Development of Techniques for Rapid Isolation and Separation of Particles in Digital Microfluidics

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PhD DissertationDevelopment of Techniques for
Rapid Isolation and Separation of Particles in
Digital Microfluidics

• By
• Mr. H. Rezaei Nejad
• Supervisor
• Dr. Mina Hoorfar

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Motivations for microfluidics
Test tubes
Robotics
Lab-on-a-chip
http//www.slideshare.net/bioflux/launching-digita
l-biology-48300995
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Lab-on-a-chip
http//www.discoveriesinhealthpolicy.com/2015/12/l
arrys-kricka-20-page-review-of-history.html
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Microfluidics

• Continuous microfluidics
• Processing fluid in a small scale
• Reduces sample and reagent consumption
• Enhances the reaction time
• Segmented-flow microfluidics
• Liquid segmentation
• Eliminates the diffusion and dispersion
• effect during flow
• Improves mixing
• Reduces reagent consumption
• Enables processing of a large number
• of reactions
• Digital microfluidics
• Operational segmentation
• Provides a programmable platform
• Provides a reconfigurable platform

https//www.youtube.com/watch?v5QVwljd04Kw
Evolution of microfluidic systems
https//www.youtube.com/watch?vxJxtQIyisns
https//www.youtube.com/watch?vxJxtQIyisns
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Digital microfluidics

• Processing fluid in a small scale
• Reduces sample and reagent consumption
• Enhances the reaction time
• Liquid segmentation
• Eliminates the diffusion and dispersion
• effect during flow
• Improves mixing
• Reduces reagent consumption
• Enables processing of a large number
• of reactions
• Operational segmentation
• Provides a programmable platform
• Provides a reconfigurable platform
• Enables parallel processing

https//www.youtube.com/watch?vxJxtQIyisns
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Digital microfluidics (DMF)
electrowetting on dielectric (EWOD)
Droplets moved by applying voltage to adjacent
cell
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Basic DMF operators
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Droplet with bio-particles

• They are not complex enough to control the
  materials (like solid particles) inside the
  droplets

As a result, it is not possible to scale down a
laboratory process on DMF platform that includes
particle or cell isolation at any of its steps.
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Background

• Droplet based devices

-Certain particles -Needs multiple channel
-Magnet particles -Moving part
-Charged particles
cross contamination
too complex
Magnetic collection and Separation for
EWOD Gaurav J. Shah 2009
Electrophoresis
Traveling-wave Dielectrophoresis (twDEP) Yuejun
Zhao 2007
Binary separation of micro particles Sung Kwon
Cho 2007
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Motivation
There is a need for the development of techniques
to control the particle motion inside the droplet
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Objectives

• Development and integration of reliable operators
  on digital microfluidics to manipulate, position,
  and separate micron size colloidal particles and
  biological cells

(1) Magnetic (2) Hydrodynamic
(3) Dielectrophoresis
particles with magnetic properties
wide range of particles but low conductivity
solution
non-buoyant particles
Project 2 (focusing) ?detection of low DNA
concentration
Project 3 (patterning) Project 4 (focusing) ?
cell/particle patterning droplet purification
Project 1 (isolation) ?purification of human
saliva DNA
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Project 1

• Magnetic collection of particles

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Magnetic collection
A neodymium round magnet integrated under the
bottom plate of digital microfluidic chip
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DNA purification with magnetic separation
technique
(a)
(b)
(c)
S
S sample droplet W washing buffers E elution
buffer
Magnet
E
DNA capturing and particle isolation (a)-(c)
W
W
(d)
(e)
(f)
Washing 2 (removing debris from the
beads) (d)-(f)
Steps
(g)
(h)
(i)
Elution (removing the DNA from the
beads) (g)-(h)
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On-chip vs. off-chip

• Different dilution of the initial samples
• 10X, 50X, 100X

1000 times less sample consumption 10 times
faster process (on-chip 3 mins off-chip 30
mins)
final DNA concentration
Purity of the final DNA sample
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Project 2

• Hydrodynamic particle separation

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Hydrodynamic focusing of particles
Spinning the droplet around a central electrode
in a controlled fashion. (each surrounding
electrodes are actuated for 125 ms)
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Initial Results
1-µm silica particle
non-buoyant
5-µm silica particle
5-µm polystyrene particle
buoyant
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Droplet Hydrodynamics (Physics)
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Geometry of central electrode
1 mm
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Particle size/density
Star design
Square design
Silica density 2.5 gr/cm3
Polystyrene density 1.05 gr/cm3
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Droplet volume
Shape of the focused region
Particle concentration
Total captured particles
Polystyrene beads (15 µm)
Silica beads (5 µm)
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Application 1Desired particle concentration
Creating droplets with different concentration of
beads on the DMF chip.
b
a
2 mm
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Application 2Particle indicator
Droplet with just 5µm polystyrene particles
Droplet with 5µm PS particles and less than 1
(of total particles) 15µm PS particles
1 mm
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Application 3Detection of very low concentration
of DNA
15µm PS particles with 18 ng/µl DNA in the
solution
15µm PS particles without DNA in the solution
1 mm
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Project 3

