Lee R Moore

Mie University, Tsu-shi, Mie-ken, Japan

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Publications (49)112.41 Total impact

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    ABSTRACT: Emerging applications of rare cell separation and analysis, such as separation of mature red blood cells from hematopoietic cell cultures, require efficient methods of red blood cell (RBC) debulking. We have tested the feasibility of magnetic RBC separation as an alternative to centrifugal separation using an approach based on the mechanism of magnetic field-flow fractionation (MgFFF). A specially designed permanent magnet assembly generated a quadrupole field having a maximum field of 1.68 T at the magnet pole tips, zero field at the aperture axis, and a nearly constant radial field gradient of 1.75 T/mm (with a negligible angular component) inside a cylindrical aperture of 1.9 mm (diameter) and 76 mm (length). The cell samples included high-spin hemoglobin RBCs obtained by chemical conversion of hemoglobin to methemoglobin (met RBC) or by exposure to anoxic conditions (deoxy RBC), low-spin hemoglobin obtained by exposure of RBC suspension to ambient air (oxy RBC), and mixtures of deoxy RBC and cells from a KG-1a white blood cell (WBC) line. The observation that met RBCs did not elute from the channel at the lower flow rate of 0.05 mL/min applied for 15 min but quickly eluted at the subsequent higher flow rate of 2.0 mL/min was in agreement with FFF theory. The well-defined experimental conditions (precise field and flow characteristics) and a well-established FFF theory verified by studies with model cell systems provided us with a strong basis for making predictions about potential practical applications of the magnetic RBC separation.
    Analytical and Bioanalytical Chemistry 10/2013; · 3.66 Impact Factor
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    ABSTRACT: A linear array of Nd-Fe-B magnets has been designed and constructed in an inverted Halbach configuration for use in separating magnetic nanoparticles. The array provides a large region of relatively low magnetic field, yet high magnetic field gradient in agreement with finite element modeling calculations. The magnet assembly has been combined with a flow channel for magnetic nanoparticle suspensions, such that for an appropriate distance away from the assembly, nanoparticles of higher moment aggregate and accumulate against the channel wall, with lower moment nanoparticles flowing unaffected. The device is demonstrated for iron oxide nanoparticles with diameters of ~5 and 20 nm. In comparison to other approaches, the inverted Halbach array is more amenable to modeling and to scaling up to preparative quantities of particles.
    IEEE Transactions on Magnetics 07/2013; 49(7):3449-3452. · 1.42 Impact Factor
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    ABSTRACT: Quadrupole Magnetic Field-Flow Fractionation (QMgFFF) is a technique for characterization of sub-micrometer magnetic particles based on their retention in the magnetic field from flowing suspensions. Different magnetic field strengths and volumetric flow rates were tested using on-off field application and two commercial nanoparticle preparations that significantly differed in their retention parameter, λ (by nearly 8-fold). The fractograms showed a regular pattern of higher retention (98.6% v. 53.3%) for the larger particle (200 nm v. 90 nm) at the higher flow rate (0.05 mL/min v. 0.01 mL/min) at the highest magnetic field (0.52 T), as expected because of its lower retention parameter. The significance of this approach is a demonstration of a system that is simpler in operation than a programmed field QMgFFF in applications to particle mixtures consisting of two distinct particle fractions. This approach could be useful for detection of unwanted particulate contaminants, especially important in industrial and biomedical applications.
    Analytical Sciences 01/2013; 29(7):761-764. · 1.57 Impact Factor
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    ABSTRACT: The emerging applications of biological cell separation to rare circulating tumor cell (CTC) detection and separation from blood rely on efficient methods of red blood cell (RBC) debulking. The two most widely used methods of centrifugation and RBC lysis have been associated with the concomitant significant losses of the cells of interest (such as progenitor cells or circulating tumor cells). Moreover, RBC centrifugation and lysis are not well adapted to the emerging diagnostic applications, relying on microfluidics and micro-scale total analytical systems. Therefore, magnetic RBC separation appears a logical alternative considering the high iron content of the RBC (normal mean 105 fg) as compared to the white blood cell iron content (normal mean 1.6 fg). The typical magnetic forces acting on a RBC are small, however, as compared to typical forces associated with centrifugation or the forces acting on synthetic magnetic nanoparticles used in current magnetic cell separations. This requires a significant effort in designing and fabricating a practical magnetic RBC separator. Applying advanced designs to the low cost, high power permanent magnets currently available, and building on the accumulated knowledge of the immunomagnetic cell separation methods and devices, an open gradient magnetic red blood cell (RBC) sorter was designed, fabricated and tested on label-free cell mixtures, with potential applications to RBC debulking from whole blood samples intended for diagnostic tests.
