Kyong Hoon Lee

Multi-walled carbon nanotubes experiencing electrical breakdown as gas sensors

This paper presents a new approach to gas sensing using a multi-walled carbon nanotube (MWCNT) subject to electrical breakdown. The electrical resistances of large-diameter MWCNTs were found to decrease in the presence of air after experiencing electrical breakdown, while pristine MWCNTs were not appreciably sensitive. The sensitivity could be controlled by manipulating the level of the electrical breakdown, and larger-diameter MWCNTs showed better sensitivity because they possess more damaged shells that can create more adsorption sites for oxygen molecules. It was suggested by theoretical calculations that the oxygen sensitivity might be associated with an oxidized junction that exists between the outer and inner shells of the nanotubes.

Immunosensor towards low-cost, rapid diagnosis of tuberculosis

A rapid, accurate tuberculosis diagnostic tool that is compatible with the needs of tuberculosis-endemic settings is a long-sought goal. An immunofluorescence microtip sensor is described that detects Mycobacterium tuberculosis complex cells in sputum in 25 minutes. Concentration mechanisms based on flow circulation and electric field are combined at different scales to concentrate target bacteria in 1 mL samples onto the surfaces of microscale tips. Specificity is conferred by genus-specific antibodies on the microtip surface. Immunofluorescence is then used to detect the captured cells on the microtip. The detection limit in sputum is 200 CFU mL(-1) with a success rate of 96%, which is comparable to PCR.

Size-specific concentration of DNA to a nanostructured tip using dielectrophoresis and capillary action.

One of the critical challenges in the fields of disease diagnostics and environmental monitoring is to concentrate extracellular DNA from a sample mixture rapidly. Unlike genomic DNA in normal cells, extracellular DNA dissolved in a biological sample can potentially offer crucial information about pathogens and toxins. The current concentration methods, however, are not able to directly concentrate extracellular DNA due to aggressive sample preparation steps. This paper presents a concentration mechanism of extracellular DNA onto a nanostructured tip using dielectrophoresis (DEP) in conjunction with capillary action. DNA immersed in a solution is captured onto a nanotip by two sequential actions: (1) attraction of DNA and other bioparticles in the vicinity of a nanotip by DEP and (2) size-specific capture of DNA onto the nanotip by capillary action. To investigate the size-specific capturing mechanism, an analytical model for the capillary action on a nanotip is presented, which is compared to the experiment for capturing polystyrene nanospheres. This analysis predicts the capture of a spherical particle smaller than 0.39 times a nanotip diameter, whereas our experiment shows that polystyrene spheres smaller than 0.84 times a nanotip diameter are captured. This discrepancy can be caused by the increase of the capturing force due to attractive DEP force. In addition, the diameter of the captured spheres can be increased by other experimental conditions including the tip geometry, the multiple particle interaction, and the contact angles. When a nanotip is used for concentrating lambda-DNA, 6.7 pg/mL (210 aM) of DNA is selectively extracted from a sample mixture containing lambda-DNA and Drosophila cells in one minute. The captured DNA is investigated by fluorescence microscopy, scanning electron microscopy (SEM), and X-ray analysis. This nanotip-based DNA concentrating method is a rapid and highly sensitive technique to detect extracellular DNA from a sample mixture.

Microfluidic one-step synthesis of alginate microspheres immobilized with antibodies.

Micrometre- and submicrometre-size functionalized beads are frequently used to capture targets of interest from a biological sample for biological characterizations and disease diagnosis. The main challenge of the microbead-based assay is in the immobilization of probe molecules onto the microbead surfaces. In this paper, we report a versatile droplet microfluidics method to fabricate alginate microspheres while simultaneously immobilizing anti-Mycobacterium tuberculosis complex IgY and anti-Escherichia coli IgG antibodies primarily on the porous alginate carriers for specific binding and binding affinity tests. The binding affinity of antibodies is directly measured by fluorescence intensity of stained target bacteria on the microspheres. We demonstrate that the functionalized alginate microspheres yield specificity comparable with an enzyme-linked immunosorbent assay. The high surface area-to-volume ratio of the functionalized porous alginate microspheres improves the detection limit. By using the droplet microfluidics, we can easily modify the size and shape of alginate microspheres, and increase the concentration of functionalized alginate microspheres to further enhance binding kinetics and enable multiplexing.

Size-selective immunofluorescence of Mycobacterium tuberculosis cells by capillary- and viscous forces

Rapid, low cost screening of tuberculosis requires an effective enrichment method of Mycobacterium tuberculosis (MTB) cells. Currently, microfiltration and centrifugation steps are frequently used for sample preparation, which are cumbersome and time-consuming. In this study, the size-selective capturing mechanism of a microtip-sensor is presented to directly enrich MTB cells from a sample mixture. When a microtip is withdrawn from a spherical suspension in the radial direction, the cells that are concentrated by AC electroosmosis are selectively enriched to the tip due to capillary- and viscous forces. The size-selectivity is characterized by using polystyrene microspheres, which is then applied to size-selective capture of MTB from a sample mixture. Our approach yields a detection limit of 800 cells mL(-1), one of the highest-sensitivity immunosensors to date.

