bluuebunny/crossref_metadata_2025_split huggingface dataset:
Flow cytometric DNA content in myelodysplastic syndromes
Authors: Montecucco Carlomaurizio , Riccardi Alberto , Traversi Egidio , Danova Marco , Ucci Giovanni , Mazzini Giuliano , Giordano Paolo 
Published: August 2005

AbstractDNA flow cytometric analysis of unfixed bone marrow cells stained with propidium iodide was carried out in 33 patients with untreated primary myelodysplastic syndromes. Patients with stable clinical course for up to 3 years had higher fractions of cells in S and G2 phases (22.7 ± 12.4% and 12 ± 3.6%) than those who developed acute leukemia and/or died early in the course of disease (14.4 ± 8.5% and 6.6 ± 4%). Median survival was more than 36 mo in patients with S+G2 cell fraction higher than 24%, and 14 mo in the remaining 16 patients with lower values (P < 0.01). Analyses repeated after 3–24 mo showed no major changes in cell proliferation pattern in ten out of 11 patients. The remaining patient had sharp decrease in S and G2 cell fraction 3 mo before the transition into acute leukemia. The DNA index (DI) of bone marrow cells was calculated to assess ploidy. However, comparative evaluation of cytologic, cytogenetic, and flow cytometric data suggest that, under our experimental conditions, the DI may be influenced by factors such as the degree of chromatin compactness.
Continuous measurement and analysis of staining kinetics by flow cytometry
Authors: Beumer Tom , Pennings Arie , Beck Hans , Wessels Johan , Haanen Clemens , Herman Chester 
Published: August 2005

AbstractThe measurement of color development with time in cells following the start of a staining reaction is of interest in a number of biological systems. These include the subsets of peripheral white blood cells after acridine orange staining, the uptake by cells and nuclei of fluorescent agents, especially antitumor drugs, and measurement of intracellular enzyme kinetics using fluorogenic or absorbing substrates. The present work describes a simple computer program for analyzing flow cytometric (FCM) data versus time, including both the population kinetics of color development and the variability of staining speed within one population of cells.A single‐channel absorption measurement in flow (Technicon Hemalog D) was used to record peroxidase kinetics in peripheral blood cells. Every 5 s, a 64‐channel absorption histogram was recorded, up to a maximum of 64 histograms. The data were then analyzed by a computer program which searched for the peak channel of each histogram. A least‐squares fit was computed for these maxima. The asymmetries of the 64 absorption histograms were compared to see if there was more than one population present with different time constants.Although developed for enzyme kinetic measurements, this program may have wider usefulness in any measurements of time‐dependent phenomena by FCM.
Characterization and sorting of mouse cytotoxic macrophages by their light scattering properties
Authors: Martin A. , Merdrignac G. , Collet B. , Genetet B. 
Published: August 2005

AbstractThe light scattering properties of mouse activated macrophages were analyzed by flow cytometry. Peritoneal adherent cells from B. abortus treated animals were found to segregate into two subpopulations as a function of their foward angle and 90° angle light scatter. The cell subpopulations were separated by automatic sorting. The strongly scattering ones contained an elevated proportion of large volume and acid phosphatase rich cells. Their nonspecific cytotoxic activity against tumor cells was more important than that of weakly light scattering cells. Thus, flow cytometry might be helpful to characterize and isolate cytotoxic macrophage populations.
Simultaneous measurement of phagocytosis and phagosomal pH by flow cytometry: Role of polymorphonuclear neutrophilic leukocyte granules in phagosome acidification
Authors: Bassøe C.‐F. , Laerum O. D. , Glette J. , Hopen G. , Haneberg B. , Solberg C. O. 
Published: August 2005

