bluuebunny/crossref_metadata_2025 huggingface dataset:
Critical look on new approach for geometrically evolving domains in geomechanics
Authors: Modaressi H. , Roux F.
Published: August 2005

Numerical modelling of geometrically evolving domains, such as multistep construction, accretion and excavation, has always been of great interest to geotechnical engineers. Stress, strain and pore‐pressure changes depend on hydromechanical properties and the loading process. Several techniques are proposed in the literature for numerical simulation of excavation or backfilling processes. These are mostly based on a natural methodology that consists in dividing the problem into several time steps, each coinciding with the addition or removal of one layer which is then modelled by means of several elements. Unfortunately, as in these methods the evolving parameters and their relative influence are not explicit, it is not possible to determine their accuracy. A more rigorous approach lies in the geometrical transformation of the studied domain, depending on the time variable for a reference domain in which the conservation equations are verified. The application of this methodology implies a great number of calculations. By using limited development techniques of the Jacobian of transformations it is possible to reduce the volume of computations. Such a technique results in approximations which can be evaluated.
Finite‐element method for EHL analysis of spherical bodies
Authors: Karami G. , Zarinchang J. , Eraghi R. Satourians , Kazeminejad H.
Published: August 2005

A finite‐element solution for elastohydrodynamic lubrication (EHL) of spherical bodies undergoing elastic deformation is presented. The Galerkin finite‐element approach is employed and the domain under investigation is discretized using isoparametric quadrilateral elements. Isothermal conditions are assumed and the generation of the pressure is assumed to be governed by the Reynolds equation. The viscosity and the density of the oil are assumed to be functions of the pressure. It is assumed that the underlying surface deforms elastically under the hydrodynamic pressure distribution. The accuracy of the technique is illustrated in particular examples.
Empirical way to improve stress accuracy at stress concentrations in finite element analysis
Authors: Tenchev R. T.
Published: August 2005

The nodal stresses in a finite element may be calculated in two ways: (a) directly at the nodes; (b) by bilinear extrapolation from the stresses at the 2 × 2 Gauss points. In both cases the von Mises stress is computed from the averaged nodal stresses. In areas of stress concentration, when the mesh is not adequately refined, there is a significant difference between the maximum von Mises stresses when methods (a) and (b) are used. By means of numerical examples it is shown that method (a) is superior to (b). Using both stress values an empirical formula, based on numerous FEM results, is presented for improving the accuracy of the maximum von Mises stress. The error reduction is considerable. Only plane stress problems with 8‐node quadrilateral isoparametric elements are considered.
Applying dispersion correction to numerical approximations of the two‐dimensional wave equation ‐ eigenproblems
Authors: Mackie R. I.
Published: August 2005

A technique is presented whereby numerical calculations of vibration modes can be improved. The paper looks at the classical two‐dimensional wave equation using finite difference approximations. Analysis of the numerical dispersion of the approximations is used to develop a correction method. In general the numerical dispersion is dependent upon both the frequency and the direction of a wave, but if a 9‐point formula is used the directional dependence is much reduced. This enables correction factors to be obtained using only the frequency of a vibration mode. The method was tested on the vibration of a square membrane and of an L‐shaped region; in both cases a marked improvement in accuracy was obtained, at very little computational cost.
A direct solver for the least‐squares problem arising from GMRES(k)
Authors: Galán Manuel J. , Montero Gustavo , Winter Gabriel
Published: August 2005

This is a study about one of the core questions in the GMRES(k) method regarding the obtaining of vector yk for the least‐squares problem, argminy |Hky ‐ β(n)e1|2 (see Saad and Schultz1). We propose a simple but efficient approach to the resolution of this problem and a low cost computation of the residual and the residual norm, including both in a complete and detailed FGMRES(k) algorithm. The whole algorithm of minimization only involves two backward substitutions with triangular matrices and a dot product. The residual and the residual norm are computed, making use of results in the least‐squares problem.
Semi‐discretization method for the heat equation with non‐local boundary conditions
Authors: Araújo A. L. , Oliveira F. Aleixo
Published: August 2005

Semi‐discretization methods with iterated corrections are considered for solving the heat equation with boundary conditions containing integrals over the interior of the interval. The given problem is transformed into an ordinary differential system of equations, when we substitute the spatial derivative by finite differences. Such a system is then solved with an implicit integrator together with a quadrature method for the boundary integrals and, for each time step, the numerical scheme ‐ implicit integrator and quadrature method ‐ is repeated iteratively in order to achieve a given accuracy. The extra work we need with the iterated corrections enables us to take care of the interrelation of the solution u(x, t) at the interior and at the boundary points and also to start the numerical process with rough approximations of u(0, t) and u(1, t).

