bluuebunny/crossref_metadata_2025_split huggingface dataset:
Carcinoma in situ specimen classification based on intermediate cell measurements
Authors: Katzko Michael W. , Pahlplatz Martin M. M. , Oud Peter S. , Vooijs G. Peter 
Published: August 2005

AbstractIn this study the possibility of classifying carcinoma in situ and normal specimens by measuring normal‐appearing intermediate cells was explored. Twenty‐five histologically verified carcinoma in situ specimens and 99 normal specimens, matched with the abnormal specimens for age and use or nonuse of an oral hormonal contraceptive, were examined. The smears were monolayer preparations stained with Thionin‐Feulgen Congored. Twenty‐one nuclear features were measured. A discrimination among the experimental groups could be made on the basis of the relationship between two features, area and average optical density (AOD). A regression of AOD on area for each smear was performed. The correlation, coefficient of variation, slope, intercept, as well as the mean of the AOD level and the age of the subject were used in a discriminant analysis. This resulted in a smear classification with a false‐positive rate of 14% and a missed‐positive rate of 32%. When contraceptive use was taken into account the overall classification was improved with a false‐positive rate of 12% and a missed‐positive rate of 20%.
Comparison of tissue disaggregation techniques of transitional cell bladder carcinomas for flow cytometry and chromosomal analysis
Authors: Smeets A. W. G. B. , Pauwels R. P. E. , Beck H. L. M. , Feitz W. F. J. , Geraedts J. P. M. , Debruyne F. M. J. , Laarakkers L. , Vooijs G. P. , Ramaekers F. C. S. 
Published: August 2005

AbstractDNA index (DI) measurements and chromosomal analysis of 42 transitional cell carcinomas were done after mechanical and enzymatical disaggregation of the tumor specimens. The results obtained with these different disaggregation techniques wee compared in the 33 cases (79%) that showed recognizable chromosomes. The enzymatically obtained cell suspensions could not be used for chromosomal analysis after short‐term culture of 24 hours. In four cases, the DI after enzymatical treatment could not be estimated. In most cases, the DI obtained from the tumor cells was similar for both aggregation techniques, with the exception of four cases of enzymatically treated cell suspensions in which the DI could not be estimated. The average DI of the aneuploid tumors was 13% higher than the corresponding chromosome count.In 19% of the aneuploid tumors the proportion of aneuploid cells could not be measured after enzymatical treatment. In the remaining suspensions the proportion of diploid cells was higher after enzymatical disaggregation than after mechanical treatment. It is concluded that for flow cytometric and direct chromosomal analysis of bladder tumors, the mechanical disaggregation technique is most suitable.
Membrane fluidity measured by fluorescence polarization using an EPICS V cell sorter
Authors: Fox Michael H. , Delohery Thomas M. 
Published: August 2005

AbstractWe have characterized the measurement of fluorescence polarization on single cells using an EPICS V cell sorter. A critical analysis is made of the balancing and calibration of the system. The system is highly linear for polarization measurements. Cellular membranes were labeled with 1,6‐diphenyl‐1,3,5‐hexatriene (DPH) to measure membrane fluidity. Fluorescence polarization histograms had coefficients of variation as low as 7%. Cells labeled with DPH after 24 hr incubation in medium lacking serum showed a significantly higher fluorescence polarization than cells in medium containing serum. The fluorescence polarization measured at 15°C was 0.311 compared to 0.270 at 25°C for cells labeled with DPH, verifying that temperature affects the membrane fluidity as measured by flow cytometry.
Spectroscopic properties of fluorescein in living lymphocytes
Authors: Kinoshita Shuichi , Fukami Tatsuya , Ido Yutaka , Kushida Takashi 
Published: August 2005

AbstractDetailed studies have been performed on various spectroscopic properties such as time dependence and excitation wavelength dependence of the fluorescence anisotropy for fluorescein molecules introduced into rat thymus lymphocytes. Experimental results have been found to be well interpreted in terms of the coexistence of two types of dye molecules, i.e., free and bound molecules. The fluorescence spectrum of only the bound molecules has been obtained from the difference in the time‐resolved spectra of fluorescence with two polarization directions. The time gate has been set at a sufficiently late time after the excitation, so that the polarization memories of the free molecules are lost. The spectrum thus determined agrees very well with that calculated from the spectral data in the stationary condition. From the above results, we come to the conclusion that the main factors which determine the fluorescence anisotropy inside the cell are the fraction and the anisotropy of the bound dye molecules. Finally, we discuss how these factors are related to biological quantities.
Flow cytometric analysis of Chlamydia trachomatis interaction with L cells
Authors: Waldman Frederic M. , Hadley W. Keith , Fulwyler Mack J. , Schachter Julius 
Published: August 2005