AbstractHuman polymorphonuclear neutrophilic leukocytes (PMNLs) phagocytosed fluorescein‐isothiocyanate (FITC)‐labelled Staphy‐lococcus aureus. Free bacteria, phagocytes, and nonphagocytes were discriminated and quantified by flow cytometry (FCM). The relative fluorescence of phagocyte‐associated and free bacteria (Nf:N) was calculated by dividing the mean phagocyte fluorescence by that of the free bacteria and the number of phagocytosed bacteria. Bactericidal capacity and chemiluminescence were measured by standard methods. The red‐to‐green fluorescence ratio of acridine orange‐stained PMNLs (R/G) was measured by FCM. Degradation of bacteria was monitored by the reduction in FITC and ethidiumbromide fluorescence of bacteria liberated from the phagocytes. Bacterial FITC fluorscence was pH dependent. Nf:N was 0.5 to 0.7. Using a standard curve for the interrelationship between bacterial fluorescence and pH, phagosomal pH was 5.0–5.5 Phagocytes, kept at 4°C for 24 h had Nf:N ∼ 1, did not degrade bacteria, but killed them and emitted chemiluminescence. NH4Cl increased phagocyte fluorescence by 27% and decreased R/G by 50%. Cyanide and azide did not affect Nf:N. Nf:N of phagocytes from a patient with chronic granulomatous disease was 32% below, and R/G was 32% higher than the controls. Acidification of the phagosomes seems to be related to discharge of PMNL granule contents and independent of the respiratory burst.
Immunofluorescence measurement in a flow cytometer using low‐power helium—neon laser excitation
Authors: Shapiro Howard M. , Glazer Alexander N. , Christenson Lisa , Williams John M. , Strom Terry B. 
Published: August 2005

AbstractHelium‐neon lasers are economical and efficient light sources; their utility in flow cytometry to date has been limited by the lack of fluorescent probes that can be excited at 633 nm. Allophycocyanin (APC), a highly fluorescent phycobiliprotein, can be used as an antibody label and has spectral characteristics suitable for use with He‐Ne lasers; we undertook to resolve whether a low‐power (7 mW) He‐Ne laser could provide sufficient excitation to permit flow cytometric detection of APC‐labeled antibodies on cell surfaces. We made an APC conjugate of monoclonal antibody 4F2, which reacts with an antigen abundant on the surfaces of activated human T‐lymphocytes; APC‐4F2 was used to stain blood mononuclear cells that had been cultured with and without phytohemagglutinin (PHA). Cells so stained were examined in a flow cytometer with orthogonal illumination at 633 nm from a 7 mW He‐Ne laser; antibody‐bearing cells were detectable by fluorescence emission above 665 nm. Cells from the same cultures were stained with fluorescein‐labeled 4F2 antibody and examined in a flow cytometer with argon ion laser excitation at 488 nm. Percentages of antibody‐bearing cells determined from APC fluorescence and from fluorescein fluorescence were in good agreement. It thus appears that He‐Ne lasers and APC‐antibodies are usable for immunofluorescence measurements; the sensitivity attainable with this technique remains to be determined.
Separation and analysis of human chromosomes by combined velocity sedimentation and flow sorting applying single‐ and dual‐laser flow cytometry
Authors: Collard J. G. , Philippus E. , Tulp A. , Lebo R. V. , Gray J. W. 
Published: August 2005

AbstractHuman chromosomes were separated on basis of size by velocity sedimentation at 52g in a specially designed sedimentation chamber. The chamber has been constructed in such a way that large numbers of chromosomes can be fractionated on a sucrose gradient while wall sedimentation, streaming, and swirling movements of the gradient during centrifugation are eliminated. Flow deflectors in the chamber allow undisturbed introduction and fractionation of the density gradient. The different chromosomal fractions obtained are highly enriched for the various human chromosomes. Individual chromosomes were subsequently sorted to purity by fluorescence activated flow sorting using a FACS IV flow sorter equipped with a 4‐W argon‐ion laser. Following this procedure, the sorting rate for specific chromosomes can be speeded up by a factor of 5–10 when the pre‐enriched chromosomal fractions are used as starting material. A high chromosome resolution could be obtained by a few simple modifications of the FACS IV. By applying fluorochromes with different DNA‐base specificity the sorting possibilities of individual human chromosomes can be improved. In addition, the chromosomal fractions were analysed by dual laser flow cytometry after staining the chromosomes with Hoechst 33258 and chromomycin A3. In this way the enrichment of virtually all individual human chromosomes in the different chromosomal fractions can be visualized.
Bivariate flout cytometric analysis of murine intestinal epithelial cells for cytokinetic studies
Authors: Pallavicini M. G. , Ng C. R. , Gray J. W. 
Published: August 2005

