bluuebunny/crossref_metadata_2025_split huggingface dataset:
Application of orthogonal light scattering for routine screening of lymphocyte samples
Authors: Terstappen L. W. M. M. , De Grooth B. G. , Greve J. , VanBerkel W. , TenNapel C. H. H. 
Published: August 2005

AbstractOrthogonal and forward light‐scattering properties of lymphocytes were measured from patients with different lymphocytic diseases in order to determine the potential value of light scattering as a screening device. Monitoring of orthogonal light scattering of lymphocytes of a B‐cell chronic lymphocytic leukemia patient during splenic irradiation (SI) revealed the selective decrease of malignant cells and the fact that the major part of the residual lymphocytes were cytotoxic lymphocytes. By combining forward and orthogonal light scattering it was shown that lymphocytes from a patient with Tλ lymphocytosis were abnormal. Orthogonal light scattering also showed an increase in cytotoxic lymphocytes in a patient with mononucleosis infectiosa and in a splenectomized patient.Orthogonal light scattering of lymphocyte subpopulations showed that the leu8+ population of a patient with mononucleosis infectiosa was bidisperse. For elderly donors the occurrence of CD3+, CD4+, CD8+, and HNK‐1+ lymphocytes with a large orthogonal light scattering varied considerably. The CD8+ lymphocytes of these donors consisted mainly of cytotoxic lymphocytes.These results show that determination of light‐scattering properties of lymphocytes may yield important diagnostic information and can indicate when further investigation of the lymphocytes by means of immunofluorescence is necessary.
Electrophoretic mobility studies on doxorubicin‐resistant and ‐sensitive murine P338 leukemic cells
Authors: Nair Swagata , Horton Allan , Leif Robert C. , Krishan Awtar 
Published: August 2005

AbstractThe electrokinetic properties of doxorubicin (DOX)‐resistant (P388/R) and ‐sensitive (P388/S) murine leukemic cells were studied in a free‐flow electrophoresis (FFE) system. The electrophoretic mobilities (EM) of P388/S and P388/R cells were 1.07 and 1.35 × 10−4cm2V−1S−1, respectively, suggesting a higher net negative charge on the P388/R cells. Neuraminidase treatment decreased the EM of both the P388/S and P388/R cells by 15–20% but had no effect on cellular doxorubicin retention. Total and cell surface sialic acid contents were similar in both the cell lines. Our studies show that no direct correlations may exist among surface charge, cell surface sialic acid content, and doxorubicin retention in DOX‐resistant and ‐sensitive P388 cells; however, differences in cell surface charge between these cell types were used to separate them by preparative FFE.
Evaluation of four methods of DNA distribution data analysis based on bromodeoxyuridine/DNA bivariate data
Authors: Lacombe F. , Belloc F. , Bernard P. , Boisseau M. R. 
Published: August 2005

AbstractFour published methods of DNA‐content histogram analysis (those of Fried, Dean and Jett, simplified Dean, and Fox) were compared using a double labeling of different cell populations. Partially synchronized and asynchronous cell populations were incubated with bromodeoxyuridine (BrdUrd) and then stained with an anti‐BrdUrd monoclonal antibody and propidium iodide (PI). The fractions of cells in the G1, S, and G2 + M phases were calculated by each method and compared with those derived from G1, S, and G2 + M areas plotted on BrdUrd/DNA bivariate histograms, taken as the “true” values. This procedure enabled an optimal choice of method for a given cell population.
Different quiescence states of three culture cell lines detected by acridine orange staining of cellular RNA
Authors: Baisch Heinz 
Published: August 2005

