bluuebunny/crossref_metadata_2025_split huggingface dataset:
Correlated measurement of platelet release and aggregation in whole blood
Authors: Ault Kenneth A. , Rinder H. M. , Mitchell J. G. , Rinder C. S. , Lambrew C. T. , Hillman R. S. 
Published: August 2005

AbstractWe have used a technique for the simultaneous measurement of platelet activation and aggregation in whole blood using two‐color immunofluorescence and flow cytometry to study the relationship between the release reaction and aggregation. A monoclonal antibody specific for the alpha granule membrane protein GMP‐140 was used to measure the release reaction, and a monoclonal antibody specific for platelet membrane glycoprotein Ib (GPIb) was used to identify platelets and platelet aggregates. Aggregates were identified as particles expressing both levels of GPIb and size larger than that of resting single platelets. Anticoagulated whole blood was incubated with platelet agonists. At various times samples of the blood were removed and immediately fixed with paraformaldehyde. Blood that had been anticoagulated with ethylenediamine tetraacetic acid showed progressive release of platelets but little or no aggregation. However, blood anticoagulated with citrate or heparin showed correlated release and aggregation. The degree of aggregation was greater in heparin than in citrate. The expression of GPIb and GMP‐140 increased in direct proportion to the size of the aggregates. Aggregates were observed varying in apparent diameter up to approximately 20 μm. During prolonged incubation there was progressive disaggregation of adenosine diphosphate (ADP)‐induced aggregates. After disaggregation the proportion of GMP‐140 negative single platelets increased, indicating that both released and nonreleased platelets participated in the aggregation. There was little or no disaggregation of phorbol myristate acetate (PMA)‐induced aggregates. The relatively small size and reversibility of platelet aggregates that we have observed in whole blood may be relevant to phenomena occurring in vivo and in extracorporeal circulation. The ability to measure such aggregates both in vitro and in vivo may be very useful.
A rapid and sensitive flow cytometric method for the detection of multidrug‐resistant cells
Authors: Herweijer Hans , Engh Ger Van Den , Nooter Kees 
Published: August 2005

AbstractMultidrug‐resistant (MDR) cells are characterized by a defect in drug accumulation caused by activity of an energy‐dependent rapid drug efflux pump. The action of this drug pump can be inhibited by specific agents, referred to as membrane transport modulating agents (MTMAs), resulting in a restoration of the intracellular drug accumulation. This paper presents a flow cytometric assay for the detection of MDR cells, which is based on the ability of these cells to respond to MTMAs. Daunorubicin net‐uptake kinetics were measured of anthracycline‐sensitive (A2780/S) and ‐resistant (A2780/R) human ovarian carcinoma cells in vitro. A2780/R cells accumulated significantly less (about a factor of 5) daunorubicin as compared to A2780/S cells. Addition of verapamil or cyclosporin A to A2780/R cells at steady‐state daunorubicin uptake led to a dose‐dependent increase in cellular daunorubicin accumulation. The sensitivity of the assay was determined by testing mixtures of A2780/S and A2780/R cells. Analysis of A2780/S cells contaminated with A2780/R cells showed that as few as 2.5% MDR cells could readily be detected in the mixture. In conclusion, this functional assay enables the detection of MDR cells in a heterogeneous cell suspension and is ideally suited for the study of the occurrence of typical MDR in human cancer.
Quantitative immunofluorescence in single Saccharomyces cerevisiae cells
Authors: Eitzman P. D. , Hendrick J. L. , Srienc F. 
Published: August 2005