AbstractThe heterogeneous nature of the small intestine and the lack of methods to obtain pure crypt populations has, in the past, limited the application of standard flow cytometric analysis for cytokinetic studies of the proliferating crypts. We describe a flow cytometric technique to discriminate crypt and villus cells in an epithelial cell suspension on the basis of cell length, and to measure the DNA content of the discriminated subpopulations. Our data indicate that bivariate analysis of a mixed epithelial cell suspension can be used to distinguish mature villus cells, G1 crypt cells, and S‐phase crypt cells. In addition, continuous labeling studies suggest that the position of a cell on the cell length axis reflects epithelial cell maturity. We applied this flow cytometric technique to determine the cvtokinetic nature of epithelial cells obtained by sequential digestion of the small intestine.
A Computer algorithm for the analysis of protein distribution in budding yeast
Authors: Martegani Enzo , Vanoni Marco , Delia Domenico 
Published: August 2005

AbstractFlow cytometry gives relevant data on cellular parameters such as DNA, RNA, and protein contents of individual cells and is therefore a powerful tool for analyzing microbial population dynamics. Relevant information about growth dynamics may be obtained from protein distribution. In fact, protein distribution is related to age distribution and depends on the law of growth of the population and the law of growth of the single cell. To extract the available information from protein distribution, we developed a computeralgorithm starting from a model for growth of Saccharomyces cerevisiae. This algorithm quantitatively fits experimental protein distributions, allows a deconvolution of these distributions, and thus yields information about temporal parameters of the cell cycle and structure of yeast populations.
Processing of staphylococcus aureus and zymosan particles by human leukocytes measured by flow cytometry
Authors: Bassϕe Carl‐Fredrik 
Published: August 2005

AbstractStaphylococcus aureus were labeled with fluorescein‐isothiocytanate (FITC), stained by ethidium bromide (EB), and measured by flow cytometry (FCM). Bacteria were identified by their FITC‐fluorescence and discriminated from the leukocyte cell nuclei by the much higher EB fluorescence and lower coefficient of variation of the latter. Following phagocytosis, both the bacterial FITC‐and EB‐fluorescence decayed. The mean FITC‐fluorescence was reduced about 20% after 15 min and 30–50% after 60 min. Zymosan particles were labeled by FITC and incubated with leukocytes for 15 min. Phagocytes were sorted by FCM and the zymosan particles were liberated by sonication. Their forward angle light scatter was reduced by about 50.6 ± 2.1% and the FITC‐fluorescence by 8.7 ± 1.0% (mean ± SD). The reduced FITC‐fluorescence and light scatter suggests degradation of proteins, and the decay of EB‐fluorescence degradation of DNA, but the specificity remains to be established. By this method phagocytes from a patient with systemic lupus erythematosus seemed to have a selective defect in DNA degradation, whereas phagocytes from a patient with acute myelogenous leukemia had a low capacity to degrade bacterial proteins. This technique offers opportunities for automatic measurements of bacteria and zymosan particle degradation by phagocytes.
Three‐color immunofluorescence analysis of mouse B‐lymphocyte subpopulations
Authors: Parks D. R. , Hardy R. R. , Herzenberg L. A. 
Published: August 2005

AbstractWe have modified a fluorescence‐activated cell sorter (FACS) to make three independent immunofluorescence measurements on each cell and used this system to study mouse B‐lymphocyte subpopulations. An argon‐ion laser (emitting at 488 nm) excites fluorescein‐and phycoerythrin‐labeled reagents, and a tunable dye laser charged with rhodamine 6G (emitting at 615 nm) excites an allophycocyanin‐labeled reagent. We report simultaneous measurements of IgM, IgD, and the recently‐defined mouse B lymphocyte antigens BLA‐1 and BLA‐2 on splenic lymphocytes of CBA/J mice and mice of the congenic strain CBA/N (which have an X‐linked immunodeficiency [xid]). These data provide information on relationships among the B‐cell populations in CBA/J “normal” mice and the defective CBA/N that could not be derived from one‐ or two‐ color immunofluorescent measurements. We believe this is the first use of allophycocyanin as an immunofluorescence label.
Differential binding of lectins to lymphopoietic and myelopoietic cells in murine marrow as revealed by flow cytometry
Authors: Reimann J. , Ehman D. , Miller R. G. 
Published: August 2005