AbstractThree cell lines (CHO, L‐929 and R1H) were investigated for their growth kinetics and the difference of exponential and quiescent state of monolayers in medium with and without serum (L‐929). The noncycling populations of L‐929 and R1H in medium with serum contained increased G1‐phase percentages but also considerable proportions of SQ and G2Q cells. Although about 90% of the cells excluded trypan blue, the viability tested by colony assay was clearly lower than for exponentially growing cultures. CHO cells showed similar fractions of cells in G1‐, S‐, and G2‐Q compartments but in addition considerable cell loss. The RNA content of these cells was reduced in plateau phase by 7–48% depending on cell type and residence time in the noncycling state. The data suggest that the cells suffered from nutrition depletion and were arrested in all phases of the cycle. In contrast, L‐929 cells in medium without serum reduced their RNA content down to one‐third that of proliferating cells and still retained the full viability as shown by the same plating efficiency in a colony assay. Since about 90% of the cells had G1 DNA content, these cells resemble true G1Q or G0 cells controlled by growth factors rather than nutritional depletion.
Bromodeoxyuridine amplifies the inhibitory effect of oxygen on cell proliferation
Authors: Poot Martin , Schindler Detlev , Kubbies Manfred , Hoehn Holger , Rabinovitch Peter S. 
Published: August 2005

AbstractThe BrdUrd‐Hoechst method was used to analyze the interaction of various oxygen concentrations with BrdUrd substituted DNA with respect to cellular proliferation. At oxygen concentrations above 5%, human diploid fibroblast‐like cells and amniotic fluid fibroblast‐like cells showed reduced proliferation rates, which resulted from an increase in noncycling cells and from a permanent arrest of cells in the G2 phase of the cell cycle. At 35% oxygen the increase in noncyling cell fraction and the permanent arrest in G2 was strongly dependent upon the concentration of BrdUrd. Incorporation of BrdUrd into DNA, therefore, amplifies the adverse effects of increasing oxygen concentrations upon cell proliferation. The mechanism of this amplification might involve a free radical attack on DNA similar to the radiation sensitizing effect of BrdUrd.
Microtubule and microfilament distribution and tubulin content in the cell cycle of indian muntjac cells
Authors: Duanshun Wang , Suwen Li , Changqing Zeng , Ruxuan Cheng , Shaobai Xue 
Published: August 2005

AbstractUsing DAPI, rabbit antitubulin antibody, FITC‐labeled goat anti‐rabbit IgG, and TRITC‐phalloidin to stain individual cells, the microspectrophotometric analysis showed that three markers that represent the nucleus, microtubules (MT), and microfilaments (MF), respectively, could be recognized in individual cells without interference. The phase of the cell cycle was determined by DNA content. We found that in Indian muntjac (IM) cells, the amount of tubulin in G2 and M phases was about twice as much as that in G1 phase. In G2 cells, the cytoplasmic microtubule complex (CMTC) became denser than in G1 cells. The cytoplasmic MT extent in basically the same orientation as MF bundles in interphase. The regions where the MT is denser also have a denser MF distribution.
Histological grading, DNA content, cell proliferation and survival of patients with astroglial tumors
Authors: Zaprianov Z. , Christov K. 
Published: August 2005

AbstractLight microscopy, image cytometry (ICM), and flow cytometry (FCM) were used to study the degree of differentiation, DNA content, and S‐phase of astrocytomas and glioblastoma multiforme in 102 patients. The postoperative real survival time (RST) was also studied.Using ICM, 62 astrocytomas were investigated. Grade I astrocytomas were composed of DNA‐diploid cell lines, while grade III and glioblastoma multiforme consisted predominantly of DNA‐aneuploid lines. Moderately differentiated astrocytomas were divided as follows: 14 DNA‐diploid and 18 DNA‐aneuploid. Forty astrocytomas were studied by FCM. Using the DNA index (DI) value, cases with abnormal DNA cell lines were established in all astrocytomas, with their number increasing in grades II and III astrocytomas. FCM indicated the same subdivision of moderately differentiated astrocytomas: 12 with DNA‐diploid and 12 with DNA‐aneuploid stem lines. Patients with DNA‐diploid cell lines in the astrocytomas and low S‐fraction survived longer than patients with abnormal DNA cell populations and higher S‐fraction.The results from this study indicate that, together with the degree of differentiation of astroglial tumors, the appearance of cell lines with abnormal DNA value and higher S‐fractions also have prognostic value.
Measurement of cell motility and morphology with an automated microscope system
Authors: Thurston Gavin , Jaggi Bruno , Palcic Branko 
Published: August 2005