    IEEE Transactions on Magnetics 01/2013; 49(1):309-315. · 1.42 Impact Factor
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    ABSTRACT: Using novel media formulations, it has been demonstrated that human placenta and umbilical cord blood-derived CD34+ cells can be expanded and differentiated into erythroid cells with high efficiency. However, obtaining mature and functional erythrocytes from the immature cell cultures with high purity and in an efficient manner remains a significant challenge. A distinguishing feature of a reticulocyte and maturing erythrocyte is the increasing concentration of hemoglobin and decreasing cell volume that results in increased cell magnetophoretic mobility (MM) when exposed to high magnetic fields and gradients, under anoxic conditions. Taking advantage of these initial observations, we studied a noninvasive (label-free) magnetic separation and analysis process to enrich and identify cultured functional erythrocytes. In addition to the magnetic cell separation and cell motion analysis in the magnetic field, the cell cultures were characterized for cell sedimentation rate, cell volume distributions using differential interference microscopy, immunophenotyping (glycophorin A), hemoglobin concentration and shear-induced deformability (elongation index, EI, by ektacytometry) to test for mature erythrocyte attributes. A commercial, packed column high-gradient magnetic separator (HGMS) was used for magnetic separation. The magnetically enriched fraction comprised 80% of the maturing cells (predominantly reticulocytes) that showed near 70% overlap of EI with the reference cord blood-derived RBC and over 50% overlap with the adult donor RBCs. The results demonstrate feasibility of label-free magnetic enrichment of erythrocyte fraction of CD34+ progenitor-derived cultures based on the presence of paramagnetic hemoglobin in the maturing erythrocytes.
    PLoS ONE 01/2012; 7(8):e39491. · 3.73 Impact Factor
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    ABSTRACT: Cell separation and fractionation based on fluorescent and magnetic labeling procedures are common tools in contemporary research. These techniques rely on binding of fluorophores or magnetic particles conjugated to antibodies to target cells. Cell surface marker expression levels within cell populations vary with progression through the cell cycle. In an earlier work we showed the reproducible magnetic fractionation (single pass) of the Jurkat cell line based on the population distribution of CD45 surface marker expression. Here we present a study on magnetic fractionation of a stem and progenitor cell (SPC) population using the established acute myelogenous leukemia cell line KG-1a as a cell model. The cells express a CD34 cell surface marker associated with the hematopoietic progenitor cell activity and the progenitor cell lineage commitment. The CD34 expression level is approximately an order of magnitude lower than that of the CD45 marker, which required further improvements of the magnetic fractionation apparatus. The cells were immunomagnetically labeled using a sandwich of anti-CD34 antibody-phycoerythrin (PE) conjugate and anti-PE magnetic nanobead and fractionated into eight components using a continuous flow dipole magnetophoresis apparatus. The CD34 marker expression distribution between sorted fractions was measured by quantitative PE flow cytometry (using QuantiBRITE PE calibration beads), and it was shown to be correlated with the cell magnetophoretic mobility distribution. A flow outlet addressing scheme based on the concept of the transport lamina thickness was used to control cell distribution between the eight outlet ports. The fractional cell distributions showed good agreement with numerical simulations of the fractionation based on the cell magnetophoretic mobility distribution in the unsorted sample.
    The Analyst 01/2010; 135(1):62-70. · 4.23 Impact Factor
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    ABSTRACT: Magnetic field-flow fractionation (MgFFF) is a technique for the separation and characterization of magnetic nanoparticles. It is explained that the analysis of polydisperse samples requires a programmed decay of field and field gradient during sample elution. A procedure for achieving reproducible field decay with asymptotic approach to zero field using a quadrupole electromagnet is described. An example of an analysis of a polydisperse sample under programmed field decay is given.
    Physics Procedia 01/2010; 9:91-95.
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    ABSTRACT: Magnetic microsphere suspensions undergo complex motion when exposed to finite sources of the magnetic field, such as small permanent magnets. The computational complexity is compounded by a difficulty in choosing a suitable choice of visualization tools because this often requires using the magnetic force vector field in three dimensions. Here we present a potentially simpler approach by using the magnetic pressure. It is a scalar quantity, pm = B2/2μ0, and its usefulness has been already demonstrated in applications to magnetohydrodynamics and ferrohydrodynamics (where B is the applied field and μ0 = 4π×10-7 T.m/A). The equilibrium distribution of the magnetic bead plug in aqueous suspension is calculated as an isosurface of the magnitude of the magnetic pressure pm = const, in the field of two permanent magnet blocks calculated from closed formulas. The geometry was adapted from a publication on the magnetic bead suspensions in microsystems and the predicted bead plug distribution is shown to agree remarkably well with the experiment.