AbstractMurine marrow cells were labeled with a panel of fluorescein‐conjugated lectins and analyzed by flow cytometry. All lectins tested [concanavalin A (Con A), phytohemagglutinin (PHA), wheat germ agglutinin (WGA), pokeweed mitogen (PWM), soybean agglutinin (SBA), peanut agglutinin (PNA), lentillectin (LL)], stained a fraction of marrow cells; the fraction was, over a defined concentration range, linearly related to the logarithm of the lectin concentration. The slope of this titration curve was characteristic of the particular lectin tested but independent of the nominal sugar specificity of the respective lectin. Con A‐, PWM‐, and PNA‐labeled marrow cells showed brightly and dimly stained subsets in relative fluorescence intensity distribution histograms at high or intermediate staining concentrations. These subsets were composed of cells with restricted and characteristic size distributions as determined by forward light scatter. Lectin staining patterns of marrow cells from normal mice were compared with those of athymic nude mice and mice with severe combined immunodeficiency (SCID). Normal and nude mice lacked a subset of small cells, dimly stained by Con A and PWM, but brightly stained by PNA. This subset, uniquely defined by the above lectins, appears to correspond to the lymphopoietic marrow compartment.
Fluorescence energy transfer measurements on cell surfaces: A critical comparison of steady‐state fluorimetric and flow cytometric methods
Authors: Szöllósi János , Trón Lajos , Damjanovich Sándor , Helliwell Stephen H. , Arndt‐Jovin Donna , Jovin Thomas M. 
Published: August 2005

AbstractThe energy transfer efficiency E was measured between fluorescein‐conjugated concanavalin A (Con A) and rhodamine‐conjugated Con A bound to homogeneous tissue culture cells, the HK22 murine lymphoma cell line. Results from a flow cytometric energy transfer method (FCET) and two different steady‐state fluorimeter methods were compared. The data were found to be in close agreement after careful correction of the steady‐state fluorimetric measurements for contributions from dissociating ligand. The biological variability of the individual cells with respect to E was calculated using an error propagation analysis and were found to be less than the variability in the absolute amount of ligand binding per cell. FCET has a number of advantages over the fluorimetric measurements using suspensions of cells: (1) relatively labile receptor‐ligand complexes can be measured; (2) the analysis can be restricted to undamaged cells by gating the data collection on the light‐scattering signals; (3) heterogeneous populations of cells with respect to donor and acceptor topology can be distinguished by the correlation of E with other cellular parameters derived from additional signals or combinations thereof; and (4) the dynamics of donor‐acceptor redistribution on subpopulations can be measured.
Increased accuracy and speed of absorption cytometric DNA measurements by automatic corrections for nuclear darkness
Authors: Allison David C. , Lawrence George N. , Ridolpho Paul F. , O'Grady Brian J. , Rasch Robert W. , Rasch Ellen M. 
Published: August 2005

AbstractWe have developed a method of calculating the average local absorbance (ALA) of a nucleus from the integrated nuclear absorbance and area. One can use the ALA, along with nuclear areas measured at different point absorbance thresholds, to determine whether a nucleus is stained too lightly or too darkly for accurate absorption measurements; this allows selection of an optimal light wavelength for the performance of these measurements. The ALA can also be used for automatic and instantaneous correction of integrated absorbance values from darkly stained cells. This allows the rapid measurement of the integrated absorbances of a large number of nuclei that are heterogeneous in stain intensity. Coefficients of variation of approximately 3% are obtained for the integrated absorbances of nuclei of nontransformed G0/G1 cells. This correction method can be applied with any image densitometer that generates both integrated absorbance and area values.
Estimation of sampling errors in a high‐resolution TV microscope image‐processing system
Authors: Harms H. , Aus H. M. 
Published: August 2005

AbstractThe basic postulate of this paper is that the commonly accepted sampling density of 2–4 pixels/μm in a high‐resolution TV microscope system is too low to digitize exactly and analyze the complex cellular detail found in stained cell images. Depending on the specific microscope system, the required sampling density is much higher, lying between 15 and 30 pixels/μm. This sampling density is derived from the aliasing error, the resolution loss, and computational limitations. The mathematical and optical methods and equipment used to obtain these results are described in detail.
Image processing for mitoses in sections of breast cancer: A feasibility study
Authors: Kaman E. J. , Smeulders A. W. M. , Verbeek P. W. , Young I. T. , Baak J. P. A. 
Published: August 2005