An improvement of the results is achieved when we estimate the local spatial truncation error and make the injection of that estimation into the ordinary differential system. Numerical experiments are presented.
Dispersion properties of simple 3D finite elements
Authors: Johnson Erland
Published: August 2005

The dispersion properties of plane waves, propagating in an infinite uniform mesh with linear cubic 3D finite elements, are investigated. Time discretization is performed with the central‐difference time‐stepping scheme in combination with lumped mass matrices. The stiffness matrices are integrated analytically with different integration schemes, and dispersion relations are derived. The results show that the stiffness integration scheme can be chosen to reduce dispersion and improve the isotropy of wave propagation. It is also observed that the numerical wave speed becomes zero along certain wave propagation directions for specific wave numbers.
Some conformal map constructs for numerical grids
Authors: Carey G. F. , Muleshkov A.
Published: August 2005

We develop some special conformal maps and related strategies for grid generation in certain ‘difficult’ domains. Of particular interest are non‐convex domains, and domains with corners and cusps.
On the pseudomaterial approach for the analysis of transient forming processes
Authors: Cruchaga M. , Oñate E. , Idelsohn S.
Published: August 2005

A fixed‐mesh method for the analysis of transient forming processes is presented. The mesh covers material regions and zones through which the material may flow. These last zones are identified by a pseudomaterial with relatively small physical parameters. During time processing, the interface between both materials is followed by an arbitrary Lagrangian mesh. This technique appears to be suitable for the treatment of moving surfaces with sharp corners. A particular boundary condition for the Navier‐Stokes equations is also introduced in order to model a porous wall.
A note on FEM buckling analysis
Authors: Wagner W.
Published: August 2005

The investigation of the non‐linear response of shell‐like structures requires insight into stability behaviour. In the paper we compare two strategies to compute singular points based on different eigenvalue problems. We show a simple algorithm to calculate critical load factors A used in engineering buckling analysis from the eigenvalues of the standard eigenvalue problem (KT – ω1) φ = 0. Some numerical examples illustrate the derived results and algorithms.
On the assumed displacement fields of a shallow curved shell finite element
Authors: Allman D. J.
Published: August 2005

An essential requirement for a satisfactory curved shell finite element is a capability for exact representation of rigid body movements: otherwise serious difficulties can occur, as is well known from early developments in shell finite element analysis. It is shown in this paper that appropriate assumed displacement fields for a particular shallow curved shell finite element with 18 degrees of freedom can be obtained in a very simple, yet apparently novel, way which permits the exact recovery of all rigid body movements. The shallow curved shell finite element considered here is constructed on a triangular flat facet approximation to a general thin elastic shell; it has quadratic middle surface geometry to allow for accurate modelling of arbitrary curvatures.
A posteriori finite element method error estimates for fourth‐order problems
Authors: Pomeranz Shirley B.
Published: August 2005

A posteriori error estimation for typical Euler‐Bernoulli beam bending problems is developed using a recovery operator based on a smoothing technique that is applied in the context of one‐dimensional fourth‐order problems. The development exploits a discrete superconvergence property of the ‘bending moments’ of the trial space interpolant of a polynomial solution of sufficiently low degree at certain Gauss points, and the superconvegence property of the energy norm difference of the trial space interpolant (of the weak solution) and the finite element approximation. An asymptotically exact error estimator is developed.
Distributed finite element structural analysis using the client–server model
Authors: Hudli Anand V. , Pidaparti R. M. V.
Published: August 2005

The paper presents a distributed computing approach for concurrent finite element analysis of truss structures. The client‐server model of distributed computation is applied to the plane frame finite element algorithm and implemented on SUN workstations. The remote procedure call combined with the ability to create threads of computation allows one to efficiently implement distributed and concurrent programs. The present approach is demonstrated through a typical truss structure to illustrate the details of the implementation. This approach can be used for very large finite element systems.
Selecting upwind parameters for improved iterative performance
Authors: Abbasian Reza O. , Carey Graham F.
Published: August 2005

The effect of upwinding on iterative performance of convection–diffusion problems is investigated. An analysis of the iterative method considered here leads to a criterion for selecting the optimal upwinding parameter to improve iterative performance for a class of two‐dimensional convection–diffusion problems. Supporting numerical experiments are presented.
The h‐, p‐ and hp‐versions of the BEM in elasticity: Numerical results
Authors: Holzer Stefan M.
Published: August 2005

The paper investigates the convergence of the h‐, p‐ and hp‐versions of a variational symmetric boundary element method (BEM) in plane elasticity by numerical experiments. The study discusses mixed boundary value problems on polygonal domains, i.e. problems for which the exact solution is analytic except in a finite number of points. The convergence of the error in energy norm is displayed for all versions of the BEM. All results are also compared with those obtained by the corresponding finite element methods. The theoretically predicted asymptotic convergence rates for all versions of the method can be observed in the numerical experiments. A comparison of computer times is given as well.
Comparison of symbolic and numerical integration methods for an assumed‐stress hybrid shell element
Authors: Rengarajan Govind , Knight Norman F. , Aminpour Mohammad A.
Published: August 2005

