bluuebunny/crossref_metadata_2025_split huggingface dataset:
A hematology data‐logging terminal
Authors: Paquette Gerard A. , Sparks Thomas E. 
Published: August 2005

AbstractIntended as a link between a central analysis facility and remote microscope analyzers at international sites, a data‐logging terminal has been developed which exploits basic analyzer control and data acquisition capabilities. The terminal depends on the analyzer's ability to assimilate, interpret, and respond to external commands transferred through its standard communications interface. Within the limits of the analyzer's high‐level control language, the data‐logging terminal is able to direct microscope operation and data acquisition as well as to initiate algorithms. Internal instrument algorithms can then establish and maintain focus, acquire specified objects, direct various scene segmentation sequences, and execute feature extractions. The data‐logging terminal can thus command the analyzer to find scenes of interest and extract data from them. User access to a limited instrument operating interface is provided through its keyboard and alphanumeric display. The terminal can also request and receive image, feature, or count data over the communications interface. Such data are recorded, together with identifiers entered through the keyboard, onto diskettes which may be transferred by mail or telecommunications for processing with the image analysis and pattern recognition techniques available at the central analysis facility. Anticipated applications include evaluation of sample‐preparation techniques, correlation of image data with sampling conditions, training of special classifiers, and analysis of instrument performance.
Effects of hoechst 33342 on survival and growth of two tumor cell lines and on hematopoietically normal bone marrow cells
Authors: Fried Jerrold , Doblin Jeffrey , Takamoto Shigeru , Perez Amaury , Hansen Herbert , Clarkson Bayard 
Published: August 2005

AbstractOur objective in this investigation was to determine whether Hoechst 33342, which is widely recognized as a DNA specific fluorochrome for living cells, is in fact nontoxic and kinetically nonperturbing at dye concentrations required to achieve acceptable DNA distributions. Three cell types were tested: HeLa S‐3; SK‐DHL2, a human lymphoma cell line; and hematopoietically normal human bone marrow cells. In the third system, only the cloning efficiencies were determined. Results differed considerably for the different cell types. While HeLa cells yielded excellent DNA distributions and were almost completely resistant to the cytotoxic and cytokinetic effects of the dye, SK‐DHL2 cells were highly sensitive to the fluorochrome even at dye concentrations which produced very poor DNA distributions. Human bone marrow cells were intermediate in their stainability and toxicity response, and acceptable DNA distributions could be obtained at the nontoxic dye concentration of 2.5 μM. Clearly, different cell types differ considerably with respect to their cytotoxic and kinetic responses to Hoechst 33342. In some cases it may not be possible to ensure adequate staining of the cells for flow cytometry without significantly altering their viability and/or proliferative behavior.
Comparative kinetics of poly (6‐thioinosinic acid) and 6‐mercaptopurine treated L1210 cells
Authors: Tomasovic Stephen P. , Higashikubo Ryuji , Cohen Arthur M. , Broom Arthur D. 
Published: August 2005

AbstractPoly(6‐thioinosinic acid), poly(s6I), a synthetic polynucleotide shown to inhibit L1210 ascites growth in vivo, was compared with 6‐mercaptopurine (6‐MP) for their effects on cellular kinetics of L1210 cells in vitro. These kinetic perturbations were monitored by changes in growth rates, cell volumes and cell‐cycle distributions. At equimolar exposure doses (based on molar concentration of purine bases), 0.1 mM poly(s6I) was more effective than 6‐MP in reducing growth rate and total cell number. A 20‐fold increase in the 6‐MP exposure dose (to 2 mM) was required to reduce total cell number similar to 0.1 mM poly(s6I). Concomitant determination of Coulter volume distributions of treated cell populations indicated 0.1 mM poly(s6I) increased the mean population volume more than either concentration of 6‐MP. Cell‐cycle distributions obtained by flow cytometry showed that, simultaneously with the increase in mean population volume, poly(s6I) exposure caused a decrease of cells in G1 and S and a pronounced accumulation of cells in G2. Treatment with 6‐MP caused similar cell‐cycle redistributions but always less than for poly(s6I). Flow sorting of these redistributed populations indicated the increase in mean population volume during treatment with 0.1 mM poly(s6I) was almost equally due to an increased percentage of G2 cells and to an increased volume of cells with the G2 DNA content. However, with 0.1 mM 6‐MP, increased volume of G2 cells was predominant. Although poly(s6I) was synthesized in an attempt to develop more effective delivery forms of biologically active compounds like 6‐MP and it has close structural similarities to 6‐MP, its mechanism of action is unknown.
A polycarbonate filter technique for collection of sorted cells
Authors: Folstad L. , Look M. , Pallavicini M. 
Published: August 2005