• Dielectrophoretic-based positioning and
  patterning of particles

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DEP on DMF
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DEP traps on DMF
1. Particle patterning
Electric field profile across the traps
2. Particle sorting
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DEP manipulation of particles on DMF
1. Particle patterning
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2. Particle sorting
Transient response
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Numerical / experimental studies
S
Single particle patterning
W
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Application Cell sorting and patterning
200 µm
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Project 4

• Dielectrophoretic-based manipulation and focusing
  of particles

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DEP manipulation triangular traps
1
Particles line up in the trap
They march toward the base of the triangle
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Effect of gap/trap vertex angle
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Numerical study
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ApplicationDroplet purification
The purification efficiency of 90 is readily
achieved
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Contributions

• A robust on-chip protocol has been developed to
  purify DNA on DMF platforms. The protocol
  effectively reduces sample consumption by 1000
  times.
• A novel hydrodynamic-based technique has been
  developed for the DMF platform. The technique
  traps particle by only actuating the droplet in a
  controlled fashion using EWOD.
• DEP has been implemented into the DMF platform
  for patterning particles and live-cells. The
  technique was optimized to pattern a single
  particle on DMF.
• Novel DEP trap geometries has been developed and
  implemented into DMF for capturing and
  manipulating the particles toward a predefined
  direction.

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Future work

• Sample preparation unit on DMF platform
• Particle focusing based detection
• DMF based bioreactor for cell study
• Blood-on-a-chip

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Peer-reviewed journal publications

• 1 H. Rezaei Nejad, Z. Goli, M. Kazemzadeh
  Narbat, N. Annabi, Y. Shrike Zhang, M. Hoorfar,
  A. Tamayol, A. Khademhosseini, Laterally
  confined micromolding for spatially defined
  vascularization, submitted to Small (January
  2016). (Lead author, generating the basic idea,
  experiment and fabrication of the device, writing
  the paper), Impact Factor 8.368.
• 2 H. Rezaei Nejad, E. Samiei, A. Ahmadi, M.
  Hoorfar, Gravity-driven hydrodynamic particle
  separation in digital microfluidic systems, RSC
  Advances, 5 (45), 35966-35975, 2015. (Lead
  author, generating the basic idea, experiment and
  fabrication of the device, writing the paper),
  Impact Factor 3.84.
• 3 H. Rezaei Nejad, M. Hoorfar, Purification of
  a droplet using negative dielectrophoresis traps
  in digital microfluidics, Microfluidics and
  Nanofluidics, 18 (3), 483-492, 2014. (Lead
  author, generating the basic idea, experiment and
  fabrication of the device, writing the paper),
  Impact Factor 2.528.
• 4 H. Rezaei Nejad, O. Z. Chowdhury, M. D. Buat,
  M. Hoorfar, Geometrical characterization of
  negative DEP for particle trapping on digital
  microfluidics platform, Lab chip, 13 (9),
  1823-130, 2013. (Lead author, generating the
  basic idea, experiment and fabrication of the
  device, writing the paper), Impact Factor 6.115.
• 5 E. Samiei, H. Rezaei Nejad, M. Hoorfar, "A
  novel particle focusing technique based on the
  cumulative effects of gravity and
  dielectrophoresis for digital microfluidics",
  Appl. Phys. Letter, 106 (20), 204101, 2015.
  (Co-author, contributed to the development of the
  basic idea and fabrication of the device, and
  participated in writing the paper), Impact
  Factor 3.302.
• 6 M. Paknahad, H. Rezaei Nejad, M. Hoorfar,
  "Development of a Digital Micropump with
  Controlled Flow Rate for Microfluidic Platforms."
  Sensors Transducers, 183 (12), 1726-5479, 2014.
  (Co-author, contributed to the development of the
  basic idea and fabrication of the device, and
  participated in writing the paper), Impact
  Factor 0.75.

