Jun Pyo Kim

Ultrasensitive carbon nanotube-based biosensors using antibody-binding fragments

We report a method to build ultrasensitive carbon nanotube-based biosensors using immune binding reaction. Here carbon nanotube-field effect transistors (CNT-FETs) were functionalized with antibody-binding fragments as a receptor, and the binding event of target immunoglobulin G (IgG) onto the fragments was detected by monitoring the gating effect caused by the charges of the target IgG. Because the biosensors were used in buffer solution, it was crucial to use small-size receptors so that the charged target IgG could approach the CNT surface within the Debye length distance to give a large gating effect. The results show that CNT-FET biosensors using whole antibody had very low sensitivity (detection limit approximately 1000 ng/ml), whereas those based on small Fab fragments could detect 1 pg/ml (approximately 7 fM level). Moreover, our Fab-modified CNT-FET could successfully block the nontarget proteins and could selectively detect the target protein in an environment similar to that of human serum electrolyte. Significantly, this strategy can be applied to general antibody-based detection schemes, and it should enable the production of label-free ultrasensitive electronic biosensors to detect clinically important biomarkers for disease diagnosis.

Enhancement of sensitivity and specificity by surface modification of carbon nanotubes in diagnosis of prostate cancer based on carbon nanotube field effect transistors

This paper presents a simple and sensitive method for the real-time detection of a prostate cancer marker (PSA-ACT complex) through label-free protein biosensors based on a carbon nanotube field effect transistor (CNT-FET). Herein, the CNT-FET was functionalized with a solution containing various linker-to-spacer ratios, the binding event of the target PSA-ACT complex onto the receptor detected by monitoring the gating effect caused by charges in the target PSA-ACT complex. Since the biosensors were used in a buffer solution, it was crucial to control the distance between the receptors through introduction of linkers and spacers so that the charged target PSA-ACT complex could easily approach the CNT surface within the Debye length to give a large gating effect. The results show that CNT-FET biosensors modified with only linkers could not detect target proteins unless a very high concentration of the PSA-ACT complex solution (approximately 500 ng/ml) was injected, while those modified with a 1:3 ratio of linker-to-spacer could detect 1.0 ng/ml without any pretreatment. Moreover, our linker and spacer-modified CNT-FET could successfully block non-target proteins and selectively detect the target protein in human serum. Significantly, this strategy can be applied to general antibody-based detection schemes and enables production of very simple and sensitive electronic biosensors to detect clinically important biomarkers for disease diagnosis.

Hydrogen production of the hyperthermophilic eubacterium, Thermotoga neapolitana under N2 sparging condition.

Gas sparging was found to be a useful technique to reduce hydrogen partial pressure in the liquid phase to enhance the hydrogen yields of strictly anaerobically fermentative bacteria. The effect of nitrogen (N(2)) sparging on hydrogen yield was investigated in sterile and non-sterile conditions using a pure strain of the hyperthermophilic eubacteria, Thermotoga neapolitana with glucose or xylose as a carbon source. The maximum hydrogen accumulations reached 41% of the gaseous mixtures after 30-40 h. Two applications of N(2) sparging after the H(2) content in the headspace reached the maximum levels gave an increase of H(2) production by 78% from 1.82 to 3.24 mol H(2)/mol glucose and by 56% from 1.41 to 2.20 mol H(2)/mol xylose. This result suggested that the removal of the produced H(2) from the gas headspace of the limited-volume, closed culture vial when it achieves the maximum level of H(2) tolerance of the bacterium is a necessary technique to improve its H(2) yield.

Enhancement of sensitivity using hybrid stimulus for the diagnosis of prostate cancer based on polydiacetylene (PDA) supramolecules

In this study, hybrid stimulus was initially introduced to improve the sensitivity of PDA vesicle chip for detection of prostate-specific antigen-α1-antichymotrypsin (PSA-ACT) complex. The strategy of hybrid stimulus on PDA vesicle chip offers the amplification method of fluorescent signal which combines a primary response by the immune reaction of antigen-antibody and a secondary response by the mechanical pressure of pAb-conjugated magnetic beads. As the primary response result on PDA vesicle chip, the PSA-ACT complex in PBS buffer was detected at 10 ng/mL. However, this detection sensitivity was insufficient for diagnosis of prostate cancer because the normal human PSA concentration is less than 4.0 ng/mL. To solve this problem, polyclonal PSA antibody-conjugated magnetic beads were used as an amplifying agent after primary immunoresponse. As a result, the PSA-ACT complex concentrations (as low as 0.1 ng/mL) could be detected in the PBS buffer sample. Therefore, this result can be applied to various fields, such as the detection of cells, proteins, and DNA for sensitive and specific biosensing based on PDA supramolecules.

Rigiflex lithography-based nanodot arrays for localized surface plasmon resonance biosensors

We present a facile and robust means of fabricating metallic nanodot arrays for localized surface plasmon resonance (LSPR) biosensors through the strategic coupling of a polymeric template prepared with rigiflex lithography and a subsequent metallization via electrodeposition. Rigiflex lithography provides the capability to realize large-scale nanosized features as well as process flexibility during contact molding. In addition, the electrodeposition process enables wet-based nanoscale metallization with high pattern fidelity and geometric controllability. Generated metallic nanodot arrays can be used as a general platform for LSPR biosensors via the sequential binding of chemicals and biomolecules. Extinction spectra of the corresponding LSPR signal are measured with UV-vis-NIR spectroscopy, from which the pattern size and shape dependence of LSPR are readily confirmed. The feasibility of a very sensitive biosensor is demonstrated by the targeted binding of human immunoglobulin G, yielding subnanomolar detection capability with high selectivity.

The strategy of signal amplification for ultrasensitive detection of hIgE based on aptamer-modified poly(di-acetylene) supramolecules

Herein, we demonstrate three strategies of signal amplification for ultrasensitive detection of human immunoglobulin E (hIgE) based on poly(di-acetylene) supramolecules. To fabricate the ultrasensitive PDA biosensor, ethylenediamine as an interlinker and aptamer as a receptor were introduced into the chip fabrication process. Using the prepared PDA liposome biosensor, the hIgE could be detected up to below 1.0 ng/ml by a primary response. In order to accomplish more ultrasensitive detection of protein on a PDA biosensor, polyclonal hIgE antibody was employed as an external mechanical force for the inducement of a secondary response. As a result, a PDA liposome biosensor sensitivity as high as 0.01 ng/ml for the target hIgE was obtained, with a sensitivity which is one hundred times of that of the method without signal amplification. These results indicate that the proposed strategies were capable of ultrasensitive quantitative and qualitative analyses of biomolecules without non-specific binding of non-target proteins.