AbstractImmunofluorescent staining and flow cytometric analysis have been investigated as means of studying the early stages of in vitro infection of Chlamydia trachomatis. The lymphogranuloma venereum strain of C. trachomatis was grown in vitro in L cells, fixed in p‐formaldehyde, stained with fluorescein isothiocyanate (FITC)‐conjugated monoclonal antibody to the chlamydial major outer membrane protein, and analyzed flow cytometrically. Infected cells stained 50–100 times more intensely than uninfected cells, and they could easily be discriminated by flow analysis. The number of infected cells and the fluorescence intensity of individual cells were proportional to the multiplicity of infection. The attachment of purified elementary bodies to L cells could be analyzed by immunofluorescence and flow cytometry. Cells exposed to 0.26 inclusion‐forming units/cell could be discriminated from an unexposed population. Flow analysis of purified elementary bodies was possible after fluorescent staining with the aid of a laser‐based cytometer and gating on low volume.
Factors governing the flow cytometric analysis and sorting of large biological particles
Authors: Harkins Kristi R. , Galbraith David W. 
Published: August 2005

AbstractWe have investigated the suitability of large flow cell tips for the flow cytometric analysis and sorting of large biological particles, including plant cells (pollen) and protoplasts. Using flow tips ranging in diameter from 79–204 μm, we have optimized conditions for the establishment of a stable hydrodynamic flow leading to accurate droplet production. We describe instrument modifications required for large particle sorting and demonstrate the use of these experimental conditions for the sorting to high purity of pollen and viable plant protoplasts possessing diameters as large as 95 μm. Our experiments have revealed a complex interaction among sorting efficiency, particle diameter, flow cell tip diameter and bimorphic crystal drive frequency. This interaction can be satisfactorily explained in terms of interference effects owing to phase differences between the particle‐induced disturbance and the undulation driven by the bimorphic crystal.
Flow cytometric measurements of fluorescence energy transfer using single laser excitation
Authors: Szöllösi János , Mátyus László , Trón Lajos , Balázs Margit , Ember István , Fulwyler Mack J. , Damjanovich Sándor 
Published: August 2005

AbstractFlow cytometric energy transfer (FCET) measurements between labeled specific sites of cell surface elements (Szöllosi et al., Cytometry, 5:210–216, 1984) have been extended in a simplified form using a flow cytometer equipped with single excitation beam. This versatile and easily applicable method has several advantages over any nonflow cytometric (i.e., spectrofluorimetric) energy transfer measurements on cell surfaces: (1) The labeled ligands can be applied in excess, without washing, thereby enahling the investigation of relatively labile receptor‐ligand complexes. (2) Contributions of signals from cell debris, from cell aggregates, or from nonviable cells can be avoided by gating the data collection on the light scatter signal. (3) The heterogeneity of the cell population with respect to the proximity of the labeled binding sites can be studied. (4) In the cases of homologous ligands or of ligands binding to the same molecule but at different epitopes, the determination of fluorescence resonance energy transfer efficiency values can be carried out on a cell‐by‐cell basis, offering data on intramolecular conformational changes.This modified FCET method enabled us to demonstrate the uniform density of glycoproteins, specific for Con A binding, in the plasma membrane of normal and Gross virus leukemic mouse cells of different sizes. The utility of this procedure has also been demonstrated by using the mean fluorescence intensities of the distribution curves in the calculation of the fluorescence energy transfer efficiency.
Interctions of pyronin Y(G) with nucleic acids
Authors: Kapuscinski Jan , Darzynkiewicz Zbigniew 
Published: August 2005