AbstractWe have developed a staining procedure that allows the simultaneous determination of intracellular amounts of DNA and an antigen in Saccharomyces cerevisiae with a single laser flow cytometer. The antigen, β‐galactosidase from a cloned lacZ gene, is inducible and is detected with an indirect immunofluorescent stain. Cell preparation procedures, specifically cell fixation and cell wall removal, have significant effects on measured levels of immunofluorescence and have been optimized to prevent cell loss and maximize immunofluorescence. Average immunofluorescent levels of cell populations expressing different levels of β‐galactosidase show excellent correlation with measurements of average β‐galactosidase activity per cell based on cleavage of o‐nitrophenyl‐β‐D‐galactopy‐ranoside. Experiments with yeast populations containing various numbers of copies of the cloned gene indicate that the relationship between immunofluorescence and antigen content also holds at the single‐cell level. Correlated measurements of DNA and β‐galactosidase content on a single‐cell level permit the investigation of cellular enzyme content as a function of cell cycle position under various conditions. The procedure can be easily modified to detect other antigens by changing the primary antibody used.
Flow cytofluorimetric determination of protein distribution throughout the cell cycle
Authors: Khochbin Saadi , Chabanas Agnès , Albert Philippe , Lawrence Jean‐Jacques 
Published: August 2005

AbstractQuantitative treatment of multiparametric determination on cells using flow cytofluorometry was made possible by the development of a computer program that allows the relative quantification of a specific protein as a function of the position of cells during the cell cycle. This type of analysis provides interesting information about the distribution of a given protein throughout the cell cycle. Four examples showing the distribution of specific proteins illustrate such a quantification during the cell cycle in two different cell lines. The program also allows for the handling of a series of histograms obtained by the analysis of protein distribution as a function of DNA content in relation to a third parameter. To illustrate possible applications for this program, the evolution of the distribution of two proteins, the oncoprotein p53 and the histone H1°, during the induced differentiation of murine erythroleukemia cells has been studied.
Flow cytometry and phytoplankton
Authors: Phinney D. A. , Cucci T. L. 
Published: August 2005

AbstractFlow cytometry and sorting are now an important technology in aquatic research. Simultaneous measurements of individual particle cell size, fluorescence, and light scatter properties are directly applicable to current topics in aquatic research. Flow sorting may be employed to obtain subsets of cells for analysis by conventional methods. The manner in which rapid, precise measurements of single cells are made is complex, and the application of this technology to aquatic samples is subject to many analytical constraints. Flow cytometric measurements of algal cell size and pigment autofluorescence are relative and are therefore dependent on the optical configuration and variability of the instrument. Specific types of reference materials are used to establish the validity of analyses: (1) instrument standards, (2) fluorescence controls, and (3) internal stain standards. The selection and application of standards and controls are discussed in the context of allometric (cell size versus pigment fluorescence) and ataxonomic (pigment color groups) methods. The widespread acceptance of particular reference materials among research groups will result in comparable data sets describing aquatic particle distributions.
Separation and concentration of phytoplankton populations using centrifugal elutriation
Authors: Pomponi Shirley A. , Cucci Terry L. 
Published: August 2005

AbstractCentrifugal elutriation is a technique for separating particles on the basis of their sedimentation velocity, an expression of size, shape, and specific gravity. Unialgal cultures, mixtures of two phytoplankton cultures, and natural seawater samples were elutriated to determine the feasibility of this technique for collecting fractions of different cell cycle phases, separating two phytoplankton species, and concentrating cells from dilute samples. Elutriation resulted in the separation of a culture of Dunaliella tertiolecta and Phaeodactylum tricornutum into homogeneous fractions of each species. Cells in the natural seawater sample were concentrated by nearly 2 orders of magnitude. Centrifugal elutriation provides an alternative cell separation and concentration technique when large numbers of cells are required.
Photobiology of natural populations of zooxanthellae from the sea anemone Aiptasia pallida: Assessment of the host's role in protection against ultraviolet radiation
Authors: Lesser Michael P. 
Published: August 2005