AbstractThis paper describes an image analysis technique for the counting of nuclei in mitosis in tissue sections. Five experienced pathologists scored mitoses in photographs of preselected areas of tissue sections of the breast. Objects consistently labelled as mitotic cells by all five pathologists were considered “mitoses” in the analysis. In total, there were 45 mitotic nuclei, 68 possible mitotic nuclei and 1,172 nonmitotic nuclei.The image analysis procedure was designed to give priority to a low false negative rate, i.e., misclassification of mitoses. The procedure consists of three steps:
Segmentation of the image.
Reduction of the number of nonmitotic nuclei by using feature values based on the brightness histogram of the objects.
Fully automatic classification of the remaining objects using contour features.
The objects remaining after the first two steps were visualized in a composite display for interactive evaluation: 10% of the mitotic nuclei were missed, and 85% of the nonmitotic nuclei were eliminated. The result of the fully automatic procedure described in this paper is rather disappointing and gave a loss of 37% of the mitoses while 5% of the nonmitotic nuclei remained.
An automated image analysis system for the detection of rare autoradiographically labelled cells in the human lymphocyte HGPRT variant assay
Authors: Stark Margaret H. , Tucker J. H. , Thomson E. J. , Perry P. E. 
Published: August 2005

AbstractThis paper describes a fast and accurate method for the identification of autoradiographically labelled human hypoxanthine‐guanine phosphoribosyl transferase (HGPRT) variant lymphocytes on slide preparations using a high‐speed computer image analysis system–the Fast Interval Processor (FIP). The system has been developed primarily for the analysis of cultured human peripheral blood lymphocyte populations in which the frequency of labelled cells may vary from less than one to more than 2,000 in every 10,000 cells. Evaluation experiments have demonstrated the excellent performance of FIP in the counting of labelled and total cells over this frequency range.
Nuclear DNA cytophotometry in prostate carcinoma
Authors: Seppelt U. , Sprenger E. 
Published: August 2005

AbstractEighty patients with prostate carcinoma underwent fine‐needle aspiration biopsy for cytologic grading and DNA‐single‐cell fluorescence photometry before and at 6‐month intervals after endocrine treatment. The histograms of DNA values showed single peaks and bimodal and scattered distribution which correlated to the different tumor grades before therapy. The DNA values were significantly different from the controls with benign prostatic hypertrophy. After start of therapy, regressive changes of the DNA‐histograms were increases of diploid and hypodiploid DNA values and disappearance of secondary peaks. Progressive changes were increased scattering of DNA values and appearance of secondary peaks. Progressive changes in the histograms were closely related to clinical remission and stable disease, but related poorly to clinical progression. The survival correlated with the pretherapeutic DNA‐histograms and with the DNA‐median, third‐quartile, and maximum parameters.
Comparative flow DNA analysis of different cell suspensions in breast carcinoma
Authors: Chassevent A. , Daver A. , Bertrand G. , Coic H. , Geslin J. , Bidabe M.‐CL , George P. , Larra F. 
Published: August 2005

AbstractThis study compared three methods of dissociation of breast lesions for DNA flow cytometry. Eleven benign lesions and 66 cancers were dissociated using mechanical, Ficoll, or enzymatic methods. DNA flow analysis showed that the DNA index did not vary from one method of dissociation to another. All benign lesions were diploid and 67% of all cancers were aneuploid. Enzymatic dissociation gave a lower percentage of aneuploid cells with a diminution of the proportion of cells in the G2 + M phase (13.2% enzymatic against 17.6% Fi‐coll); on the other hand, it provided cell populations of greater viability than the other methods (32.6% enzymatic, 17.2% Ficoll; P < 0.01). The mechanical and Ficoll suspensions did not differ significantly when they were analyzed on the basis of their DNA content and their cellular viability. When compared with mechanical preparation, Ficoll suspension showed a lower recovery of tumor cells, but this inconvenience was compensated for by a more homogenous aspect where the contribution of aggregates and debris was clearly lessened. Therefore, this study led us to choose Ficoll suspension for subsequent flow analysis of breast tumors.