AbstractWe describe the application of polycarbonate filters to the collection of cells sorted by flow cytometry. Cells are sorted directly onto polycarbonate filters and transferred to microscope slides which are then processed to dissolve the filter and remove pore outlines. The technique results in cell preparations which retain cellular morphology and have high cell recovery of 82–100%.
Is the central dogma of flow cytometry true: That fluorescence intensity is proportional to cellular dye content?
Authors: Kerker M. , Van Dilla M. A. , Brunsting A. , Kratohvil J. P. , Hsu P. , Wang D. S. , Gray J. W. , Langlois R. G. 
Published: August 2005

AbstractMeasurements and theoretical calculations of fluorescent emission from four samples of polystyrene microspheres (diameter 0.92, 1.63, 1.90 and 4.18 m̈m) containing the same fluorescent dye show a general dependence upon particle size, emission angle, and polarization conditions. However, for the excitation and detection conditions used in flow cytometry, the relative fluorescent intensities measured for the four particle sizes are proportional to the dye content to +10% accuracy, independent of particle size. Accordingly, the central dogma of flow cytometry ‘that fluorescence is proportional to cellular dye content’ is valid to this accuracy for these solid, highly refractive polymer particles. Most mammalian cells are much less refractive, therefore, should conform more closely to the central dogma.
Rapid, one step staining procedures for analysis of cellular DNA and protein by single and dual laser flow cytometry
Authors: Crissman H. A. , Steinkamp J. A. 
Published: August 2005

AbstractDetailed, simplified techniques are described for simultaneous staining and analysis of DNA and protein in a number of mammalian cell types. Cell staining involves the addition of appropriate dye mixtures to unfixed or ethanol‐fixed cells and subsequent analysis of cell populations in the staining reagents generally within 10 to 20 minutes. The approach is novel in that no centrifugation steps are involved during the staining procedure, thus, eliminating cell clumping and cell loss and making the procedures appropriate for samples containing limited numbers of cells.For single wavelength analysis, staining of DNA and protein in ethanol‐fixed cells was accomplished with a dye solution containing propidium iodide, fluorescein isothiocyanate and RNase. After 20 minutes at room temperature cells were analyzed using the 488 nanometer (nm) laser excitation line. For dual laser analysis the following dye combinations were employed without RNase: mithramycin‐rhodamine 640, mithramycin‐substituted rhodamine isothiocyanate, Hoechst 33342‐rhodamine 640 and Hoechst 33342‐rhodamine isothiocyanate. Unfixed cells were also stained with the Hoechst 33342‐rhodamine 640 dye combination. Mithramycin was excited at 457.9 nm, Hoechst 33342 at 333–363 nm, and the rhodamine dyes at 568 nm.Cell types analyzed included Chinese hamster ovary cells, cultured mouse colon 26 cells, mouse embryo forelimb bud cells, and rat cell samples obtained by lung lavage.
Flow cytometry of human testicular tissue
Authors: Pfitzer Peter , Gilbert Peter , Rölz Gero , Vyska Karel 
Published: August 2005

AbstractFlow cytometry of testicular tissue from autopsies showed differences in the proportion of substained spermatozoa (1 CA), spermatids (1 CB), diploid 2C nuclei and premeiotic spermatozoa in the 4 C class. The evaluation of the histograms was performed with computer assistance. Healthy accident cases or those delivered in forensic medicine furnished the control values. Patients with hepatic cirrhosis showed a nearly complete loss of the haploid class, while cases with fatty degeneration of the liver demonstrated a reduction. Circulatory insufficiency in cases of acute shock resulted in an increase in the 4C values of meiotic prophase I and a decrease of spermatids. Spermatozoa were normal in these cases. After prolonged shock a nearly complete breakdown of spermatogenesis was observed. In cases of atherosclerosis an irregular degree of reduction was seen. Cancer patients showed a reduction of spermatogenesis in correlation to therapy.
Cell cycle operation during batch growth of fission yeast populations
Authors: Agar D. W. , Bailey J. E. 
Published: August 2005