AbstractNatural populations of the sea anemone Aiptasia pallida containing endosymbiotic dinoflagellates were acclimated to different irradiance regimes, with and without ultraviolet radiation (UV). They showed a compensatory response in the amount of chlorophyll and the activities of enzymes responsible for detoxifying active species of oxygen that are produced by the interaction between visible or ultraviolet radiation and photosynthetically produced oxygen. Protection from active species of oxygen is essential to prevent the photooxidation of chlorophyll and the concomitant loss of productivity. Bulk analyses of chlorophyll showed differences between the populations exposed to varying irradiance regimes, but revealed no significant independent effect of UV. However, analysis by flow cytometry of the individual cells from treated populations did detect statistically significant differences in cell size and the amount of chlorophyll fluorescence per cell, which could be attributed to treatment with ultraviolet radiation. With flow cytometry we are able to detect the population variability that is undetectable by bulk measurements which is important in assessing the effects of environmental parameters in both symbiotic and free‐living microalgae. Research using simultaneous measurements by flow cytometry could add considerable insight into the population dynamics of both zooxanthellae and host cells.
A simple and rapid method for determining the linearity of a flow cytometer amplification system
Authors: Bruce Bagwell C. , Baker David , Whetstone Sherry , Munson Mark , Hitchcox Shelly , Ault Kenneth A. , Lovett Edmund J. 
Published: August 2005

AbstractWe describe a simple and rapid method for determining the linearity of a flow cytometer amplification system. The method is based on a fundamental characteristic of linear amplifiers: The difference between two amplified signals increases linearly with increasing amplifier gain. Two populations of beads or cells, differing slightly in fluorescence intensity, are analyzed by the flow cytometer at increasing photomultiplier tube high‐voltage settings. The distribution of the populations' mean difference versus mean position is a straight line intersecting the origin for linear amplifiers. Although some types of nonlinearities cannot be detected with this technique, deviations from linearity indicate nonlinear components in the flow cytometer amplification system. The correlation coefficient is used to quantify degree of nonlinearity. We also describe a method for amplifier nonlinearity compensation.
Statistical evaluation of cell kinetic data from DNA flow cytometry (FCM) by the EM algorithm
Authors: Baldetorp Bo , Dalberg Mats , Holst Ulla , Lindgren Georg 
Published: August 2005

AbstractFlow cytometric DNA measurements yield the amount of DNA for each of a large number of cells. A DNA histogram normally consists of a mixture of one or more constellations of G0/G1‐, S‐, G2/M‐phase cells, together with internal standards, debris, background noise, and one or more populations of clumped cells. We have modelled typical DNA histograms as a mixed distribution with Gaussian densities for the G0/G1 and G2/M phases, an S‐phase density, assumed to be uniform between the G0/G1 and G2/M peaks, observed with a Gaussian error, and with Gaussian densities for standards of chicken and trout red blood cells. The debris is modelled as a truncated exponential distribution, and we also have included a uniform background noise distribution over the whole observation interval. We have explored a new approach for maximum‐likelihood analyses of complex DNA histograms by the application of the EM algorithm. This algorithm was used for four observed DNA histograms of varying complexity. Our results show that the algorithm works very well, and it converges to reasonable values for all parameters. In simulations from the estimated models, we have investigated bias, variance, and correlations of the estimates.
Detection of bladder carcinoma in females by flow cytometry and cytology
Authors: Hermansen Dane K. , Badalament Robert A. , Fair William R. , Kimmel Marek , Whitmore Willet F. , Melamed Myron R. 
Published: August 2005

AbstractThe sensitivity of bladder wash flow cytometry (BWFCM), voided urinary cytology (VUC), and cytology of cathetrized urine obtained at the time of cystoscopy (CUC) were reviewed on all women evaluated for bladder cancer at Memorial Sloan‐Kettering Cancer Center between June 1985 and December 1986. This comprised sixty‐four episodes of pathologically proven bladder cancer in 48 women. Considering positive and suspicious results jointly the sensitivities of BWFCM, CUC and 3 VUC were 75%, 64% and 56%, respectively. If only positive results were considered (i.e., suspicious results considered as negative), the sensitivities of BWFCM, CUC and 3 VUC were 64%, 31% and 32%, respectively. The sensitivities of these tests are less than for a predominantly male population, presumably related to the presence of squamous epithelium and greater frequency of pyuria. However, bladder wash flow cytometry and conventional cytology are still a very valuable addition to cystoscopic examination, and the combination of BWFCM with conventional cytology is more sensitive than either procedure alone.
Separation of pigmented and albino melanocytes and the concomitant evaluation of endogenous peroxide content using flow cytometry
Authors: Boissy Raymond E. , Trinkle Linda S. , Nordlund James J. 
Published: August 2005