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Conference proceedings

• 1 H. Rezaei Nejad, M. Hoorfar, R. Samanipour,
  W. Zongjie, K. Kim, M. Hoorfar, Cell-Patterning
  and Culturing on Digital Microfluidics (DMF),
  µTAS 2015, Gyeongju, Korea. (Lead author,
  developing the basic idea, fabrication of the
  device).
• 2 E. Samiei, H. Rezaei Nejad, M. Hoorfar, A
  novel density-based dielectrophoretic particle
  focusing technique for digital microfluidics,
  IEEE MEMS 2015, January, Portugal. (Co-author,
  contributed in the development of the basic idea
  and device fabrication, and participated in
  writing the paper)
• 3 H. Rezaei Nejad, E. Samiei, A. Ahmadi, M.
  Hoorfar, Hydrodynamic Density-Based Particle
  Focusing in digital Microfluidic Systems, µTAS
  2014, Texas, US. (Lead author, generating the
  basic idea, experiment and fabrication of the
  device, writing the paper)
• 4 H. Rezaei Nejad, E. Samiei, M. Hoorfar,
  Droplet Dispensing From Open to Close Digital
  Microfluidics, Microtech 2014, Washington DC,
  US. (Lead author, generating the basic idea,
  experiment, writing the paper).
• 5 H. Rezaei Nejad, M. Paknahad, M. Hoorfar,
  Droplet Actuation on a Digital Microfluidic Chip
  Using a Portable DC Voltage Source, Microtech
  2014, Washington DC, US. (Lead author, generating
  the basic idea, experiment and fabrication of the
  device, writing the paper).
• 6 H. Rezaei Nejad, M. Paknahad, M. Hoorfar,
  Microtech 2014, Indirect Pumping of a Droplet in
  a Microfluidic Channel on a DMF Platform,
  Washington DC, US. (Lead author, generating the
  basic idea, experiment and fabrication of the
  device, writing the paper).
• 7 E. Samiei, H. Rezaei Nejad, M. Hoorfar,
  Effect of electrode geometry on droplet
  splitting in digital microfluidic platforms",
  ICNMM 2014, Chicago, US. (Co-author, contributed
  in the development of the basic idea and device
  fabrication, and participated in writing the
  paper).
• 8 O. Zaman, H. Rezaei Nejad, G. Sikander, M.
  Hoorfar, DNA Purification on Digital
  Microfluidics Platforms, CSME International
  Congress 2014, June1-4, Toronto, Ontario, Canada
  (Co-author, contributed in the development of the
  basic idea and device fabrication, and
  participated in writing the paper)
• 9 O. Zaman, H. Rezaei Nejad, M. D. Buat, M.
  Hoorfar, Digital Microfluidics a Possible
  Approach for Controlling Bimolecular Adsorption,
  ICNMM 2012, Puerto Rico. (Co-author, contributed
  in the development of the basic idea and device
  fabrication, and participated in writing the
  paper)

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Any Question?
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(No Transcript)
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Hydrodynamic approach(particle hydrodynamics)
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Scale of biological particles
Physical forces gravity, hydrodynamics
electromagnetic,
functionalize micro-particles
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Ferrites with a shell
The surface of a maghemite or magnetite magnetic
nanoparticle is relatively inert and does not
usually allow strong covalent bonds with
functionalization molecules. The silica shell can
be easily modified with various surface
functional groups via covalent bonds between
organo-silane molecules and silica shell.

• Higher chemical stability (crucial for biomedical
  applications)
• Narrow size distribution (crucial for biomedical
  applications)
• Higher colloidal stability since they do not
  magnetically agglomerate
• Magnetic moment can be tuned with the
  nanoparticle cluster size
• Retained superparamagnetic properties
  (independent of the nanoparticle cluster size)
• Silica surface enables straightforward covalent
  functionalization

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Particle Isolation
Hydrodynamic collection of particles

• Magnetic
• Separation

Dielectrophoresis collection of non-charged
particles
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Quantification and Parametrical Study

• Analytical study
• Predicting particle focusing behavior

• Image analysis technique development
• Calibration curves
• Image processing

• Studied parameters
• Actuation scheme
• Central electrode geometry
• Droplet volume
• Particle size/density

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Actuation Scheme
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Numerical / Experimental Studies

• Numerical Studies
• The physic is studied
• CM factor, Electric field, DEP force, Particle
  motion
• Optimization

• Experimental studies
• Droplet volume/gap ratio
• Efficiency
• Speed of the process
• Particle/electrode ratio

• Application
• Single particle trapping Droplet purification
  Changing particle concentration Cell
  patterning/sorting. (all on DMF)

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Patterning Kidney cancer cell
b
a
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Dielectrophoresis theory
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