Hybrid shell elements have long been regarded with reserve by the commercial finite element developers despite the high degree of reliability and accuracy associated with such formulations. The fundamental reason is the inherent higher computational cost of the hybrid approach as compared to the displacement‐based formulations. However, a noteworthy factor in favour of hybrid elements is that numerical integration to generate element matrices can entirely be avoided by the use of symbolic integration. In this paper, the use of the symbolic computational approach is presented for an assumed‐stress hybrid shell element with drilling degrees of freedom, and the significant time saving achieved is demonstrated through an example.
On the evaluation of nearly singular kernel integrals in boundary element analysis
Authors: Wu Shukai
Published: August 2005

The paper concentrates on the numerical evaluation of nearly singular kernel integrals commonly encountered in boundary element analysis. Limitations of the method developed recently by Huang and Cruse (1993) for the direct evaluation of nearly singular kernel integrals are analysed and pointed out. A general method is developed that extends the idea for all practical cases. Example results are presented, demonstrating the effectiveness of the method.
Pressure‐dependent plasticity models: Loading–unloading criteria and the consistent linearization of an integration algorithm
Authors: Govindarajan R. M. , Aravas N.
Published: August 2005

A general methodology for the numerical integration of pressure‐dependent plasticity models is developed, and the corresponding ‘linearization moduli’ are derived.
New stress recovery formula for B‐bar methods
Authors: Jirásek Milan
Published: August 2005

An alternative justification of the B‐bar procedure using the Hu–Washizu variational principle is presented. In contrast to previous work, the derivation starts from the three groups of algebraic equations (strain‐displacement equations, stress‐strain equations and equations of equilibrium) obtained by discretization of the Hu–Washizu functional. Based on an appropriate assumption, these equations can be manipulated to yield the element stiffness matrix in the form typical of the B‐bar procedure. This approach directly leads to a stress‐recovery formula different from the one suggested before. The theory is illustrated by application to a family of quadrilateral plane elements.
On the convergence of the generalized matrix multisplitting relaxed methods
Authors: Zhongzhi Bai
Published: August 2005

In this paper, I discuss the convergence and divergence rates for a class of generalized matrix multisplitting relaxation methods in a detailed manner. In particular, when the coefficient matrix is an L‐matrix, I obtain several sufficient and necessary conditions ensuring the convergence of these methods.
Modal reaction method for modal participation factors in support motion problems
Authors: Chen J. T. , Hong H.‐K. , Yeh C. S.
Published: August 2005

An efficient method, called the modal reaction method, for calculating the modal participation factors for support motion problems is proposed. It is found that modal reactions at supports proportionally describe the magnitudes of the modal participation factors. Thus, the modal participation factor for support motion problems can be calculated very efficiently, saving more than 99 per cent, for the given example with 640 degreees of freedom, of the CPU time compared with using the popular quasistatic decomposition method.
Evaluation of some error estimators for the four‐noded lagrangian quadrilateral
Authors: Ramsay A. C. A. , Sbresny H.
Published: August 2005

A comparative study of error estimators using a patch recovery scheme with those using simple nodal averaging is made for the four‐noded Lagrangian quadrilateral element through two plane stress elasticity problems. It is demonstrated that error estimators using a patch recovery scheme are generally more effective and have an estimated stress field that is closer to the exact one than those using simple nodal averaging.
Parallel computation of integral equation methods for three‐dimensional transient wave propagation
Authors: Walker S. P. , Yeung C. Y.
Published: August 2005

Electromagnetic and acoustic scattering from bodies many wavelengths long presents some of the largest computational problems, with costs scaling with the fourth to sixth power of incident frequency. Parallel processing offers one contributory route to the increase in computational power required for such analyses. The paper presents a parallel implementation of the integral formulation of the transient wave equation. It is demonstrated that it is the repeated evaluation of far field effects which is the dominant cost, and that with appropriate domain partitioning this process can be parallelised very effectively. Results for scattering problems comprising up to 1658 nodes are presented, evaluated on 64‐ and 128‐processor systems.
Finite hyperelements: A 4D geometrical framework using covariant bases and metric tensors
Authors: Perrochet P.
Published: August 2005

A method is presented to evaluate space‐time finite element matrices in a general 4D context. Unlike the classical Jacobian‐based ‘textbook’ approach, the method described herein uses curvilinear gradient operators to properly handle finite elements having fewer local dimensions (i.e. two or three) than the global embedding domain. This unconventional, but powerful, method is described in details and is illustrated by an application simulating 4D pure convection in a curved diverging‐converging flow field.
The checkerboard patterns of P2 – P1 discontinuous pressure elements
Authors: Xiao‐Liang Cheng , Jin‐Sheng Jiang
Published: August 2005

In this note we obtain the checkerboard patterns of P2 – P1 discontinuous pressure elements for the Stokes problem The results are more complete than those previously presented.
Calculation of fully developed flows of complex fluids in pipes of arbitrary shape, using a mapped circular domain
Authors: Ahmeda M. , Normandin M. , Clermont J.‐R.
Published: August 2005