AbstractFlow cytometry (FCM) has been used extensively to analyze various biological properties of the cell. In this report, we describe a method by which FCM was used to determine the light scattering profile of a mixed population of pigmented and non‐pigmented melanocytes, plus its subsequent use for the sorting and separation of the two cell types. In addition, the relative peroxide content in pigmented and non‐pigmented melanocytes was compared by flow cytometry.Cultured avian melanocytes from a pigmented control and from three genetically distinct albino sources were studied. FCM analysis of forward versus side light scatter within a mixed suspension of pigmented and amelanotic melanocytes distinguished two overlapping populations of cells. Sorting of these two populations demonstrated that the population exhibiting much side and minimal forward light scatter was primarily pigmented melanocytes, while conversely the population exhibiting less side and more forward scatter was principally non‐pigmented cells. These two melanocyte types also demonstrated differences in levels of endogenous peroxides. The intracellular content of peroxide in the two subpopulations of cells was measured utilizing the nonfluorescent compound, 2′,7′‐dichlorofluorescin diacetate (DCFH‐DA), which within the cell is oxidized by intracellular peroxides to a fluorescent dichlorofluorescein (DCF). Non‐pigmented albino melanocytes had the highest quantity of endogenous peroxides, while heavily pigmented cells had considerably less peroxide‐related fluorescence. The amount of this DCF fluorescence could be enhanced by increasing concentrations of DCF used in the assay.These flow cytometric methods are useful for isolating and culturing subpopulations of melanocytes expressing various pigment levels and to investigate the relationship between melanin and its precursors with hydrogen and lipid peroxides in melanocytes.
Sample station modification providing on‐line reagent addition and reduced sample transit time for flow cytometers
Authors: Kelley Keith A. 
Published: August 2005

AbstractA flow cytometer was equipped with a modified sample station to facilitate online addition of mediators to the sample and reduce the time of delivery of the sample to the interrogation point. The ready availability of materials and straightforward nature of this design make modifications simple and facilitate measurements of cellular activation. Parameters such as pH, membrane fluidity, and calcium mobilization are easily measured in this system, because detection can be made less than 4 s after addition of mediator with no interruption of sample flow. The sample station modification is described in detail along with methods for mixing and temperature control.
Fluorescent staphylococci as microbeads
Authors: Szabó Gábor , Damjanovich Sándor 
Published: August 2005

AbstractFixed bacteria of the protein A‐rich Cowan I Staphylococcus strain were labelled with fluorescein isothiocyanate and used as the second‐step reagent in an indirect immunofluorescent assay of specific cell‐surface antigen expression. The results are documented with fluorescence microscopy and flow cytometry.
The athena semi‐automated karyotyping system
Authors: van Vliet Lucas J. , Young Ian T. , Mayall Brian H. 
Published: August 2005

AbstractIn this article we describe Athena, a system that provides for semi‐automated karyotyping of metaphase spreads. The system is based upon the Macintosh II computer. It uses software that is written entirely in C and consists of approximately 200 Kbytes of executable code. Athena provides automated segmentation of metaphase images into individual chromosomes, automated measurements on each banded chromosome, and automated classification into the standard Paris‐convention karyotype. Furthermore, the system provides the ability to construct one or more chromosome data bases to represent the types of metaphase spreads and staining techniques that may be used in a given laboratory. Because we believe that it is impossible to construct a system that can achieve perfect segmentation, perfect separation of touching and overlapping chromosomes, perfect localization of the centromeres, and perfect classification, the system offers the possibility for interaction at each of the above stages using the well‐accepted Macintosh user interface.
Multivariate chromosome analysis and complete karyotyping using dual labeling and fluorescence digital imaging microscopy
Authors: Arndt‐Jovin Donna J. , Jovin Thomas M. 
Published: August 2005