AbstractSpectral properties of pyronin Y (PY) alone or in complexes with natural and synthetic nucleic acids of various base compositions have been studied in aqueous solution containing 10 or 150 mM NaCl and 5 mM Hepes at pH 7.0. The dimerization constant (KD = 6.27 × 103, M−1) and the absorption spectra of the dye in monomeric and dimeric form were established. The complexes of PY with single‐stranded (ss) nucleic acids show a hypsochromic shift in absorption, and their fluorescence is quenched by over 90% compared to free dye. In contrast, complexes with double‐stranded (ds) RNA or DNA (binding by intercalation) exhibit a bathochromic shift in their absorption (excitation) spectrum, and their fluorescence is correlated with the base composition of the binding site. Namely, guanine quenches fluorescence of PY by up to 90%, whereas A, C, I, T, and U bases exert a rather minor effect on the fluorescence quantum yield of the dye. The intrinsic association constant of the dye to ds RNA (Ki = 6.96 × 104, M−1) and to ds DNA (Ki = 1.74 × 104, M−1) was measured in 150 mM NaCl; the binding site size was 2‐3 base pair for both polymers. Implications of these findings for qualitative and quantitative cytochemistry of nucleic acids are discussed.
Application of pyronin Y(G) in cytochemistry of nucleic acids
Authors: Darzynkiewicz Zbigniew , Kapuscinski Jan , Traganos Frank , Crissman Harry A. 
Published: August 2005

AbstractChinese hamster ovary (CHO) cells or isolated nuclei were stained with pyronin Y (PY) and analyzed by absorption or fluorescence microscopy, as well as by flow cytometry. Specificity of the staining reaction was assayed by testing sensitivity of the stainable material to RNase or DNase. The colored complexes detected by light absorption in fixed cells stained with PY are nonfluorescent and are most likely the products of condensation of single‐stranded (ss) RNA by PY; the poly(rA) and poly(rA, rG) are the most sensitive to condensation.The products of PY interaction with double‐stranded (ds) nucleic acids are fluorescent and can be detected in cells by cytofluorometry. PY used alone stains both DNA and RNA, and the staining capabilities of these nucleic acids vary depending upon the PY concentration at equilibrium; at a concentration above 330 μM, the RNA stainability decreases, perhaps due to its denaturation and condensation caused by the dye. In the presence of Hoechst 33342, PY can specifically stain RNA in fixed cells or isolated cell nuclei. Because only complexes of PY with ds RNA are fluorescent, this dye can be used as a probe of RNA conformation, e.g., to monitor denaturation of RNA in situ. The RNA stainability of mitotic cells is about 25% lower than that of cells in G2 phase, which indicates that during mitosis proportionately less cellular RNA is in the ds conformation. The advantages and limitations of the two cytochemical methods for DNA/RNA detection, one based on the use of Hoechst 33342 and PY, and another employing the metachromatic properties of acridine orange, are compared.
Method for detection and isolation of cholesteryl ester‐containing “foam” cells using flow cytometry
Authors: Kruth Howard S. , Cupp James E. , Khan Mushtaq A. 
Published: August 2005

AbstractAccumulation of cholesteryl ester within vascular cells is a defining characteristic of atherosclerotic lesions. Therefore, it is of interest to be able to monitor this critical event in the development of atherosclerosis. With this objective in mind, we have developed a method for the detection of cholesteryl ester‐containing cells (i.e., foam cells) in cell suspensions prepared from enzymatically dissociated aortas.Cholesteryl ester in aortic cells was selectively stained with the fluorescent dye filipin. Because filipin binds to unesterified cholesterol but not to esterified cholesterol, it was necessary first to remove unesterified cholesterol from cells by ethanol extraction so that its presence would not interfere with the specific detection of cholesteryl ester. Then unesterified cholesterol made available by enzymatic hydrolysis of cellular cholesteryl ester could be specifically stained with filipin. The filipin‐stained cell suspensions were analyzed using flow cytometry. With a flow cytometer it was possible to detect and sort cholesteryl ester‐containing cells onto glass slides for microscopic analysis.Cell suspensions prepared from either grossly normal or atherosclerotic swine aortas contained cells with cholesteryl ester inclusions. As expected, these cells were more numerous in the atherosclerotic aortas. Cells with higher levels of fluorescence contained more numerous cholesteryl ester inclusions. Flow cytometric detection of cholesteryl ester‐containing cells should be generally useful in studies of cellular cholesterol metabolism as well as in specific studies of cellular cholesterol accumulation in atherosclerotic vessels.
In vivo cell kinetic effects of cis‐platinum on human ovarian cancer xenografts measured by dual parameter flow cytometry
Authors: Sevin Bernd‐uwe , Pollack Alan , Averette Hervy E. , Ramos Reinaldo , Donato Daniel 
Published: August 2005

