural effusion were strongly positive for PAS reaction and immunostainings revealed strong reactivities with anti-S-100 protein and antimelanoma specific HMB-45 antibodies, as did the tumor cells in the original tumor. The findings of tumor cells in this case was, therefore, similar to those of amelanotic malignant melanoma, and support the melanocytic origin of CCSa.

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Cytologic features of insulinoma and their histopathologic background observed in 6 cases, including a malignant case, were reported. Though cellular morphologies were not always similar among cases each other, the most characteristic features were found in the coagulation form of the chromatin like as grains of sand. This feature seemed to be analogous to a mixture with Salt and Pepper, that has been reported by Hsiu et al. In cases associated with a few anisonucleosis and anisocytosis epithelial arrangements, including rosette formation and indian file arrangement, have been often seen, while cellular conjunction were relatively loose in cases with anisonucleosis and anisocytosis. In all cases, cellular features observed was not variables in paralleld with histpathologic types and the abilities of hormon productions such as insulin, glucagon, somatostatin detected immunohistochemically.

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A 62-year-old woman was admitted because of an upper abdominal mass. Computed tomography showed a multicystic tumor in the body of the pancreas. Histological examination of the excised tumor revealed serous cystadenoma.
Analysis of cytologic smears showed tumor cell clustering like sheets and sometimes glands. The cuboidal cells had clear cytoplasm. The nuclei of the tumor cells were small, round to oval, and uniform. The chromatin structure was finely granular, with one to two small nucleoli. Periodic acid-Schiff (PAS) staining suggested intracytoplasmic glycogen, which was confirmed by diastase digestion. Mucin was not detected in the tumor cells
Ultrastructurally, the cytoplasm of the tumor cells contained numerous glycogen granules and scanty organelles. Immunohistochemically, epithelial membrane antigen, MAM 6 antigen and CA 19-9 were present on the luminal surface of the tumor cells, and keratin, α₁-antitrypsin and α₁-antichymotrypsin in the cytoplasm.

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We reported cytologic and histologic findings in six cases of mediastinal cystic lesions, which included four congenital cysts and two lymphangiomas.
In an aspirate of the thymic cyst, few clusters of thymic epithelial cells were found in a background of lymphocytes. Ciliated columnar epithelium in the bronchial cyst and mesothelium in the pericardial cyst were diagnostic. In the two cases of lymphangioma, many small lymphocytes and a few foamy macrophages were aspirated.
In diagnosis of mediastinal cystic lesions, it was important to macroscopically observe fluid from the cyst. Almost all mediastinal cystic lesions were benign. As the aspirated fluid sometimes contained only a small number of cells, thespecimen had to be carefully studied.

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