AbstractThe combination of multiple dye‐DNA interactions, a fluorescence digital imaging system with a scientific CCD camera, and multivariate image analysis allows the rapid karyotyping of fluorescent human metaphase chromosome spreads. Chromosomes are stained with the bisbenzimidazole dye Hoechst 33342 and chromomycin A3, a dye pair used frequently in bivariate flow analysis and sorting of metaphase chromosomes in suspension. The use of ratio functions involving the total and peak intensities of the two dyes provides increased resolution of the karyotype in the microscope, and it can be anticipated that the same approach could lead to improved performance with flow systems as well. High pass filtering with a Laplace operator yields characteristic banded images of the individual chromosomes, even with total fields that are less than 200 pixels on a side.
Discrepancies between flow cytometric and cytogenetic studies in the detection of aneuploidy in human solid tumors
Authors: Shackney Stanley E. , Burholt Dennis R. , Pollice Agnese A. , Smith Charles A. , Pugh Reginald P. , Hartsock Robert J. 
Published: August 2005

AbstractParallel flow cytometric (FCM) cell DNA studies and cytogenetic studies were performed on clinical samples from twenty human solid tumors of various types and on cell lines established in tissue culture from three of these tumors. Six of twenty clinical samples (30%) showed concordance between flow cytometry and cytogenetics with respect to the presence or absence of aneuploidy. Among the fourteen cases with discrepancies between the two methods, 8 (40% of all cases) showed hypodiploidy by cytogenetics and had diploid DNA histograms. Three cases (15%) had prominent discrete peaks in the triploid to tetraploid region by cytogeneties but had only barely discernible corresponding peaks in the DNA histogram. In two cases (10%)cytogenetic studies revealed diffuse aneuploidy. Cytogenetic studies demonstrated near‐tetraploidy in three samples, but only one of these was detected by FCM; all three cases exhibited other numerical chromosomal abnormalities. In one case aneuploidy was demonstrated by FCM and not by cytogenetics. Among the tumor cell lines established in culture, the DNA Index was often higher than the cytogenetic index. Overall, 13/20 or 65% of patients with solid tumors in this, study had numerical chromosomal abnormalities that were not detected by flow cytometry. Eleven of these patients had distant metastases at the time of tumor sampling, and nine of these died of their disease within 1–11 months of the time of study.
Rapid metaphase and interphase detection of radiation‐induced chromosome aberrations in human lymphocytes by chromosomal suppression in situ hybridization
Authors: Cremer T. , Popp S. , Emmerich P. , Lichter P. , Cremer C. 
Published: August 2005

AbstractChromosomal in situ suppression (CISS)‐hybridization of biotinylated phage DNA‐library inserts from sorted human chromosomes was used to decorate chromosomes 1 and 7 specifically from pter to qter and to detect structural aberrations of these chromosomes in irradiated human peripheral lymphocytes. In addition, probe pUC1.77 was used to mark the Iq12 subregion in normal and aberrant chromosomes 1. Low LET radiation (60Co‐γ‐rays; 1.17 and 1.33 MeV) of lymphocyte cultures was performed with various doses (D = 0, 2, 4, 8 Gy) 5 h after stimulation with phytohaemagglutinin. Irradiated cells were cultivated for an additional 67 h before Colcemid arrested metaphase spreads were obtained. Aberrations of the specifically stained chromosomes, such as deletions, dicentrics, and rings, were readily scored after in situ hybridization with either the 1q12 specific probe or DNA‐library inserts. By the latter approach, translocations of the specifically stained chromosomes could also be reliably assessed. A linear increase of the percentage of specifically stained aberrant chromosomes was observed when plotted as a function of the square of the dose D. A particular advantage of this new approach is provided by the possibility to delineate numerical and structural chromosome aberrations directly in interphase nuclei. These results indicate that cytogenetic monitoring of ionizing radiation may be considerably facilitated by CISS‐hybridization.