The development of an automated system for electrical characterization of cells using a novel force balance method

Dielectrophoresis (DEP), a cell separation technique based on the dielectric properties, has significantly advanced biomedical research in diverse applications ranging from blood stem cells purification to cancer cells isolation from heterogeneous populations. The ability to accurately measure the dielectric properties of individual cells is not only critical for effective sorting applications, but is also advantageous for enhancing the current knowledge of cell biology. This thesis proposes a novel method: the n-DEP spring, which applies an electrical field gradient upon continuously flowing cells to distinguish them based on their individual DEP properties. Specifically, the method uses the equilibrium position originating from the force balance between hydrodynamic and DEP forces to infer the cellular dielectric properties. For thorough DEP characterization, changing different conditions of cells is an essential but time-consuming process which usually takes hours to days. Especially for DEP characterization of time-sensitive events, such as neutrophil activation or cell apoptosis, short characterization time is required. This thesis describes the automation of the fluidic, optics, and electronics components of the DEP characterization system, which shortens the characterization time within an hour. We first demonstrated the automated DEP characterization of a mammalian cell type in thirty-nine conditions within an hour. Subsequently, we characterized the neutrophils with different activation states and successfully found out the right conditions to discriminate the activated neutrophils and non-activated neutrophils. With this system and method, we now have the potential to rapidly screen through a variety of system parameters, and optimize conditions for effective cell sorting. Thesis Supervisor: Joel Voldman Title: Associate Professor of Electrical Engineering and Computer Science 3

[1]  H. Amini,et al.  Label-free cell separation and sorting in microfluidic systems , 2010, Analytical and bioanalytical chemistry.

[2]  Koji Asami,et al.  Characterization of heterogeneous systems by dielectric spectroscopy , 2002 .

[3]  Hermine Hitzler,et al.  Force measurements of optical tweezers in electro-optical cages , 1998 .

[4]  Yuri Feldman,et al.  Study of normal and malignant white blood cells by time domain dielectric spectroscopy , 2001 .

[5]  Jonghyun Oh,et al.  Comprehensive analysis of particle motion under non-uniform AC electric fields in a microchannel. , 2009, Lab on a chip.

[6]  Frederick F Becker,et al.  Microsample preparation by dielectrophoresis: isolation of malaria. , 2002, Lab on a chip.

[7]  Thomas Laurell,et al.  Acoustic whole blood plasmapheresis chip for prostate specific antigen microarray diagnostics. , 2009, Analytical chemistry.

[8]  F F Becker,et al.  Dielectric properties of human leukocyte subpopulations determined by electrorotation as a cell separation criterion. , 1999, Biophysical journal.

[9]  H Morgan,et al.  Analytical electric field and sensitivity analysis for two microfluidic impedance cytometer designs. , 2007, IET nanobiotechnology.

[10]  Siyang Zheng,et al.  Membrane microfilter device for selective capture, electrolysis and genomic analysis of human circulating tumor cells. , 2007, Journal of chromatography. A.

[11]  R. Pethig Review article-dielectrophoresis: status of the theory, technology, and applications. , 2010, Biomicrofluidics.

[12]  P. Renaud,et al.  Characterization and optimization of liquid electrodes for lateral dielectrophoresis. , 2007, Lab on a chip.

[13]  Michael D. Vahey,et al.  A microfluidic platform for the genome-wide analysis of electrical phenotype : physical theories and biological applications , 2010 .

[14]  J. Voldman,et al.  An equilibrium method for continuous-flow cell sorting using dielectrophoresis. , 2008, Analytical chemistry.

[15]  Y. Huang,et al.  Introducing dielectrophoresis as a new force field for field-flow fractionation. , 1997, Biophysical journal.

[16]  Noo Li Jeon,et al.  Unique Dielectric Properties Distinguish Stem Cells and Their Differentiated Progeny , 2008, Stem cells.

[17]  Chulhee Choi,et al.  Microfluidic self-sorting of mammalian cells to achieve cell cycle synchrony by hydrophoresis. , 2009, Analytical chemistry.

[18]  Dielectric cell response in highly conductive buffers. , 2012, Analytical chemistry.

[19]  R. Pethig,et al.  Differentiation of viable and non-viable bacterial biofilms using electrorotation. , 1995, Biochimica et biophysica acta.

[20]  R. Pethig,et al.  Separation of viable and non-viable yeast using dielectrophoresis. , 1994, Journal of biotechnology.

[21]  G. Kass,et al.  Rapid, automated measurement of dielectrophoretic forces using DEP‐activated microwells , 2011, Electrophoresis.