The problem of determination of fully developed duct flows of arbitrary shape is considered for fluids obeying various constitutive equations. A numerical method based on the transformation of the physical flow (simply connected) domain into a mapped circular domain is set up. The numerical determination of the unknown transformation between the two domains leads to knowledge of the velocities and stresses in the duct, starting from circular contour values of the velocity in the mapped computational domain. Different constitutive equations are investigated for several flow geometries. The comparisons with existing analytical solutions are found to be consistent for all the geometries tested. The method may provide a useful tool for determination, in steady flow conditions, of the flow characteristics for various types of ducts and constitutive equations.
The use of conjugate gradients methods with a segregated finite volume procedure for solving transient, incompressible Navier–Stokes equations
Authors: Nieplocha J. , Schreiber W. C.
Published: August 2005

PISO (pressure‐implicit with splitting of operators) is an algorithm devised to solve the transient incompressible Navier–Stokes and energy equations. In that it uses separate equations for each of two pressure corrections, velocity components and transported scalars, PISO is known as a segregated solution procedure. Preconditioned, generalized conjugate gradient (GCG) methods in combination with schemes for vectorization and storage minimization are described in the paper for the purpose of solving both the symmetric and non‐symmetric algebraic equation systems that result from the PISO algorithm. Of particular interest in the paper is a comparison of the present procedure with a procedure described by Chin et al. (1992) in which preconditioned conjugate gradient methods are used to solve the Jacobian matrix used in the Newton iteration of the fully coupled non‐linear equations. The two methods are similar in that both techniques are based on a discretization using staggered control volumes with the power‐law method for modelling advection‐diffusion transport.
Theoretical formulations for adaptive finite element computations
Authors: Li L.‐Y. , Bettess P. , Bull J. W. , Bond T. , Applegarth I.
Published: August 2005

A new mesh refinement formulation for h‐adaptive solutions of linear elliptic problems is presented. The reliability of the methods employed for estimating errors and the convergence rate of the errors during an adaptive process are investigated by means of an asymptotic analysis method. The present adaptive mesh refinement strategy significantly improves adaptivity performance and has higher error convergence rates than those usually used. Numerical examples are shown to illustrate the efficiency of the method shown.
Computation of the turbulent flow field around an aerofoil by k‐ϵ‐γ model in a non‐staggered, C‐type single‐grid system
Authors: Kim Ki Sup , Chung Myung Kyoon
Published: August 2005

The whole turbulent flow field around a two‐dimensional aerofoil is solved with a consistent numerical method on a single‐grid system and with an intermittency implemented two‐equation model, namely the k‐ϵ‐γ turbulence model in elliptic form. In order to solve the numerical instability problem arising from the branch‐cut in C‐type grids using the non‐staggered finite‐volume method, a new numerical technique is proposed to guarantee the correct velocity‐pressure coupling along the branch‐cut. The computational results are compared with those by the standard k‐ϵ model and experimental data. This study demonstrates the relative simplicity of the present C‐type single grid system, and the capability is assessed of the k‐ϵ‐γ equations for predicting turbulent flows consisting of both wall‐free and wall‐bounded regions.
Numerical simulation of viscoelastic complex flows using the stream‐tube method and optimization algorithms
Authors: Bereaux Y. , Clermont J. R. , Yassine A.
Published: August 2005

Numerical simulation of complex flows of viscoelastic fluids requires us to use efficient algorithms for solving the non‐linear governing equations. The stream‐tube analysis permits calculation of such flows by considering elementary subdomains in a mapped computational domain where the transformed streamlines are parallel and straight. The consideration of subregions leads us to use optimization procedures. The paper concerns the application of such algorithms to compute main flows in axisymmetric converging geometries, for fluids obeying memory‐integral constitutive equations.
Finite element nodal ordering algorithms
Authors: Kaveh A. , Behfar S. M. R.
Published: August 2005

Graph theoretical algorithms are presented for finite element nodal ordering to reduce the bandwidth of their stiffness matrices. Five different graphs are defined for representing the connectivity of FE models, and the problem of finite element nodal ordering is transformed into that of a graph nodal ordering. The efficiencies of these methods are compared through examples of two‐ and three‐dimensional finite element models.
Infinite boundary element for elastodynamic problems of anti‐plane motion in an elastic half‐space
Authors: Fu T. M. , Cheung Y. K.
Published: August 2005

For elastodynamic problems in an elastic half‐space, the idea of describing the displacement variation on the free ground surface outside the region being studied with the fundamental solution in a full space is presented in the paper. Then a corresponding mapped infinite boundary element is constructed for the problems of anti‐plane motion. The computed results show that such an element can be applied effectively to obtain high‐precision solutions.
A finite difference algorithm for plotting electric force surface in three‐dimensional field computation
Authors: Slimani D. , Djellal K. , Chemali H. , Kolli K. , Mesli R.
Published: August 2005

A finite difference algorithm for plotting the electric force surface is presented in three‐dimensional electrostatic field computation. Based on the flux concept, the nodal electric flux is calculated numerically through the curve integral of the electric displacement vector which was obtained by a numerical differentiation of the nodal electric potential. This latter is calculated by a finite difference method. The electric force surface may also be plotted.
Rapid growth of KDP crystal from aqueous solutions with additives and its optical studies
Authors: Li Guohui , Xue Liping , Su Genbo , Li Zhengdong , Zhuang Xinxin , He Youping
Published: August 2005

