Diagnosis of bladder cancer with urinary cytology, immunocytology and DNA-image-cytometry.

DNA-image-cytometry and antibodies directed against the Lewis X- and the 486p 3/12 antigen were applied to improve diagnostic accuracy of urinary cytology for the detection of bladder cancer. Cytology, immunocytology and DNA-image-cytometry were performed in spontaneously voided urine samples and barbotage bladder washings from 71 patients. The DNA content was determined using the CM-1 Cytometer according to the recommendation of the ESCAP Consensus Report on Standardization of DNA-image-cytometry (1995). For immunocytological examination we used the monoclonal anti Lewis X antibody P-12 and antibody 486p 3/12. All patients underwent subsequent cystoscopy and for any suspicious lesion biopsy or transurethral resection was done. Histological findings revealed 31 patients with transitional cell carcinomas of different stages and grades of malignancy. 40 patients had various benign diseases of the urinary bladder. Cytology yielded a sensitivity of 68% and a specificity of 100%. DNA aneuploidy was detected in 81% of cancer patients with a specificity of 100%. By combination of these two methods the overall sensitivity increased to 87%. Immunocytology with Lewis X and 486p 3/12 antibodies showed reactivity in 84% and 87% in combination with a specificity of 80% and 70%, respectively. By combining urinary cytology, immunocytology and/or DNA-image-cytometry the overall sensitivity increased to 94% with no change in specificity. DNA-image-cytometry should be used to evaluate particularly urothelial cells suspicious for malignancy in urinary specimens. Because of low specificity the monoclonal antibodies against Lewis X- and 486p 3/12 antigens are not helpful in screening for bladder cancer. Nevertheless, their high sensitivity may justify their use in case DNA image cytometry is not available and in the follow up of patients with transitional cell carcinoma.     

Diagnostic accuracy of cytology and immunocytology in carcinomatous effusions

Background: Immunocytology substantially improves the diagnostic accuracy of conventional cytology in the diagnosis of carcinomatous effusions. Due to the unequivocal characterization of the various cell populations, a sensitivity of 92% and specificity of 100% was achieved by immunocytology, examining samples of 1234 serous effusions. Objective: Cytology plays a central role in the aetiological clarification of serous effusions. The sensitivity of this method for the diagnosis of carcinomatous effusions varies between 40% and 80%. The aim of the present study was to investigate whether immunocytology substantially improves the diagnostic quality of the cytological examination in the diagnosis of carcinomatous effusions. Method: Consecutive serous effusions were examined by conventional cytology and by immunocytology. The immunocytological examination was performed on smears, using a standard panel of three antibodies against pancytokeratin, human epithelial antigen 125 and calretinin. Results: Altogether, 1234 effusion samples were examined. A total of 603 effusions were caused by carcinomas, five by malignant mesotheliomas, 11 by malignant lymphomas and 615 by non‐malignant disorders. In conventional cytology, carcinomatous effusions were correctly diagnosed in 314 samples, corresponding to a sensitivity of 52%. In 31 specimens (5%) tumour cells without further specification were described and in 161 samples (27%) the presence of tumour cells was suspected (84% overall sensitivity). A total of 97 carcinomatous effusions (16%) were diagnosed false‐negatively and 50 (8%) of the 615 non‐malignant effusions false‐positively (92% specificity). In immunocytology, 561 carcinomatous samples were correctly diagnosed, representing a sensitivity of 93%. In six cases (1%) the presence of tumour cells was suspected. A total of 36 carcinomatous effusions (6%) were diagnosed false‐negatively (94% over‐all sensitivity). Out of the 615 non‐malignant specimens, there were no false‐positive diagnoses (100% specificity). Conclusion: Immunocytology is a simple, cost‐effective, routinely practicable method which substantially improves the diagnostic accuracy of conventional cytology in the diagnosis of carcinomatous effusions. Therefore, we recommend the use of immunocytology in all those cases where cytology on its own is not completely unequivocal.     

Regional cytology and cytochemistry of the crayfish kidney tubule

Cytological and cytochemical methods were used to identify and characterzie six distinct regions of the crayfish kidney: coelomosac, labyrinth I and II, and nephridial canal I, II, and III. Cells of the coelomosac possess cytoplasm which is strongly PAS-positive, but poor in RNA and protein. Their nuclei possess unusual projections which extend to the basal plasmalemma. Labyrinth I contains columnar cells rich in glycogen. Labyrinth II is characterized by a distended lumen and by shorter cells with high cytoplasmic RNA, many possessing a large intracellular vacuole. A PAS-positive brush border is unique to the two portions of the labyrinth. Cells in the nephridial canal show strong reactions for RNA and Mg⁺⁺-dependent ATPase. In nephridial canal I and II, cells are flattened to cuboidal with the lumen being more distended in nephridial canal I than anywhere else in the tubule. In nephridial canal III, the cells are large and columnar, and the cytoplasmic RNA concentration is greatest apically. Nuclei in all regions of the tubule epithelium, except coelomosac, are large and react strongly for protein. Coelomosac nuclei and those in blood cells are condensed and contain little protein. The cytoplasm of blood cells displays a significant amount of RNA, and traces of polysaccharide material. These observations demonstrate the presence of highly specialized morphological and histochemical alterations along the length of the kidney tubule and indicate sequential modification of urine in the lumen. Evident morphological and cytochemical likenesses between analogous regions of the mammlian nephron and the crayfish kidney tubule suggest that basic physiological similarities may also exist.     

Tissue transfers: substrates for cytology and cytochemistry of animal tissues.