AbstractBatch cultivation provides a continuous sequence of different environments useful for studying responses of cell cycle controls. Flow cytometry measurements have been made of the frequency functions for protein, RNA, and DNA at different times during batch growth of the fission yeast Schizosaccharomyces pombe. The mean cellular protein and RNA contents and their variances tend to increase with increasing population specific growth rates. Analysis of the midexponential phase DNA frequency function data indicates that DNA synthesis occupies 12% of the total cell cycle time and is completed at the same time as cell separation. Coordination of DNA synthesis and cell separation is less precise when population growth rate is low in late lag and early stationary phases.
The effect of some β‐lactam antibiotics on escherichia coli studied by flow cytometry
Authors: Martinez Octavio V. , Gratzner Howard G. , Malinin Theodore I. , Ingram Marylou 
Published: August 2005

AbstractThe effects of three beta‐lactam antibiotics on Escherichia coli were studied by means of flow cytometry. Since these agents block bacterial cell wall synthesis in such manner as to prevent septal formation without appreciably affecting nucleic acid synthesis, the resulting cell elongation caused by these agents can be assessed by nucleic acid fluorescent staining. It was shown by this technique that the somatic effects of cefazolin, cefamandole and moxalactam were related both to the antibiotic concentration and time of exposure to the drugs and were observable within 30 minutes of the initial exposure of the cultures to these agents. These results demonstrate that fluorescent cytometry can provide accurate assessment of the effects of compounds that inhibit cell wall formation. This technology could be a useful tool for comparing antibiotic somatic effects on bacteria and for rapidly and reliably determining their sensitivity and resistance to these agents.
Conversion of linear histogram flow cytometry data to a logarithmic display
Authors: Brunk C. F. , Bohman R. E. , Brunk C. A. 
Published: August 2005

AbstractA routine is described that readily allows the rescaling of linear histographic data to a corresponding logarithmic histogram. This procedure significantly improves data display, particularly where a wide range in the measured parameter is encountered. The logarithmic scale displays peaks with band widths more proportional to their respective coefficients of variation than is the case in a linear display. Rescaling several linear histograms to a common logarithmic scale allows the combination of these linear data even though the linear ranges are different. This routine is presented as a program written in BASIC for execution on a microcomputer.
The analysis and interpretation of DNA distributions measured by flow cytometry
Authors: Dean Phillip N. , Gray Joe W. , Dolbeare Frank A. 
Published: August 2005

AbstractA principal use of flow cytometers is for the measurement of fluorescence distributions of cells stained with DNA specific dyes. A large amount of effort has been and is being expended currently in the analysis of these distributions for the fractions of cells in the G1, S, and G2 + M phases. Several methods of analysis have been proposed and are being used; new methods continue to be introduced. Many, if not most, of these methods differ only in the mathematical function used to represent the phases of the cell cycle and represent attempts to fit exactly distributions with known phase fractions or unusual shapes. In this paper we show that these refinements probably are not necessary because of cell staining and sampling variability. This hypothesis was tested by measuring fluorescence distributions for Chinese hamster ovary and KHT mouse sarcoma cells stained with Hoechst‐33258, chromomycin A3, propidium iodide, and acriflavine. Our results show that: (a) single measurements can result in phase fraction estimates that are in error by as much as 40% for G2 + M phase and 15–20% for G1 and S phases; (b) different dyes can yield phase fraction estimates that differ by as much as 40% due to differences in DNA specificity; (c) the shapes of fluorescence distributions and their interpretation are very dependent on the dye being used and on its binding mechanism.
Flow cytometric analysis of G1 exit kinetics in asynchronous L1210 cell cultures with the constant transition probability model
Authors: Sharpless T. K. , Schlesinger F. H. 
Published: August 2005