    AIP Conference Proceedings 01/2010; 1311:111-117.
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    ABSTRACT: Magnetic deposition microscropy (MDM) combines magnetic deposition and optical analysis of magnetically tagged cells into a single platform. Our multistage MDM uses enclosed microfabricated channels and a magnet assembly comprising four zones in series. The enclosed channels alleviate the problem plaguing previous versions of MDM: scouring of the cell deposition layer by the air-liquid interface as the channel is drained. The four-zone magnet assembly was designed to maximize capture efficiency, and experiments yielded total capture efficiencies of >99% of fluorescent- and magnetically-labeled Jurkat cells at reasonable throughputs (10(3) cells/min). A digital image processing protocol was developed to measure the average pixel intensities of the deposited cells in different zones, indicative of the marker expression. Preliminary findings indicate that the multistage MDM may be suitable for depositing cells and particles in successive zones according to their magnetic properties (e.g., magnetic susceptibilities or magnetophoretic mobilities). The overall goal is to allow the screening of multiple disease conditions in a single platform.
    Analytical Chemistry 01/2009; 81(1):43-9. · 5.70 Impact Factor
  • 11/2007: pages 331-412; , ISBN: 978-0444527547
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    ABSTRACT: Positive selection of CD34+ blood progenitor cells from circulation has been reported to improve patient recovery in applications of autologous transplantation. Current magnetic separation methods rely on cell capture and release on solid supports rather than sorting from flowing suspensions, which limits the range of therapeutic applications and the process scale up. We tested CD34+ cell immunomagnetic labeling and isolation from fresh leukocyte fraction of peripheral blood (leukapheresis) using the continuous quadrupole magnetic flow sorter (QMS), consisting of a flow channel (SHOT, Greenville, IN) and a quadrupole magnet with a maximum field intensity (B(o)) of 1.42 T and a mean force field strength (S(m)) of 1.45 x 10(8) TA/m(2). Both the sample magnetophoretic mobility (m) and the inlet and outlet flow patterns highly affect the QMS performance. Seven commercial progenitor cell labeling reagent combinations were quantitatively evaluated by measuring magnetophoretic mobility of a high CD34 expression cell line, KG-1a, using the cell tracking velocimeter (CTV). The CD34 Progenitor Cell Isolation Kit (Miltenyi Biotec, Bergisch Gladbach, Germany) showed the strongest labeling of KG-1a cells and was selected for progenitor cell enrichment from 11 fresh and 11 cryopreserved clinical leukapheresis samples derived from different donors. The CD34+ cells were isolated with a purity of 60-96%, a recovery of 18-60%, an enrichment rate of 12-169, and a throughput of (1.7-9.3) x 10(4) cells/s. The results also showed a highly regular dependence of the QMS performance on the flow conditions that agreed with the theoretical predictions based on the CD34+ cell magnetophoretic mobility.
    Biotechnology and Bioengineering 05/2007; 96(6):1139-54. · 3.65 Impact Factor
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    ABSTRACT: To evaluate a negative selection technique for the hematopoietic progenitor cell enrichment from clinical leukapheresis product using continuous magnetophoresis. The leukapheresis product was labeled with a tetrameric antibody cocktail (TAC) and magnetic colloid against nonprogenitor leukocytes (StemSep enrichment cocktail kit, Stem Cell Technologies, Vancouver, Canada). The separation of hematopoietic progenitor cells was performed by flow-through magnetophoresis in an annular channel placed coaxially inside a quadrupole magnetic field, in a split-flow thin-cell fractionation configuration (referred to as quadrupole magnetic flow sorter, QMS). The TAC antibody cocktail and the magnetic colloid were titrated to determine minimum effective antibody and magnetic reagent concentrations by measuring cell magnetophoretic mobility (m) distribution using cell tracking velocimetry. Leukapheresis products from eight donors having initial CD34+ cell purity between 0.37 and 9.7% were enriched to the final purity of 30 to 85% and yield of 49 to 84% with a maximum throughput of 6.7 x 10(4) cells/s. The progenitor cell enrichment was accompanied by a more than 3.5 log(10) T-lymphocyte depletion, a significant factor considering the intended application to allogeneic transplantation. Cell colony-forming unit assays showed that there was no deterioration of progenitor cell proliferation and differentiation following the QMS enrichment process. The negative selection method of hematopoietic progenitor cells by continuous magnetophoresis is a promising approach to a process scale-up, important for clinical applications.