[22]  F F Becker,et al.  Differential analysis of human leukocytes by dielectrophoretic field-flow-fractionation. , 2000, Biophysical journal.

[23]  R. Tompkins,et al.  Equilibrium separation and filtration of particles using differential inertial focusing. , 2008, Analytical chemistry.

[24]  Mehmet Toner,et al.  Microfluidic system for measuring neutrophil migratory responses to fast switches of chemical gradients. , 2006, Lab on a chip.

[25]  G. Fuhr,et al.  Paired microelectrode system: dielectrophoretic particle sorting and force calibration , 1999 .

[26]  S. Takayama,et al.  Gravity-driven microfluidic particle sorting device with hydrodynamic separation amplification. , 2007, Analytical chemistry.

[27]  J. Wikswo,et al.  Microfluidic switching system for analyzing chemotaxis responses of wortmannin-inhibited HL-60 cells , 2008, Biomedical microdevices.

[28]  Zachary R. Gagnon,et al.  Cellular dielectrophoresis: Applications to the characterization, manipulation, separation and patterning of cells , 2011, Electrophoresis.

[29]  Jonathan Cohen The immunopathogenesis of sepsis , 2002, Nature.

[30]  Y. Huang,et al.  Dielectrophoretic separation of cancer cells from blood , 1995, IAS '95. Conference Record of the 1995 IEEE Industry Applications Conference Thirtieth IAS Annual Meeting.

[31]  S. Gawad,et al.  Single cell dielectric spectroscopy , 2007 .

[32]  N. Otsu A threshold selection method from gray level histograms , 1979 .

[33]  Christian H. Reccius,et al.  Leukocyte analysis and differentiation using high speed microfluidic single cell impedance cytometry. , 2009, Lab on a chip.

[34]  R. Hölzel,et al.  Electrorotation of single yeast cells at frequencies between 100 Hz and 1.6 GHz. , 1997, Biophysical journal.

[35]  B. Kirby,et al.  Automated dielectrophoretic characterization of Mycobacterium smegmatis. , 2011, Analytical chemistry.

[36]  Minoru Seki,et al.  Microfluidic devices for size-dependent separation of liver cells , 2007, Biomedical microdevices.

[37]  R. Pethig,et al.  CORRIGENDUM: Dielectric measurements using non-uniform electric field (dielectrophoretic) effects , 1977 .

[38]  Ronald Pethig,et al.  The removal of human leukaemia cells from blood using interdigitated microelectrodes , 1994 .

[39]  Michael P Hughes,et al.  Dielectrophoresis-activated multiwell plate for label-free high-throughput drug assessment. , 2008, Analytical chemistry.

[40]  Rudolf Höber,et al.  Eine Methode, die elektrische Leitfähigkeit im Innern von Zellen zu messen , 1910, Pflüger's Archiv für die gesamte Physiologie des Menschen und der Tiere.

[41]  R. Tompkins,et al.  A microfluidics approach for the isolation of nucleated red blood cells (NRBCs) from the peripheral blood of pregnant women , 2008, Prenatal diagnosis.

[42]  R. Pethig,et al.  The dielectrophoresis enrichment of CD34+ cells from peripheral blood stem cell harvests. , 1996, Bone marrow transplantation.

[43]  H. P. Schwan,et al.  Surface conductance and other properties of latex particles measured by electrorotation , 1987 .

[44]  M. Stelzle,et al.  Numerical modelling and measurement of cell trajectories in 3‐D under the influence of dielectrophoretic and hydrodynamic forces , 2011, Electrophoresis.

[45]  S Takashima,et al.  Dielectric properties of mouse lymphocytes and erythrocytes. , 1989, Biochimica et biophysica acta.

[46]  Ulrich Zimmermann,et al.  Electro-rotation: development of a technique for dielectric measurements on individual cells and particles , 1988 .

[47]  Francis Lin,et al.  Generation of dynamic temporal and spatial concentration gradients using microfluidic devices. , 2004, Lab on a chip.

[48]  Emmanuel Picard,et al.  Determination of Clausius–Mossotti factors and surface capacitances for colloidal particles , 2011 .

[49]  H. P. Schwan,et al.  Linear and nonlinear electrode polarization and biological materials , 2006, Annals of Biomedical Engineering.

[50]  Thomas B. Jones,et al.  Electromechanics of Particles , 1995 .

[51]  A. Irimajiri,et al.  A dielectric theory of "multi-stratified shell" model with its application to a lymphoma cell. , 1979, Journal of theoretical biology.

[52]  Michael D. Vahey A novel method for the continuous separation of microorganisms based on electrical properties , 2006 .