AbstractA method is presented which provides quantitative descriptions of the morphology and motility behaviour of a large number of unperturbed individual cells. For that, a microscope system has been developed that performs automated tracking of motile mammalian cells in tissue culture and the concurrent measurement of cell morphology. Experiments using 3T3 mouse fibroblasts show that simple global morphological features, such as degree of cell elongation, correlate with the motile behaviour of individual cells. Combined data from many cells following trypsinization and replating show the time course of the spreading of cells, measured by increase in average cell area, and the onset of cell polarization, measured by change in average cell circularity. Methods for describing fibroblast morphology and changes in morphology are discussed.
Intracellular pH measurements using flow cytometry with 1,4‐diacetoxy‐2,3‐dicyanobenzene
Authors: Cook John A. , Fox Michael H. 
Published: August 2005

Abstract1,4 ‐ Diacetoxy ‐ 2,3 ‐ dicyanobenzene (ADB) has been increasingly used for measurement of intracellular pH by flow cytometry. ADB rapidly enters cells and is cleaved to the fluorescent pH indicator 2,3‐dicyano‐hydroquinone (DCH). We have analyzed several potential problems that can affect its usefulness as a pH indicator. Hydrolysis of ADB in aqueous solutions reveals the temporary presence of a fluorescent species blue‐shifted from DCH at the same pH. The presence of this species with DCH can lead to erroneous pH measurements. Stable pH measurements with ADB depend on the incubation conditions and esterase activity. Heated cells required 20 min for stable measurements, whereas control cells required 5 to 10 min. The reproducibility of pH measurements was excellent, with a resolution of ≤ 0.05 pH units in the range of 6.4 to 8.0. Absolute calibration curves of intracellular pH using the ionophore nigericin depended on matching the intracellular K+ concentration with the buffer, but relative measurements of intracellular pH were insensitive to K+. ADB was nontoxic to Chinese hamster ovary cells at up to 20 μg/ml. However, when cells loaded with dye were passed through a UV laser beam, concentrations of dye > 5 μg/ml were highly toxic. Viable cells could be sorted on the basis of intracellular pH if ADB were used at low concentrations.
Identification of inflammatory cells in bovine milk by flow cytometry
Authors: Redelman Doug , Butler Sara , Robison Jon , Garner Duane 
Published: August 2005

AbstractCells recovered from normal or mastitic bovine milk were examined by flow cytometry. All milk samples contained particulate material that was heterogeneous in size and that produced a right‐angle light‐scatter signal equal to or greater than that produced by human or bovine neutrophils. Although this material labeled with Hoechst 33342, it produced fluorescence intensities below that of intact bovine cells, suggesting that it consisted of cell fragments. Mastitic milk additionally contained other populations of cells that were poorly resolved from the normal particulate material by size (electronic volume sensor) and right‐angle light scatter. In order to improve this resolution, the milk cells were incubated with carboxydimethylfluorescein diacetate (CMFDA) to label intact cells. When milk samples labled with CMFDA were examined by dual‐parameter analysis using green fluorescence and right‐angle light scatter, five or more populations of cells could be identified in mastitic milk. These populations included intact and degenerate neutrophils, lymphocytes, including both small and activated cells, monocytes, and large activated macrophages containing many vacuoles and phagocytosed particles. Using this procedure, all the animals in the University of Nevada‐Reno Holstein dairy herd were tested once a month for 6 months. In addition, individual animals with mastitis were examined one or more times each day during the course of the inflammatory process. In the routine screening, the flow cytometric examination detected mastitis before overt symptoms developed. In cows identified to have mastitis, the flow cytometric examination provided prognostic information regarding the success of treatments.
Localization of monoclonal B‐cell populations through the use of Komogorov‐Smirnov D‐value and reduced chi‐square contours
Authors: Bagwell C. Bruce , Lovett Edmund J. , Ault Kenneth A. 
Published: August 2005