    Experimental Hematology 05/2007; 35(4):662-72. · 2.91 Impact Factor
  • 01/2007;
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    ABSTRACT: Cell separation is important in medical and biological research and plays an increasingly important role in clinical therapy and diagnostics, such as rare cancer cell detection in blood. The immunomagnetic labeling of cells with antibodies conjugated to magnetic nanospheres gives rise to a proportional relationship between the number of magnetic nanospheres attached to the cell and the cell surface marker number. This enables the potential fractionation of cell populations by magnetophoretic mobility (MM). We exploit this feature with our apparatus, the Dipole Magnet Flow Fractionator (DMFF), which consists of an isodynamic magnetic field, an orthogonally-oriented thin ribbon of cell suspension in continuous sheath flow, and ten outlet flows. From a sample containing a 1:1 mixture of immunomagnetically labeled (label+) and unlabeled (label-) cells, we achieved an increase in enrichment of the label+ cell fraction with increasing outlet numbers in the direction of the magnetic field gradient (up to 10-fold). The total recovery of the ten outlet fractions was 90.0+/-7.7%. The mean MM of label+ cells increased with increasing outlet number by up to a factor of 2.3. The postulated proportionality between the number of attached magnetic beads and the number of cell surface markers was validated by comparison of MM measured by cell tracking velocimetry (CTV) with cell florescence intensity measured by flow cytometry.
    Journal of Biochemical and Biophysical Methods 08/2006; 68(1):1-21. · 2.33 Impact Factor
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    ABSTRACT: During intra-erythrocytic development, malaria trophozoites digest hemoglobin, which leads to parasite growth and asexual replication while accumulating toxic heme. To avoid death, the parasite synthesizes insoluble hemozoin crystals in the digestive vacuole through polymerization of beta-hematin dimers. In the process, the heme is converted to a high-spin ferriheme whose magnetic properties were studied as early as 1936 by Pauling et al. Here, by magnetophoretic cell motion analysis, we provide evidence for a graduated increase of live cell magnetic susceptibility with developing blood-stage parasites, compatible with the increase in hemozoin content and the mechanism used by P. falciparum to avoid heme toxicity. The measured magnetophoretic mobility of the erythrocyte infected with a late-stage schizont form was m = 2.94 x 10(-6) mm3 s/kg, corresponding to the net volume magnetic susceptibility (relative to water) of Deltachi = 1.80 x 10(-6), significantly higher than that of the oxygenated erythrocyte (-0.18x10(-6)) but lower than that of the fully deoxygenated erythrocyte (3.33x10(-6)). The corresponding fraction of hemoglobin converted to hemozoin, calculated based on the known magnetic susceptibilities of hemoglobin heme and hemozoin ferriheme, was 0.50, in agreement with the published biochemical and crystallography data. Magnetophoretic analysis of live erythrocytes could become significant for antimalarial drug susceptibility and resistance determination.
    The FASEB Journal 05/2006; 20(6):747-9. · 5.70 Impact Factor
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    ABSTRACT: A spatially uniform magnetic energy density gradient (∇B2) distribution offers a controlled environment to separate magnetically tagged cells or biomolecules based on their magnetophoretic mobility [ L. R. Moore et al., J. Biochem. Biophys. Methods 37, 11 (1998) ]. A design to obtain a uniform ∇B2 distribution for a microelectromechanical-systems-based magnetic cell separator was developed. The design consists of an external magnetic circuit and a microfabricated channel (biochip) with embedded discrete pole pieces on the channel walls. The two-dimensional and three-dimensional magnetostatic simulation softwares utilizing boundary element methods were used to optimize the positions and the dimensions of the discrete pole pieces, as well as the external magnetic circuit—the combination of which would generate a uniform ∇B2 profile over the channel cross section. It was found that the discrete pole pieces required specific magnetic properties (saturation magnetization constant >1.55 T) to affect the overall ∇B2 distribution. Investigating different positions of the discrete pole pieces inside the external magnetic field indicated that the proposed design could generate uniform ∇B2 distribution with ±100 μm displacements along the height/width and ±1° inclination from the optimum position.
    Journal of Applied Physics 04/2006; 99(8):08R905-08R905-3. · 2.21 Impact Factor
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    ABSTRACT: The need for innovative separative techniques suitable for the fractionation of biomaterials prompted this investigation into the performance of the gravitational split-flow thin channel (G-SPLITT) system as a cell sorter. The rigorous mathematical description of the separation mechanism allows achievement of fast separation of several million myeloma cells from healthy splenocytes using flow conditions calculated from theory. Separation in G-SPLITT is based on differences in sedimentation rate. For accurate prediction of the optimal working conditions, this parameter was directly measured by cell tracking velocimetry rather than relying on a measure of diameter (by Multisizer) and an assumed density for each cell population. We also discuss the influence of different flow conditions on the effectiveness of separation.