The Tissue Transfer Technique (TTT) is a novel method of sampling animal tissue that can be used to study tissue morphology, chemistry and physiology. This review provides an overview of the technique and demonstrates its use to detect the tissue distribution of specific epitopes, lectin binding sites and nucleic acids as well as its application as an organ monolayer in culture. These applications are compared and contrasted with standard histological techniques including the "Tissue Printing Technique" developed to sample plant tissue.     

Routine immunophenotyping of acute leukaemias

Recent progress in immunophenotyping includes the availability of monoclonal antibodies (MAbs), knowledge of specificity and reactivity patterns of these reagents, and the technical improvements and standardization of immunofluorescence and immunocytology staining procedures, including flow cytometry. These advances have contributed significantly to the establishment of immunophenotyping as an essential diagnostic tool in the differential diagnosis of types of acute leukaemia. Immunophenotyping allows for the objective and reproducible distinction of acute lymphoblastic leukaemia (ALL) from acute myeloblastic leukaemia (AML) and of T-lineage from B-lineage ALL. Immunologically defined ALL and AML subtypes have been found to convey prognostic significance. Using cell lineage-specific and differentiation stage-specific MAbs, cases of T- and B-lineage ALL and of AML can be further classified into a number of different subtypes. Routine immunophenotyping concentrates on the diagnostic enquiry into a few major, clinically relevant subtypes; only a limited number of crucial reagents are employed that are commercially available. The simplification and standardization of discriminatory immunomarker panels make immunophenotyping a reliable diagnostic instrument for the provision of critical data to make a differential diagnosis. An effort to identify the nature and origin of the blast cells precisely, immunological typing definitely plays an important part in the multiple-marker analysis of acute leukaemia (morphology, cytochemistry, karyotyping, genotyping) for applied diagnostic and fundamental research purposes.     

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.     

Cytochemistry of the acute leukaemias

Summary The use of the Romanowsky staining technique, Sudan Black B, the periodic acid-Schiff reaction and methods for revealing peroxidase, acid and alkaline phosphatases and butyrate, acetate or chloroacetate esterases for identifying and discriminating subvarieties of acute leukaemia at the light microscope level is reviewed and the results of their application in a recent study of the first 720 cases admitted to the Medical Research Council's 8th Acute Myeloid Leukaemia trial summarized. The distribution of varieties of acute myeloid leukaemia and the relevance of age and cytochemical findings to clinical prognosis is presented. Identification of the predominant primitive cell—myeloblast, promyelocyte, monoblast, and others—appears to have little prognostic significance. In fact, the presence of periodic acid-Schiff positive erythroblasts is a bad prognostic sign.The association of certain cytochemical findings with the 15;17 translocation and acute promyelocytic leukaemia, especially the patterns of esterase positivity in Auer rods and the Sudan Black, peroxidase, periodic acid-Schiff and esterase findings characteristic of the 8;21 translocation are illustrated. Cytochemical features helpful in distinguishing acute megakaryoblastic leukaemia, notably the periodic acid-Schiff reaction, differential esterase reactivities and 5-nucleotidase, are discussed and illustrated.Brief reference is made to the cytochemical differentiation of lymphoblastic leukaemias. Details of a technical method for the demonstration of 5-nucleotidase are given.     

Ultrastructural cytochemistry of chronic T-cell leukaemias. A study with four acid hydrolases

The ultrastructural localization of four acid hydrolases (acid phosphatase, beta-glucuronidase, beta-glucosaminidase and alpha-naphthylacetate esterase) has been studied in lymphocytes from 16 patients with three types of chronic T-cell leukaemia, namely, T-prolymphocytic leukaemia (T-PLL), T-chronic lymphocytic leukaemia (T-CLL) and adult T-cell lymphoma leukaemia (ATLL). Different patterns of enzyme distribution were observed in the leukaemic T-cells from these disorders. In T-PLL, reactivity for the four acid hydrolases was confined to single or a few large granules. Gall bodies were reactive for beta-glucuronidase, b-glucosaminidase and alpha-naphthylacetate esterase but apparently unreactive for acid phosphatase. In T-CLL, scattered small- to medium-size cytoplasmic granules and many parallel tubular arrays were strongly reactive for acid phosphatase, beta-glucuronidase and beta-glucosidase but showed no reactivity for alpha-naphthylacetate esterase. Intermediate features were observed in ATLL. The observed differences in enzyme reactivity reflect a different content of lysosomal granules in the various types of leukaemic T-cells. They also suggest that similar differences may be found in normal T-lymphocyte subsets.     

The cytology and cytochemistry of the hemocytes of Mytilus edulis and their responses to experimentally injected carbon particles

Hemocytes of Mytilus edulis were examined cytologically and cytochemically. On the basis of structure, staining reactions, and phagocytic behavior, they were divided into two main groups: basophilic hemocytes and eosinophilic granular hemocytes (granulocytes). The basophilic cells were further divided into small lymphocytes and larger phagocytic macrophages reactive for lysosomal hydrolases. Mitosis was observed in granulocytes and in small lymphoid cells, believed to be the stem cells for the basophilic cell line. A few cells appeared to be intermediate between lymphocytes and small granulocytes. Macrophages were the main cell type involved in the clearance of injected carbon particles. However, granulocytes did show some phagocytic activity. Brown cells displaying apparent amoebocytic behavior were found to contain Fe³⁺ and Pb²⁺ in cytoplasmic inclusions, some of which were also reactive for β-glucuronidase and glucosaminidase. These cells appear to have a separate origin from the hemocytes.