1. Pencil lead microelectrode and the application on cell dielectrophoresis Name:Tsung-Han Lin Teacher:Pofessor Hsu Class:Introduction to the Nano-electromechanical Systems Date: Nov. 4

2. Outline Introduction Experimental Results Conclusions

3. Introduction Dielectrophoresis (DEP) refers to the migration of dielectric particles under the net force (F) exerted by an asymmetric electric field: F=2r3εmRe[K(ω)] ∇ E2 where F is the dielectrophoretic force; r is the radius of the particle;εm is the permittivity of the medium; ∇ E is the gradient of the root mean square of the electric field strength; Re[K(ω)] is the real part of Clausius–Mossotti factor.

4. Migration towards high electric field regions was defined as positive DEP, whilst the movement away from such regions is the negative one. Since the Clausius–Mossotti factor is a function of frequency, it is possible to “tune” the direction and strength of the dielectrophoretic force by the frequency of the electric field for separating different particles

5. Manipulating micro-scaled particles such as cells by DEP was once considered to be difficult since the DEP force was too weak to overcome the Brownian motion in a device of regular size [3].Raising the applied voltage is a possible solution, but the additional joule heating and electrochemical problems restrict the bio-applications. Nowadays, using micro-devices, several low voltage DEP systems have been developed for separating bioparticles including viruses [4], proteins [5], and DNAs [6,7], but the fabrication of those microsystems required sophisticated instruments and procedures such as photolithography

6. Commercially available graphite pencil leads have been directly employed as the electrodes for various catalytic and analytical applications; it might be a convenient and inexpensive alternative material for other electrochemical applications. To our knowledge, there are only two studies concerning the miniaturization of graphite pencil leads by mechanical grinding and electrochemical etching]; however, the documented sizes are still too large (>40 m) for a low voltage cell DEP purpose. In the present approach, we propose a convenient and reproducible electrochemical etching procedure for sharpening a pencil lead tip to micro- scale in diameter. A DEP system based on the pencil lead microelectrode was then constructed and the manipulation of latex microbeads and human red blood cells were demonstrated.

7. Experimental A pencil lead (0.5 mm in diameter) 1.0 M NaOH aqueous An ac voltage (3.0 Vrms for 10 min) Polypropylene micro-pipette tip (2–200 L volume range) 2.0 mM ferricyanide/ferrocyanide buffer solution (pH 7.0) polystyrene latex microbeads (nominal diameter of 3 m) and human red blood cells conducting glass (indium tin oxide coated glass,40 mm × 40 mm ×1 mm) Frequency 20 Hz to 2 MHz Stainless Plate

8. Schematic diagram of the etching bath. Both graphite pencil lead and stainless plate were submerged into the solution with an angle of about 30 ◦ to avoid the bubbling effect.

9. Dielectrophoretic system configuration. The lower left inset is the photo of the DEP chamber with the microelectrode positioning at the center.

10. Results Under microscopic observation, 200 uL of latex microbeads or human whole blood sample solution with different conductivities were delivered into the DEP separation chamber. By adjusting the sinusoidal frequency from 20 Hz to 2 MHz, the range of positive DEP and negative DEP was identified as in Tables 1 and 2. For the latex microbeads, positive DEP separations occurred at low frequency ranges with low buffer conductivities. In contrast, the blood cells suffered positive DEP separations at high frequency ranges with high buffer conductivities.

11. By tuning the separation frequency at suitable experimental conditions, latex beads and human red blood cells can be attracted towards the microelectrode and then expelled away, the behaviors are similar to the published reports [38,39]. Microparticles can be easily manipulated with the simple microelectrode and experimental design

12. Latex microbeads in a buffer with conductivity of 10 S cm−1: after 30 s of 1 kHz positive DEP; after 60 s of 2 MHz negative DEP

13. human blood cells in a buffer with conductivity of 5 S cm−1: after 90 s of 1 MHz positive DEP after 90 s of 1 kHz negative DEP

15. Conclusions With a simple electrochemical etching process, our group has successfully fabricated a graphite pencil lead microelectrode with its diameter approaching 10um. The insulation for the microelectrode was achieved by shielding with a polypropylene layer. The low-cost microelectrode can be easily manufactured with a reproducible quality and high analytical performance. Equipped with a conductive glass as the counter electrode and the microscopic window, a simple dielectrophoretic chamber was designed using the mentioned microelectrode to generate an asymmetrical electric field. The economic and simple system was proven to be suitable for manipulating microparticles under direct microscopicobservation.

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