A compact microfluidic chip with integrated impedance biosensor for protein preconcentration and detection

Abstract

In this study, a low-cost, compact biochip is designed and fabricated for protein detection. Nanofractures formed by self-assembled gold nanoparticles at junction gaps are applied for ion enrichment and depletion to create a trapping zone when electroosmotic flow occurs in microchannels. An impedance measurement module is implemented based on the lock-in amplifier technique to measure the impedance change during antibody growth on the gold electrodes which is caused by trapped proteins in the detection region. The impedance measurement results confirm the presence of trapped proteins. Distinguishable impedance profiles, measured at frequencies in the range of 10-100 kHz, for the detection area taken before and after the presence of proteins validate the performance of the proposed system.

FIG. 1.

Schematic of the proposed EEE-based protein concentration with integrated impedance sensing electrodes for protein detection. (a) Highly sensitive lock-in amplifier technique is employed for quantitatively recognizing proteins at the designated detection window. (b) Details of microfluidic channels and gold nanoparticle region.

FIG. 2.

Process of protein preconcentration based on EEE and electroosmotic flow. (a) Nanofractures generated by applying 50 VDC to two inlets and 0 V to other inlets. (b) Nanodepletion zone formation. (c) Electroosmotic flow activation by bias voltage +V. (d) Protein concentration by the depletion zone and electroosmotic flow after bias voltage application.

FIG. 3.

Impedance biosensor based on the impedance measurement method. (a) Model of the impedance biosensor based on the impedance measurement method. The impedance includes the surface impedance and resistance (from PBS). Gold electrodes and PBS form surface impedance, which consists of capacitance C_f and resistance R_f. R_s is the resistivity of PBS solution. (b) Equivalent circuit of the biosensor consisting of surface impedance and solution resistance. (c) Block diagram of the impedance measurement module. A microcontroller is used to adjust the frequency and phase through two AD 9805 function generator ICs and for data acquisition. The impedance measurement is implemented using IC OPA2350 for the balance-bridge conditioning circuit. AD 630 is used for analog lock-in amplification. (d) An actual image of the printed circuit board of the impedance measurement module.

FIG. 4.

Protein distribution when DC voltages are applied. Proteins are concentrated at the designated detection window after 30 min. (a) The protein is trapped in the detection region after supplying 0 V, 50 DC voltage, and 48 DC voltage. The trap works based on depletion force and EOF. (b) Successful protein preconcentration obtained after 30 min low DC voltages were supplied to the microfluidic channels.

FIG. 5.

Impedance of the biosensor before and after protein injection in the detection region. (a) Impedance profiles of four biochips after antibody immobilization on the electrode surface (the amplitude of the exciting signal is 600 mV_p-p). (b) Presence of targeted proteins confirmed by the impedance change before and after protein injection and incubation in 20 min (measured with an amplitude of the exciting signal of 600 mV_p-p). (c) Impedance change measured at 50 kHz and 600 mV_p-p amplitude of the exciting signal for four prototypes before and after protein injection and incubation in 20 min. Distinguishable lines confirm the presence of the targeted protein. (d) Impedance versus time used to investigate proteins captured on electrodes in the case where 100 mV_p-p amplitude of the exciting signal is applied. Profiles were captured before and after protein immobilization and after 10 and 20 min after proteins were trapped on electrodes.

FIG. 6.

Impedance change during antibody binding and protein trapping on gold electrodes. (a) Difference in the impedance change of the biochip before and after antibody growth on gold electrodes and after protein preconcentration. (b) Change in the imaginary part of biosensor impedance after antibody binding on gold electrodes and after protein trapping by antibodies.

FIG. 7.

Serial capacitance of biosensor at multiple frequencies when PBS solution fully filled the channel, antibodies were on electrodes, and proteins were trapped.