    Analytical Chemistry 09/2005; 77(16):5294-301. · 5.70 Impact Factor
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    ABSTRACT: Continuous flow immunomagnetic separation is an attractive alternative to current batch mode immunomagnetic separation methods because it is capable of high sorting speeds at mild cell conditions, and grants the operator better control of separation process. The control of the separation is dependent on knowledge of the amount of magnetic label attached to the cell (magnetic labeling intensity), however. Determination of the magnetic labeling is accomplished by measuring cell magnetophoretic mobility using a newly developed technique of Cell Tracking Velocimetry (CTV). Flow cytometry was used to define the antibody binding characteristics of a fluorescently tagged primary antibody. Subsequently, CTV was used to measure antibody-binding characteristics of a magnetically tagged secondary antibody. The results of this study show that CTV is capable of providing valuable information concerning the cell labeling by magnetically tagged antibodies. It was demonstrated that the magnetically conjugated antibody binding curve exhibits the same exponential increase to saturation characteristics as that seen with the fluorescently tagged antibody. Further, it was shown that the intensity of the secondary magnetic labeling is directly proportional to the intensity of the primary fluorescent label. CTV is an accurate tool for evaluation of magnetically conjugated antibodies. The ability to determine the intensity of magnetic labeling is necessary for the development of continuous flow immunomagnetic separations based on cell magnetophoresis.
    Cytometry Part A 09/2005; 66(2):103-8. · 3.71 Impact Factor
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    ABSTRACT: Field-flow fractionation (FFF) is an analytical scale separation and characterizatio technique for macromolecules and particles. A quadrupole magnetic FFF device has bee constructed for analyzing magnetic nanoparticles. It is shown to give reproducible results and be capable of distinguishing between different lots of a commercial magnetic nanoparticle material.
    Journal of Physics Conference Series 08/2005; 17(1):174.
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    ABSTRACT: Magnetic micro and nanoparticles conjugated to affinity labels have become a significant, commercial reagent. It has been demonstrated that the performance of cell separation systems using magnetic labels is a function of the magnitude of the magnetic force that can be generated through labeling. This magnetic force is proportional to the number of magnetic particles bound to the cell, the magnetic energy gradient, and the particle-field interaction parameter. This particle-field interaction parameter, which is the product of the relative volumetric, magnetic susceptibility and the volume of the micro or nanoparticle, is a fundamental parameter which can be used to characterize the magnetic particles. An experimental technique is presented which measures the volumetric magnetic susceptibility of particles through the use of susceptibility modified solutions and an experimental instrument, Cell Tracking Velocimetry, CTV. Experimental studies were conducted on polystyrene microspheres alone and those bound to four different magnetic nanoparticles. The experimentally determined values of the magnetic susceptibility of the polystyrene microspheres are consistent with values found from literature. Consequently, magnetic susceptibility measurements of these polystyrene microspheres bound with the magnetic nanoparticles combined with particle size measurements using commercial dynamic light scattering instrument allowed estimates of the particle-field interaction parameter to be made for four commercial, magnetic nanoparticles. The value found for MACS beads is close to what is reported from an independent study. The values for MACS beads and Imag beads are found to agree with what is observed from experiments. Finally, an experimental demonstration of the impact that differences in this field interaction parameter has on the labeling of human lymphocytes is presented.
    The Analyst 05/2005; 130(4):514-27. · 3.97 Impact Factor

Publication Stats

641 Citations
278 Downloads
2k Views
112.41 Total Impact Points

Institutions

  • 2013
    • Mie University
      • Department of Chemistry for Materials
      Tsu-shi, Mie-ken, Japan
  • 2006–2013
    • Lerner Research Institute
      Cleveland, Ohio, United States
    • Cleveland Clinic
      • Department of Biomedical Engineering
      Cleveland, OH, United States
  • 2005
    • Sapienza University of Rome
      • Department of Chemistry
      Roma, Latium, Italy
  • 1996–2005
    • The Ohio State University
      • William G. Lowrie Department of Chemical and Biomolecular Engineering
      Columbus, OH, United States
  • 2000
    • Bar Ilan University
      • Department of Chemistry
      Ramat Gan, Tel Aviv, Israel
  • 1999
    • Case Western Reserve University
      • Department of Biomedical Engineering
      Cleveland, OH, United States