AbstractDual parameter flow cytometry, measuring DNA and nuclear protein of individual cell nuclei simultaneously, makes it possible to follow cell kinetic perturbations in six distinct compartments of the cell cycle following chemotherapy in vivo. Human ovarian cancer xenografts in nude mice from a primary and recurrent tumor of the same patient were studied. The response to intraperitoneal application of cis‐platinum was assessed by tumor volume measurements, changes in labeling indices by autoradiography, and dual parameter flow cytometry. Sequential tissue samples were taken from each tumor using fine needle aspirations as a microbiopy method. Pretherapy samples were compared to multiple specimens collected up to 18 days after therapy. Morphologic changes of each specimen were also assessed. Cis‐platinum affects malignant cells in the G1B, S, G2A, and G2B compartments with various intensities and different time frames, depending on the drug sensitivity of each individual tumor.
Evaluation of contextual analysis for computer classification of cervical smears
Authors: Garcia Gerardo L. , Kuklinski Walter S. , Zahniser David J. , Oud Peter S. , Vooys Peter G. , Brenner John F. 
Published: August 2005

AbstractA procedure for automated analysis of cervical smears has been implemented in an image cytometry system. Smears are described exclusively in terms of global and contextual information extracted by pattern‐recognition algorithms and represented by a vector of proportions of cellular object types. Linear discriminant functions, based on a Fisher criterion, are derived to classify smears with a cross‐section of diagnoses into two broad categories, normal and abnormal. Results obtained from 83 smears indicate 78% correct classification. In contrast to most automated systems, good classification results were obtained in normal smears with benign changes caused by inflammation and with postmenopausal atrophia and in abnormals with mild dysplasia. These findings suggest that contextual analysis may be sensitive to subtle changes in cellular morphology and to progressive patterns of dysplasia. When used with standard isolated cell analysis, contextual analysis may provide additional complementary information for automated cervical prescreening.
The reliability of microspectrophotometric and flow cytometric nuclear DNA measurements in adenocarcinomas of the breast
Authors: Fallenius Anders G. , Askensten Ursula G. , Skoog Lambert K. , Auer Gert U. 
Published: August 2005

AbstractThe reliability of microspectrophotometric (MSP) and flow cytometric (FCM) nuclear DNA measurements has been studied in 50 human breast adenocarcinomas. The tumor material was obtained by means of fine‐needle aspiration biopsy, and all samples except one were found to be highly representative. The results confirm earlier observations that a good correlation exists between modal value (MV) determined by MSP and DNA index (DI) determined by FCM. However, when tumors were classified into low and high malignant variants according to FCM/DI, FCM/S‐phase percentages, and MSP histogram types, the concordance was less pronounced. This was found to be due mainly to the fact that in near‐diploid tumors a discrepancy exists between MSP and FCM ploidy, as well as between MSP distribution pattern and the estimated percentages of cells in the S‐phase region. Another source of discrepancy was observed in tumors with stemlines in the normal tetraploid region, including cells with highly scattered aneuploid DNA values. These tumors were judged by MSP as aneuploid/high malignant and by FCM as euploid/low malignant. In view of this discrepancy, we conclude that the simple determination of the stemline position by MSP/MV or FCM/DI is not sufficient for adequate cytochemical malignancy grading of breast carcinomas. We suggest that a combination of ploidy and percentage of cells scattered outside the modal peaks is a more sensitive method for optimal cytochemical malignancy grading in breast carcinomas.
Improved model for specimen classification based on single‐cell classifiers
Authors: Cox C. , Wheeless L. L. , Reeder J. E. , Robinson R. D. , Berkan T. K. 
Published: August 2005

AbstractWe consider probabilistic models for specimen classification procedures based on systems which classify individual cells as normal or abnormal. The models which we consider generalize those discussed previously by Castleman and White (Anal. Quant. Cytol. 2:117–122, 1980; Cytometry 2:155–158, 1981) and by Timmers and Gelsema (Cytometry 6:22–25, 1985). In particular, they include the biologically plausible possibility that the specimen contains cells which are intermediate between the extremes of normal and abnormal. We find that if these additional cells occur differentially in normal and abnormal specimens, then specimen classification can become substantially more efficient when the cell classifier has different error rates for these cells.
Centromeric index versus DNA content flow karyotypes of human chromosomes measured by means of slit‐scan flow cytometry
Authors: Lucas J. N. , Gray J. W. 
Published: August 2005