Potassium Chloride (KCl) and Ethylene diamine tetraacetie acid (EDTA) as new additives were added into the KDP solutions in a small amount (5M% and 0.01M% respectively). The solubility curve and metastable zone width of KDP solution with 5M%KCl was determined and we explained the mechanism of rapid growth of KDP crystal with KCl additve. The clear transparent crystal of KDP with a dimension of 54 × 54 × 42 mm have been grown rapidly by cooling solution method in 2 days. The crystal grown from additives added solution was subjected to optical transmission and laser damage threshold studies as compared with the crystal grown by traditional growing method. (© 2005 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)
A novel metal‐organic coordination complex crystal: tri‐allylthiourea zinc chloride (ATZC)
Authors: Sun H. Q. , Yuan D. R. , Wang X. Q. , Cheng X. F. , Gong C. R. , Zhou M. , Xu H. Y. , Wei X. C. , Luan C. N. , Pan D. Y. , Li Z. F. , Shi X. Z.
Published: August 2005

A new metal‐organic coordination nonlinear optical crystal, tri‐allylthiourea zinc chloride (ZnCl2(AT)3, where AT is CH2=CHCH2NHCSNH2, abbreviated as ATZC), is reported. It was synthesized in water and recrystallized in ethanol. For the crystal structure, optical and thermal characterization were determined by elemental analyses, X‐ray diffraction, infrared spectroscopy, differential scanning calorimeter, thermogravimetric analysis, and powder SHG efficiency meansurement. It belongs to the trigonal system, space group R3, with a = 11.0498(4) Å, c = 16.0416(11) Å, z = 3 and V = 1696.24(15) Å3. It exhibits powder SHG efficiency the same order as Urea crystal. (© 2005 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)
High pressure electrical resistivity study on nonlinear bis thiourea cadmium chloride (BTCC) single crystal
Authors: Ariponnammal S. , Selva Vennila R. , Radhika S. , Arumugam S.
Published: August 2005

The Bis Thiourea Cadmium Chloride (BTCC) crystals have been crystallized by slow evaporation technique. The lattice parameters of the grown crystals have been determined by the Energy dispersive x‐ray diffraction technique (EDXRD) and the structure has been confirmed. The high pressure electrical resistivity study have been carried out on this crystal and the results have been reported here. (© 2005 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)
Conduction mechanisms in TlSbSe2 chalcogenides
Authors: Ulutas K. , Kalkan N. , Yildirim S. , Deger D.
Published: August 2005

TlSbSe2 samples used for electrical measurements were cleaved from larger crystals grown by using the Bridgman ‐ Stockbarger method. Electrical properties of the samples are investigated with a view of determining the dominant conduction mechanism. DC conduction studies reveal that Poole – Frenkel conduction is predominant. The DC conduction studies revealed the non‐ohmic type conduction. The activation energies are also calculated. It is seen that the activation energy decreases with increase of applied voltage. (© 2005 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)
Dielectric properties of thallium gallium diselenide layered crystal in the incommensurate phase
Authors: Şentürk E. , Tümbek L. , Mikailov F. A.
Published: August 2005

The dielectric measurements of the layered crystal were studied in temperature range of successive phase transitions. The measurements revealed that the phase transition occurred in 242 K is an incommensurate phase transition. When the sample is annealed at a stabilized temperature in the incommensurate phase, a remarkable memory effect has been observed on cooling run. The mechanism of the memory effect in the incommensurate phase of the semiconducting ferroelectric TlGaSe2 can be interpreted in the frame of the theory of defect density waves. This theory claims that the memory effect is the result of pinning of the incommensurate structure by the lattice inhomogeneities. With decreasing the annealing temperature the phase transition temperature shifts to lower temperatures gradually. Moreover, the peak intensities also increase gradually. In addition to these effects, the phase transition temperature shifts to lower temperatures with increasing annealing time. (© 2005 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)
Microstructure of a genuine Damascus sabre
Authors: Levin A. A. , Meyer D. C. , Reibold M. , Kochmann W. , Pätzke N. , Paufler P.
Published: August 2005

The surface and cross‐section of a genuine Damascus sword was characterised by means of wide‐angle X‐ray scattering techniques complemented by optical and transmisssion electron microscopy. Position‐resolved X‐ray phase analysis revealed that, unlike ferrite and martensite, the distribution of cementite is inhomogeneous in different spatial zones parallel to the cutting edge of the blade. For the first time a quantitative X‐ray phase and texture analysis was made possible by averaging all spatial zones of the surface and the cross‐section of the sword and applying the Rietveld method with code TOPAS. Differences concerning texture and structure characteristics of ferrite, martensite and cementite are discussed. More evidence for the structure of cementite nanowires is supplied. (© 2005 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)
Bacterial growth studied by flow cytometry
Authors: Steen Harald B. , Boye Erik
Published: August 2005