AbstractIn this study three different flow cytometric analysis techniques are woven together into a single system that permits improved detection of small percentages of monoclonal B cells in a milieu of either normal blood leukocytes or bone marrow cells.This analysis is an extension of the concept of clonal excess, which is used to detect the presence of a tumor that is a clonal expansion of B cells expressing either kappa or lambda light chains.The technique also utilizes “multiple list‐mode processing,” which is defined in this context as the simultaneous analysis of two or more listmode files that share one or more common parameters. This type of data structure enables the segmentation of two parameter light scatter displays into regions from which numerous kappa and lambda histograms subsequently can be analyzed for their respective Komogorov‐Smirnov D‐values or R‐values (reduced chi‐square value).The final technique makes use of a calculated parameter display system. Superimposed on the light scatter dot density plot are D‐value or R‐value contours. The contours target the location of the population that is abnormal, thus providing information for setting optimal bitmap gates for clonal excess studies, other phenotypic analyses, or cell sorting.In experiments using model systems, the sensitivity of this assay is estimated to be between 0.25% and 2.5%. The technique's distribution information and sensitivity may prove useful for staging, treatment monitoring, and relapse detection of B‐cell leukemia and lymphoma.This application illustrates the potential of combining multiple listmode processing and calculated parameter display to expand the effective dimensionality of listmode data.
How reproducible are flow cytometry data from paraffin‐embedded blocks?
Authors: Kute T. E. , Gregory B. , Galleshaw J. , Hopkins M. , Buss D. , Case D. 
Published: August 2005

AbstractThe purpose of this technical report is to determine the reproducibility of flow cytometry data for ploidy and cell cycle kinetics using paraffin‐embedded blocks of breast cancer tissue. One block from each of 39 tumors was studied in this report with each block having multiple sections analyzed independently. All of these sections gave ploidy analyses, while only 34 gave cell kinetic values. The standard deviation for the DNA index value in the multiple analysis study was less than 0.1 in all but three cases. The cell kinetic values gave larger variability, and the actual values were dependent on the method of analysis. Comparison of the variability for each method of analysis could not predict which procedure was superior. These results would indicate that ploidy is a reproducible value, while cell kinetic parameters should only be used as an indicator of proliferative activity that has been normalized to the mean or median of a large set of observations processed and analyzed by the same procedure.
A flow cytometric assay for intracellular nonprotein thiols using mercury orange
Authors: O'Connor Jose Enrique , Kimler Bruce F. , Morgan Michael C. , Tempas Kathryn J. 
Published: August 2005

AbstractThe level of nonprotein thiols was assayed in individual mammalian cells using flow cytometry. Previous determinations of glutathione (GSH, the most abundant nonprotein thiol in most cells) by flow cytometry were based on UV laser excitation of fluorochromes. Because of several shortcoming of UV excitation, an assay for GSH using visible light is of interest. Selective staining of nonprotein thiols with mercury orange (a mercurial compound that binds stoichiometrically to sulfhydryl groups) was obtained by restricting the staining time. By using various drugs that affect GSH levels and overall thiol levels in cells, it was shown that GSH is the primary thiol group being stained. Thus a quick, specific technique using mercury orange has been developed for the flow cytometric determination of nonprotein thiols and preferentially for GSH in individual mammalian cells.
Conversion of logarithmic channel numbers into relative linear fluorescence intensity
Authors: Schmid Ingrid , Schmid Peter , Giorgi Janis V. 
Published: August 2005

AbstractWe describe a simple, reproducible, and generally applicable method to assess the performance of log amplifiers by using a fluorescent sample that provides multiple peaks of different intensities. The channel differences between multiple peaks are used to evaluate the logarithmic behavior of the fluorescence signal amplifier on the flow cytometer. A calibration curve can be created to correct the channel numbers for deviations from true logarithmic behavior and then convert data into relative linear intensities. By using these linear fluorescent intensities, we compared the capacity of different antisera against HIV‐1 (human immunodeficiency virus type 1) peptides to inhibit the binding of HIV‐1 to CEM, a CD4‐positive T‐cell line. A wide range of applications for this calibration procedure can be envisioned and the method is valuable for monitoring instrument performance over time.
Fluorescence response and sensitivity determination for ATC 3000 flow cytometer
Authors: Gaucher J. C. , Grunwald D. , Frelat G. 
Published: August 2005