AbstractWe have applied slit‐scan flow cytometry (SSFCM) to classify human chromosomes according to their centromeric index (CI) and relative DNA content. The resulting bivariate—CI vs. DNA content—distributions shows 14 peaks for normal human chromosomes. Distinct peaks are produced by chromosomes 1, 2, 3, 4+5, 6+7+X, 8, 13+14+15, 16, 17+18, 19+20, and 21+22+Y. In addition, chromosomes 9 through 12 are resolved into three peaks. The identity of the chromosomes comprising each peak was determined by comparing CI vs. DNA content distributions measured for normal human chromosomes by means of SSFCM with CI and DNA content values measured for human chromosomes with image analysis. The accuracy of CI measurement by SSFCM was verified by measuring CIs for human chromosomes isolated from human/rodent hybrid cell lines containing only a few known human chromosomes. These studies showed CIs measured for human chromosomes 1–19 and 21 to be inclose agreement with the CIs calculated by means of image analysis. We further confirmed the chromosome assignments for each peak by showing that the relative volumes of the peaks in the CI vs. DNA content distributions for chromosomes from normal cells are similar to the relative frequencies of chromosomes expected for these peaks based on the peak assignments.
Quantitative analysis of a nuclear antigen in interphase and mitotic cells
Authors: Clevenger Charles V. , Epstein Alan L. , Bauer Kenneth D. 
Published: August 2005

AbstractThe quantification of an interchromatin‐as‐sociated antigen, designated p105, during cellular passage through mitosis is described. Indirect immunofluorescence microscopy and immunogold electron microscopy demonstrated a qualitative increase in p105 within the mitotic cytoplasm. Multiparameter flow cytometric analysis was performed on fixed cells sequentially stained with anti‐p105 immunofluorescence and/or propidium iodide. This analysis demonstrated approximately a tenfold increase in intracellular p105 content as a function of progression from the G2 to the M phase. This increase was corroborated by the quantitative immunoblot analysis of colchicine‐treated cell cultures and of cells sorted on the basis of anti‐p105 immunofluorescence. The data reveal that the increased levels of anti‐p105 immunofluorescence in conjuction with flow cytometry may be used effectively to quantitate mitotic index and isolate mitotic cells. The function and modulation of p105 throughout the cell cycle is discussed.
Comparison of synchronized Chinese hamster ovary cells obtained by mitotic shake‐off, hydroxyurea, aphidicolin, or methotrexate
Authors: Fox Michael H. , Read Robert A. , Bedford Joel S. 
Published: August 2005

AbstractSynchronized cell populations are necessary to study many aspects of cell biology. We have developed a method to obtain highly synchronized Chinese hamster ovary cell populations in S phase or G2 phase by utilizingmitotic selection followed by incubation with either hydroxyurea, aphidicolin, or methotrexate for 12 h. Flow cytometry analysis shows that the coefficient of variation in the spread of the cell population in S phase is as low as 6%. Drug toxicity studies compare the effects of the various drugs on G1 and S phase cells. The use of aphidicolin or hydroxyurea results in the most highly synchronized cell populations, but methotrexate yields inadequate synchronization. These results demonstrate that both aphidicolin and hydroxyurea are useful drugs for obtaining highly synchronized cell populations after an initial synchrony in mitosis. Aphidicolin is perhaps the best choice because of less toxicity to S phase cells when used in low concentrations.
Monoclonal antibody isolation of type II pneumocytes
Authors: Funkhouser Jane D. , Cheshire L. Bryan , Read Robert J. , Hester Raymond B. , Peterson Raymond D. A. 
Published: August 2005

AbstractA monoclonal antibody that identifies a membrane molecule unique in rat lung for type II alveolar epithelial cells was used to isolate these cells from enzymatically dispersed lung cells by fluorescence‐activated cell sorting. Although multistep physical separation techniques have permitted the isolation of large quantities of these cells and flow cytometry has been used by others to isolate lamellar body‐containing cells, the application of this antibody‐directed sorting has distinct advantages. Because the marker molecule is expressed on immature type II cells prior to the development of lamellar bodies, the antibody will also permit their isolation and study.