AbstractFollowing addition of mitotic inhibitors to asynchronous cultures of mouse lymphoid cell line L1210, the rate of cell exit from G1 phase accelerates during the first 2–3 hr then declines with apparent first order kinetics. The simplest mathematical model that calls for biphasic G1 exit kinetics is the two‐state model with constant transition probability. It posits an exponential distribution of residence times in the first state, which is wholly contained in the G1 phase, and a fixed duration, Tb, for the rest of the cycle.We derive the distribution of expected times‐to‐division predicted by the model for a population in exponential steady state growth, and integrate this to obtain the predicted G1 exit curve for the blocked population as a function of two parameters, Tb and the population doubling time, Td.The experimental exit curves were fractions of the population remaining below an early S phase DNA content threshold, as a function of time after adding Colcemid, vincristine or vinblastine to exponentially growing cultures. Doubling times were determined from the rate of accumulation of blocked cells in G2 + M. An iterative method was then used to obtain a least‐squares fit of Tb and an observational parameter, Tt, representing the average time‐to‐division of cells at the chosen DNA threshold.The fitted G1 exit curves match the data well, showing that the constant transition probability model is a useful mathematical description of the G1 kinetics of these cells. The method described is a convenient way to estimate the parameters of this model for asynchronous, exponentially growing populations.
Design and operation of a dual laser chromosome sorter
Authors: Lebo Roger V. , Bastian Araxy M. 
Published: August 2005

AbstractWe describe the design and operation of a dual laser fluorescence activated chromosome sorter that can analyze and sort chromosome suspensions stained with two complementary DNA stains. We used two 18‐watt argon‐ion lasers to provide maximal excitation of the DNA‐specific stains Hoechst 33258 and chromomycin A3. The lasers are focused onto the sample stream by independent optics permitting optimal focusing of any pair of laser wavelengths. We incorporated beam expanders to decrease the energy density on the optics and improve the focal spot uniformity. Adjustable focusing lens holders are used to set the optimal focal spot size. The redesigned sorter separates suspensions of normal human fibroblast chromosomes into 21 fractions and can reliably sort more pure chromosome fractions than was previously possible.
Simple methods to determine and compare the sensitivity of flow cytometers
Authors: Pinkel D. , Steen H. B. 
Published: August 2005

AbstractKnowledge of the sensitivity of a flow cytometer is important both in choosing an instrument for a particular application and in determining whether the resolution obtained on real samples is limited by instrumental factors or sample variability. The coefficient of variation (CV) obtained for a measurement is given by CV2 = CVS2 + CVI2 + 1/n + K(δVB2)/n2, where CVS is the variability due to the sample, CVI that due to variations in illumination of the sample particles, n the number of photoelectrons emitted from the photomultiplier photo cathode per sample particle (which is the signal intensity), δVB2 the mean square background noise voltage (usually due to background illumination), and K a constant. Measurement of the CV of light flashes from a light emitting diode as a function of their intensity allows development of standard curves which can be used to directly determine the degree to which the resolution obtained on actual samples is limited by signal intensity and background light. The use of standard particles allows relative calibration of the curves for different instruments and which also ranks their sensitivities.
Multistation multiparameter flow cytometry: A critical review and rationale
Authors: Shapiro Howard M. 
Published: August 2005

AbstractThe capacity for fluorescence excitation by beams of different wavelengths at separate points along the sample stream, and the capacity for computer analysis of multiparameter data thus obtained, are now available in flow cytometer/sorter systems from commercial producers. It is now readily apparent to most experienced users of flow cytometers that such multiparameter analysis offers the most convenient solution to the problem of characterizing subpopulations of cells within a mixed population. The use of multiple beams facilitates resolution of fluorescence signals from several probes within or upon a single cell and widens the range of analytical alternatives available to experimenters. This critical review discusses the history of the instrumentation, the parameters now measurable and the probes used for their measurement, and the methods for data analysis. Required sensitivity and precision are discussed, leading to the conclusion that many of the advantages of multistation, multiparameter flow cytometry can be made available in less complex and less costly instruments using less powerful sources and less elaborate computer hardware than are presently incorporated in commercial apparatus.
Application of the CYTOMIC 12 flow cytometric compact analyzer for automatic kinetic measurements
Authors: Kachel V. , Schneider H. , Bauer J. , Malin‐Berdel J. 
Published: August 2005