AbstractThe fluorescence emitted by labeled particles after interaction with exciting light is conditioned by laser beam geometry and by the mode of fluorescence collection and filtration. A laser elliptic focusing mode is described, and the fluorescence characteristics of the sample cell flow are calculated. Fluorescence collection and detection through optical filters were analyzed, and efficiency was calculated for the ATC 3000 flow cytometer (Odam‐Bruker, Wissembourg, France). A mathematical model is proposed for calculation of the fluorescence signal and its fluctuations. The background noise for the ATC 3000 was quantified experimentally using fluorescent microspheres of a known number of bound equivalent fluorescein isothiocyanate (FITC) molecules. These experimental measurements were found to fit the theoretical predictions, thus validating the proposed model.
Imaging system for morphometric assessment of absorption or fluorescence in stained cells
Authors: Jaggi B. , Poon S. S. S. , Macaulay C. , Palcic B. 
Published: August 2005

AbstractAn image acquisition and processing system has been developed for quantitative microscopy of absorption or fluorescence in stained cells. Three different light transducers are used in the system to exploit the best characteristics of these sensors for different biological measurements. A digital scanner, in the form of a linear array charge‐coupled device (CCD), acquires data with high spatial and photometric resolution. A color (RGB) camera is employed when spectral information is required for the segmentation of cellular subcomponents. An image‐intensified charged‐injection device (CID) camera provides for very low light intensity measurements, primarily for fluorescence‐labeled cells. Properties of these transducers, such as contrast transfer function, linearity, and photo‐response nonuniformity, have been measured.Two dedicated image processing units were incorporated into the system. The front‐end processor, based on a digital signal processor, provides functions such as object detection, raw image calibration, compression, artifact removal, and filtering. The second image processor is associated with the frame memory and includes a histogram processor, a dedicated arithmetic logic unit for image processing functions, and a graphics module for onebit overlay functions. An interactive program was developed to acquire cell images and to experiment with a range of segmentation algorithms, feature extractions, and other image processing functions. The results of any image operation are displayed on the video monitor. Once a desired processing sequence is determined, the sequence may be stored to become part of a command library and can be executed thereafter as a single instruction.
Image cytometry of estrogen receptors in breast carcinomas
Authors: Cohen Olivier , Brugal GÉRard , Seigneurin Daniel , Demongeot Jacques 
Published: August 2005

AbstractA significant level of estrogen receptors (ER) in breast cancer cells is an indication of tumor differentiation and suggests that a homeostatic control of cell growth may persist in these cancers. In medical practice, the Dextran‐coated charcoal assays (DCCA) are still the most frequently used test to characterize patients having ER‐positive malignant breast tumors and for whom hormonal therapy is justified. Nevertheless, this routine biochemical technique is not satisfactory because it is a broad method unsuitable for revealing receptor tissue heterogeneity. However, immunocytochemical labeling, such as the ERICA method, which involves a monoclonal antibody linked to peroxidase, is a specific reaction for this purpose but which until now was not quantitative. The present study uses an original cell preparation technique combining the PAP reaction with toluidine blue counterstain for image analysis on the SAMBA system. Special software has been developed for the quantitative analysis of immunocytochemistry in cancers. Results obtained showed a high correlation between the DCCA values and the score derived from the mean ER concentration per positive tumor cell and the labeling index. In addition, intracell and intratumor heterogeneity can be displayed according to several parameters and were shown to vary according to tumor and to antiestrogen (Tamoxifen) presurgical therapy.
Modified histogram subtraction technique for analysis of flow cytometry data
Authors: Overton W. Roy 
Published: August 2005

AbstractAnalysis of flow cytometry histogram data by the subjective selection of an integration window can be a tedious and time‐consuming task and is often inaccurate. A new method for automated calculation of the percent positive from immunofluorescence histograms is presented. This new method is a modification of the currently used method of channel‐by‐channel histogram subtraction. Its accuracy is compared to that of the channel‐by‐channel histogram subtraction method and to another currently used automated method, which selects an integration window by finding the channels that contain the most fluorescent 2% of a control histogram.The new histogram subtraction method is objective, easy to use, and is more accurate than other currently used automated analysis methods. PASCAL source code is given for each method of analysis.