The feasibility of flow cytometry for measurements on bacteria has been demonstrated by measurements of DNA‐associated fluorescence of Escherichia coli K‐12 in various phases of cell growth. Bacteria were stained with a combination of ethidium bromide and mithramycin after fixation in 70% ethanol. Cultures grown to stationary phase accumulated in two peaks representing cells with two and four chromosomes. Qualitatively similar histograms were obtained with cells grown in the presence of chloramphenicol, whereas cells of the temperature sensitive strain E 177 (dnaA) ended up with only one chromosome per cell at the restrictive temperature. The fluorescence intensity of cells with one chromosome was about 103 times smaller than that of human diploid cells. Instrumental resolution at this level of intensity was CV = 5%, whereas peak widths corresponded to CV = 7–8%. Dyes bound to RNA did not appear to contribute significantly to the fluorescence.
SV40‐Infected muntjac cells: Cell cycle kinetics, cell ploidy and T antigen concentration
Authors: Gershey Edward L.
Published: August 2005

Muntjac cells in which the SV40 virus neither readily causes transformation nor replicates were used to study the effect of SV40 infection on cell ploidy and the influence of ploidy on the concentration of T antigen, which is required for the initiation of viral DNA synthesis. Both the DNA content, as measured by flow microfluorometry of propidium iodide‐DNA fluorescence, and the average number of chromosomes per cell indicated that infection with SV40 did not alter the ploidy of the host cell. SV40 infection had no effect on the ploidy distribution of muntjac cells. After immunofluoresceinlabeled anti‐γG, infected and uninfected cultures were compared. In uninfected cells incubated with a 1:20 dilution of anti‐T serum no fluorescence could be observed by fluorescence microscopy, but when examined by flow microfluorometry, fluorescence was detected after standing with as little as 1000‐fold diluted antiserum. Determination of the amount of T antigen and DNA content in the same cell by simultaneous measurement of fluorescein isothiocyanate‐conjugate and propidium iodide fluorescence, indicated that the cellular concentration of T antigen did not vary with the ploidy of the genome or the number of nuclei per cell. These results suggest that gene dosage is not a factor which determines the permissive environment for SV40 replication.
Correlation of the growth of chinese hamster V279‐171b multicellular spheroids with cytokinetic parameters
Authors: Johnson T. S. , Bain E. , Raju M. R. , Martin J. C.
Published: August 2005

The volumes of Chinese hamster V279‐171b multicellular spheroids grown in spinner culture were measured and correlated to DNA distribution and cell volume distribution information. Spheroid growth curves were obtained over a 21‐day period using both electronic cell volume measurements and microscope micrometer sizing. The spheroids were collected from the aperture tube after electronic sizing, and single cell suspensions were prepared via trypsinization. Saline‐washed cells were then stained with mithramycin and analyzed as unfixed or ethanol‐fixed cells for both cell volume and DNA fluorescence measurements. Spheroid growth was determined to have an initial 8‐ to 9‐day rapid growth phase, and a slow phase from 10 to 21 days. The single cell volume distributions of cells obtained from spheroids were dependent on spheroid volume and the relative proportions of small volume noncycling cells to large volume cycling cells comprising the spheroids were also dependent on spheroid volume. DNA distribution data and dual parameter DNA distribution cell volume contour patterns obtained for single cells dissociated from spheroids were closely related to the measured spheroid volume and revealed that the fraction of small, hypoxic G0‐like cells was greatest in old, large spheroids.
Characterization of the cell cycle and synchrony in a CV‐1 cell line
Authors: D'Alisa Rose M. , Gershey Edward L.
Published: August 2005

Synchronous populations of CV‐1 cells have been prepared by the selective detachment of mitotic cells. An approach, using a combination of standard techniques, has allowed evaluation of the degree of synchrony throughout the cell cycle. CV‐1 cells have a cycle of ∼19‐hr duration, throughout which synchronous populations progress with a dispersion of 3 hr, i.e., ≥90% of the cells enter S phase and subsequently divide within a 3‐hr period. A major cause of this dispersion is the differential attachment times of individual cells within the population when a synchronous culture is initiated.
Collodion membrane secures cells sorted by flow cytofluorometry onto microscope slides
Authors: Meck Robert A. , Benson Neal A. , Ng Alan G. , Price Brandon J. , Ingram Marylou
Published: August 2005

Quantitative microscopic cytology of cells previously sorted by flow cytofluorometry has been hindered by the loss of cells from the microscope slide during staining procedures. The simple application of a semipermeable membrane of collodion over fixed or unfixed cells sorted directly onto a microscope slide secured virtually 100% of the cells onto the slide. Cells covered with the collodion membrane stained with Papanicolaou's stain as well as routine clinical cervical cytologic preparations. In contrast, fewer than one half the cells sorted onto uncoated or albumin coated slides were retained after staining.
Analytical cytometric approaches to heterogeneous cell populations in solid tumors: A review
Authors: Dethlefsen Lyle A. , Bauer Kenneth D. , Riley Reba M.
Published: August 2005