Dielectrophoretic concentration of low-abundance nanoparticles using a nanostructured tip

Electric field-induced concentration has the potential for application in highly sensitive detection of nanoparticles (NPs) for disease diagnosis and drug discovery. Conventional two-dimensional planar electrodes, however, have shown limited sensitivity in NP concentration. In this paper, the dielectrophoretic (DEP) concentration of low-abundance NPs is studied using a nanostructured tip where a high electric field of 3 × 10(7) V m(-1) is generated. In experimental studies, individual 2, 10, and 100 nm Au NPs are concentrated to a nanotip using DEP concentration and are detected by scanning transmission and scanning electron microscopes. The DEP force on Au NPs near the end of a nanotip is computed according to the distance, and then compared with Brownian motion-induced force. The computational study shows qualitative agreement with the experimental results. When the experimental conditions for DEP concentration are optimized for 8 nm-long oligonucleotides, the sensitivity of a nanotip is 10 aM (10 attomolar; nine copies in a 1.5 μl sample volume). This DEP concentrator using a nanotip can be used for molecular detection without amplification.

Electric field guided assembly of one-dimensional nanostructures for high performance sensors

Various nanowire or nanotube-based devices have been demonstrated to fulfill the anticipated future demands on sensors. To fabricate such devices, electric field-based methods have demonstrated a great potential to integrate one-dimensional nanostructures into various forms. This review paper discusses theoretical and experimental aspects of the working principles, the assembled structures, and the unique functions associated with electric field-based assembly. The challenges and opportunities of the assembly methods are addressed in conjunction with future directions toward high performance sensors.

Cryopreservation of Mycobacterium tuberculosis Complex Cells

ABSTRACT Successful long-term preservation of Mycobacterium tuberculosis cells is important for sample transport, research, biobanking, and the development of new drugs, vaccines, biomarkers, and diagnostics. In this report, Mycobacterium bovis bacillus Calmette-Guérin and M. tuberculosis H37Ra were used as models of M. tuberculosis complex strains to study cryopreservation of M. tuberculosis complex cells in diverse sample matrices at different cooling rates. Cells were cryopreserved in diverse sample matrices, namely, phosphate-buffered saline (PBS), Middlebrook 7H9 medium with or without added glycerol, and human sputum. The efficacy of cryopreservation was quantified by microbiological culture and microscopy with BacLight LIVE/DEAD staining. In all sample matrices examined, the microbiological culture results showed that the cooling rate was the most critical factor influencing cell viability. Slow cooling (a few degrees Celsius per minute) resulted in much higher M. tuberculosis complex recovery rates than rapid cooling (direct immersion in liquid nitrogen) (P < 0.05). Among the three defined cryopreservation media (PBS, 7H9, and 7H9 plus glycerol), there was no significant differential effect on viability (P = 0.06 to 0.87). Preincubation of thawed M. tuberculosis complex cells in 7H9 broth for 20 h before culture on solid Middlebrook 7H10 plates did not help the recovery of the cells from cryoinjury (P = 0.14 to 0.71). The BacLight LIVE/DEAD staining kit, based on Syto 9 and propidium iodide (PI), was also applied to assess cell envelope integrity after cryopreservation. Using the kit, similar percentages of “live” cells with intact envelopes were observed for samples cryopreserved under different conditions, which was inconsistent with the microbiological culture results. This implies that suboptimal cryopreservation might not cause severe damage to the cell wall and/or membrane but instead cause intracellular injury, which leads to the loss of cell viability.

Electrolyte-free amperometric immunosensor using a dendritic nanotip

Electric detection using a nanocomponent may lead to platforms for rapid and simple biosensing. Sensors composed of nanotips or nanodots have been described for highly sensitive amperometry enabled by confined geometry. However, both fabrication and use of nanostructured sensors remain challenging. This paper describes a dendritic nanotip used as an amperometric biosensor for highly sensitive detection of target bacteria. A dendritic nanotip is structured by Si nanowires coated with single-walled carbon nanotubes (SWCNTs) for generation of a high electric field. For reliable measurement using the dendritic structure, Si nanowires were uniformly fabricated by ultraviolet (UV) lithography and etching. The dendritic structure effectively increased the electric current density near the terminal end of the nanotip according to numerical computation. The electrical characteristics of a dendritic nanotip with additional protein layers was studied by cyclic voltammetry and I-V measurement in deionized (DI) water. When the target bacteria dielectrophoretically captured onto a nanotip were bound with fluorescence antibodies, the electric current through DI water decreased. Measurement results were consistent with fluorescence- and electron microscopy. The sensitivity of the amperometry was 10 cfu/sample volume (103 cfu/mL), which was equivalent to the more laborious fluorescence measurement method. The simple configuration of a dendritic nanotip can potentially offer an electrolyte-free detection platform for sensitive and rapid biosensors.

Microfabricated glucose sensor based on single-walled carbon nanotubes

This paper describes a novel glucose sensor that uses the hydrogen-specific gas sensing capability of single walled carbon nanotubes (SWCNTs) assembled on microelectrodes. Highly specific glucose sensing was demonstrated using buffered sample solutions with clinically significant concentrations. The proposed approach enables a simple but powerful bio-sensor reliably operating with a completely new principle, and opens up novel device applications where functional nano-components are integrated into a bioMEMS device.