AbstractFlow cytometry, usually applied to cells which have time independent features, can also be used for kinetic experiments where the change of cell populations with time is investigated. Dedicated time sequencing programs written in Assembler and incorporated in the CYTOMIC 12 analyzer (4) are described. A sequence of 64 one parameter histograms can be automatically acquired and immediately displayed as a pseudo‐two‐parameter histogram. The acquisition time for each of the subsequent histograms can be selected between 1 and 32 seconds. Kinetics lasting up to 34 minutes are resolved into 64 time intervals. Two parameter kinetics can be resolved into 12 32 × 32 channel, two parameter histograms which are displayed and evaluated immediately on the analyzer screen in groups of 4 without using complicated list mode procedures. The standard CYTOMIC 12 software can be applied for processing and printing of the sequence distribution curves.
Nondiscrete heterogeneity of human erythrocytes: Comparison of coulter‐principle flow cytometry and Soret‐hemoglobinometry image analysis
Authors: Bessman J. David , Hurley Edward L. , Groves Michael R. 
Published: August 2005

AbstractIn the blood of normal subjects, the volumes of single erythrocytes are distributed with a coefficient of variation (cv) of 10.8 ± 1.8%; while in hemoglobinopathies, cv increases proportionately to the degree of anemia produced. Using single cell Soret‐band hemoglobinometry and focused‐aperture impedance counting, we compared the distribution of red cell volume, area, hemoglobin content, and hemoglobin concentration in normals and subjects with anemic disorders. The cv, nondiscrete heterogeneity, is first, a general characteristic of biologic measurement, second, a sensitive indicator of abnormality of erythropoiesis, and third, consistently less for hemoglobin concentration than for volume, area, or hemoglobin content of the same cells.
Behavior of elliptocytes under shear stress in the rheoscope and ektacytometer
Authors: Bull Brian , Feo Claude , Bessis Marcel 
Published: August 2005

AbstractWhen ellipsoidal red cells from patients with hereditary elliptocytosis are subjected to shear stress in a rheoscope the cells rotate around their short axis and align themselves perpendicular to the flow. At high shear stress they deform into an ellipsoid of revolution with the major axis of the ellipse normal to the flow. This change in the direction of the long axis is not the result of simple reorientation of the cells. Rather, the short axis of an elliptocyte exposed to increasing shear stress expands while its long axis contracts. It is this process that eventually brings about the apparent reorientation.Elliptocytes in a well mixed suspension are randomly oriented. As a consequence, the initial diffraction pattern displayed by the ektacytometer is a hazy circle. As the shear stress is increased this becomes rhomboid due to expansion of the central region of each cell (a change confirmed by the rheoscope). Central expansion coexisting with polar remnants of the resting elliptocytic cell shape gives rise to the rhomboid outline. As the shear stress in the ektacytometer approaches 120 dynes/cm2 the characteristic, elliptical diffraction pattern seen with normal cells makes its appearance. As the shear stress is decreased, the rhomboid pattern reappears followed by a new elliptical pattern oriented at right angles to the one seen under high shear. This pattern arises from the resting shape of the elliptocytes which have all been oriented at right angles to the shear stress as it decreased to zero. As a consequence of this uniform cellular orientation, all exposures to shear stress after the initial one start with a deformability index of less than zero if it is calculated in the usual fashion from the axial ratio of the shear deformed cells. Certainly under these circumstances and possibly in other conditions as well the use of the less‐specific term “ektacytometric index” would be less misleading.
Setting up and running a microscope‐based flow cytometer
Authors: Petersen Steffen E. 
Published: August 2005

AbstractA noncommercial flow cytometry system was set up in a laboratory with no engineering staff attached. The system is based upon a conventional fluorescence microscope and a new flow principle reported by H. Steen and T. Lindmo (Science 204:403, 1979). A few modifications and additions have been introduced. The instrument has proven to be a high‐resolution system, easy to adjust and control, giving coefficients of variation of cellular DNA down to 0.8% and of fluorescent polyethylene microspheres down to 0.9%.