The problems encountered in studying the heterogeneity of cells in solid tumors is reviewed with emphasis on the role of various analytical cytometric assays for studying both the biology and the dynamics of proliferating, quiescent and dead malignant cells in vitro and in vivo. Due to advances in cytometric technology, many interesting in vitro studies on tumor cell heterogeneity have been and will be conducted over the next several years. For example, the acidic acridine orange staining of HeLa cells in suspension culture does readily discriminate between proliferating and quiescent cells. Some of these assays have been and others will be extended to in vivo studies. However, it is obvious that either the current analytical cytometric techniques must be modified and refined to permit better resolution for the complex situation in vivo or other new analytical cytometric techniques will have to be developed before many interesting studies on tumor cell heterogeneity in vivo can be addressed with reasonable efficiency.
New cell cycle compartments identified by multiparameter flow cytometry
Authors: Darzynkiewicz Zbigniew , Traganos Frank , Melamed Myron R.
Published: August 2005

Simultaneous measurements of cellular DNA and RNA as well as estimates of the sensitivity of DNA in situ to denaturation by acid (which correlates with the degree of chromatin condensation) and cell ability to incorporate 5‐bromodeoxyuridine (BUdR), performed on over 40 different cell systems enabled us to subclassify cells into 12 functionally distinct cell cycle compartments. Quiescent cells had low RNA values, DNA very sensitive to denaturation, and they did not incorporate BUdR. Although in most cell systems they had 2C DNA content (G1Q), quiescent cells with higher DNA values (SQ and G2Q) were also seen. The G1 phase of exponentially growing cells had two distinct compartments, A and B. G1 cells entered S phase only from the B compartment. An increase in RNA and a decrease in the sensitivity of DNA to acid denaturation beyond a specific level characterized the transition of cells from the A to B compartments. Since the G1A to G1B transition was not linear but exponential, it may be assumed that a non‐deterministic event triggering cell progression into the cycle resides within G1A. Under adverse growth conditions the probability of a G1A to G1B transition decreased. Cells In G2 had DNA more sensitive to acid denaturation than S or G1 phase cells. Mitotic cells had the most condensed chromatin and their RNA content was twice that of G1A cells. Differentiated cells were characterized by 2C DNA (G1D) but, depending on the cell type, had varying RNA content and different degrees of chromatin condensation. A hitherto undescribed category of cells undergoing transition from quiescence to the cycle (or vice versa) was distinguished based on their intermediate values of RNA, sensitivity of DNA to acid denaturation and inability to incorporate BUdR at the rate characteristic for S cells. Depending on their DNA content (C) at the time of transition these cells could be classified as G1T (2C), ST (4>C>2) or G2T (4C).
Evaluation of the S phase distribution of flow cytometric DNA histograms by autoradiography and computer algorithms
Authors: Sheck L. E. , Muirhead K. A. , Horan P. K.
Published: August 2005

Cell sorting and tritiated thymidine autoradiography were used to define the distribution of S phase cells in flow cytometric DNA histograms obtained from exponential mouse lymphoma cells (L5178Y). The numbers of labeled S phase cells, autoradiorgraphically determined from cells sorted at 2‐channel intervals in the G1/early S and late S/G2M regions of the histogram, were compared with the numbers of computed S phase cells in comparable 2‐channel intervals as predicted by several computer algorithms used to extract cell cycle phase distributions from DNA histograms. Polynomial and multirectangle algorithms gave computed estimates of total %S in close agreement with the tritiated thymidine labeling index for the cell population, while multi‐Gaussian algorithms underestimated %S. Interval autoradiographic and algorithm studies confirmed these results in that no significant differences were found between the autoradiographic S phase distribution and S phase distributions calculated by the polynomial and multirectangle models. However, S phase cells were significantly underestimated in G1/early S by a constrained multi‐Gaussian model and in both G1/early S and late S/G2 by an unconstrained multi‐Gaussian model. For the particular cell line (L5178Y), staining protocol (mithramycin following ethanol fixation) and instrumentation (Coulter TPS‐2 cell sorter) used in this study, close agreement between computed %S and tritiated thymidine labeling index was found to be a reliable indicator of an algorithm's success in resolving S phase cells in the G1/S and S/G2 transition regions of the DNA histograms.
A fluorescence polarization flow cytometer
Authors: Keene J. P. , Hodgson B. W.
Published: August 2005

The effect of mitogens on the degree of polarization of the fluorescence from viable human lymphocytes, labeled with fluorescein, have been reported to be different in cells from subjects with malignant disease as compared with those from healthy donors. This has potential applications in the field of clinical diagnosis and a flow cytometer could be the instrument of choice for this purpose. We believe that such an instrument should be simple and easy to use and a flow cytometer was assembled with this in mind and with the object of gaining experience which would provide the basis for a future finalized design. The instrument is suitable for use in other biologic applications where polarization is of interest and also as a general purpose flow cytometer. Its main features, its characteristics and a preliminary performance report are presented here.
A dual laser flow sorter utilizing a CW pumped dye laser
Authors: Arndt‐Jovin Donna J. , Grimwade Brian G. , Jovin Thomas M.
Published: August 2005

A dual laser flow sorter has been constructed from an existing single laser system by incorporating a dye laser as the second laser source. The argon ion laser emission was used both as a pump laser and as a source by beam splitting before entrance to the dye laser. The emissions of the dye laser and pump laser beams were recombined and focused with the same optical train used in the single laser system. The imaging on the stream of the flow sorter provided for spatial and thus temporal separation of the exciting beams and subsequent sample emissions.
Simultaneous quantitation of hoechst 33342 and immunofluorescence on viable cells using a fluorescence activated cell sorter
Authors: Loken Michael R.
Published: August 2005

The cytochemical stain Hoechst 33342 has been used to quantify DNA in viable cells and has been used under nonsaturating conditions to discriminate between lymphoid cell types. In order to correlate the quantitative emission from Hoechst 33342 with cell surface antigens, a fluorescence activated cell sorter was modified to simultaneusly detect emission from the UV excited Hoechst dy and fluorescein attached to the cell surface by immunofluorescence techniques. A special set of laser mmirrors was installed in an argon ion laser so that all the lines from 351‐488 nm could be used to illuminate the cells. Appropriate emission filters were used to separate the light emitted by Hoechst 33342 from the fluorescein. An electronic cross‐over circuit was used to compensate for special overlap between the two dyes. Analysis of murine lymph node cells stained both with Hoechst 33342 under nonsaturating conditions and anti‐Thy 1.2 indicated that the cells that stained dimly with the Hoechst dy expressed the Thy 1.2 marker while the cells that were brightly stained with Hoechst 33342 lacked this differentiation antigen. The correlation of cell surface myeloma protein with cell cycle on an in vitro cell line indicated that the heterogeneity of cell surface antigen expression could not be accounted for solely by variations occurring during the cell cycle.
Enzyme kinetic studies in cell populations using fluorogenic substrates and flow cytometric techniques
Authors: Watson James V.
Published: August 2005

Two flow cytometers, an Ortho Cytofluorograf model 4800A and a “custom” built dual laser instrument, have been interfaced with a PDP 11/40 computer to assay the rate of accumulation of fluorescent products in populations of single cells due to enzymatic hydrolysis of fluorogenic substrates. Esterase activity assayed with fluorescein diacetate exhibited abnormal Michaelis‐Menten kinetic behavior in EMT6 cells. This was in marked contrast to normal kinetics obtained with homogenates assayed with a conventional spectrofluorimeter. Moreover, the homogenates were 4 5x less efficient at converting substrate than were intact cells. Fluorescein diacetate is lipophilic and the induced fluorescence was evenly distributed throughout intact cells. In contrast a second substrate studied, 3‐O‐methyl fluorescein phosphate, is lipophobic and highly polar and the induced fluorescence was located at the cell surface. Results obtained with this substrate suggested that membrane, but not intracellular phosphatases were being assayed. Although the reaction product appeared to be lost rapidly from the site of hydrolysis it was possible to extract Km and Vmax for the dephosphorylation which followed normal kinetics. The dual laser flow cytometer was built to study 4‐methylum‐belliferone based substrates (UV excitation) simultaneously with those based on fluorescein. An assay using fluorescein diacetate and 4‐methylumbelliferyl acetate simultaneously was able to discriminate between a mouse mammary tumour (EMT6) and a human colonic adenocarcinoma cell line (HT 29). Furthermore, incubation of both cell lines for up to 6 hr with these substrates maintained viability at about 90%.
DNA measurements of chromosomes 9 and 22 of six patients with t(9;22) and chronic myeloid leukemia
Authors: Geraedts J. P. M. , Van der Ploeg M.
Published: August 2005

More than 90% of the cases of chronic myeloid leukemia are characterized by the presence of a so‐called Philadelphia chromosome. This Philadelphia chromosome has been shown to be a product of a translocation between chromosomes 9 and 22 in the majority of cases. It is present in the bone marrow and unstimulated blood cultures. Visual examination of this chromosome gave the impression that small differences in length might exist between patients. Therefore, DNA measurements were undertaken on six patients with t(9;22) and chronic myeloid leukemia. Metaphases obtained from unstimulated peripheral blood cultures were Q‐banded for photography and identification of chromosomes 9 and 22. Feulgen staining was performed, and the preidentified cells were photographed again. Cytophotometry of the photographic negatives was done using a Hidacsys‐Arrayscan computer program. Relative DNA contents were calculated for the normal chromosomes 9 and 22 and both chromosomes involved in the translocation. Between 7 and 17 cells were measured per patient. The results indicated that no differences existed between the patients regarding the amount of material translocated from chromosome 22 to 9. In all patients the sum of the DNA contents of the normal chromosomes 9 and 22 was less than that of the translocation chromosomes. Possible explanations for this observation are discussed.