A novel surface plasmon resonance imaging (SPRi) biosensor based on waveguide coupled surface pla... more A novel surface plasmon resonance imaging (SPRi) biosensor based on waveguide coupled surface plasmon resonance (WCSPR) measurements is reported. Simply fabricated by gold evaporation deposition and polymer spin-coating, the WCSPR chips can be easily adapted on an commercial SPRi monitoring instrument. The chips were designed by using Fresnel method and fabricated according to its optimized design, the calculation and experimental results fits well. Over an 8 mm × 8 mm rectangle imaging area, the WCSPR chips realized about 52% increase of sensitivity, 57% enhancement of resolution and 55% improvement of signal to noise ratio compared to conventional SPR chips. Furthermore, high throughput protein-protein interaction analyses based on WCSPR chips were successfully accomplished by applying Protein A and rabbit antibody interaction in an array format. The reflectivity responses observed from the WCSPR chips were about 50% higher than the conventional SPR chip while the binding kinetic parameters were within the same range as with conventional SPR chip. (J. Zhu). 1 Equal contribution, Zhiyou Wang designed and fabricated WCSPR chips, Lusheng Song performed the parameters analysis of the WCSPR chips and the applications for high-throughput bio-interaction measurements. 0925-4005/$ -see front matter His current study is focused on optical sensing, material fabrication process, research and application of new SPR structure to imaging technique. . His current study is focused on microfluidic devices and biosensors.
Accurate measurement of inter-peptide interactions is beneficial for in-depth understanding disea... more Accurate measurement of inter-peptide interactions is beneficial for in-depth understanding diseaserelated protein folding and peptide aggregation, and further for designing and selecting potential peptide drugs to the target antigen. Herein, we demonstrate a 3D polyrotaxane (PRX) surface for detecting peptides interactions by surface plasmon resonance imaging (SPRi). This surface is supramolecular selfassembly monolayer (SAM) structure fabricated by threading α-cyclodextrans (α-CD) through a linear polyethylene glycol (PEG) chain fixed on gold chip surface to form pseudopolyrotaxane, and further capping the pseudopolyrotaxane with bulky terminated group to form PRX film. The hydroxyl groups of α-CD can provide more active sites to increase molecules immobilization density, and PEG chain has unique protein non-fouling feature. We chose Alzheimer's disease marker β-amyloid 40 (Aβ40) as model peptide, and detected the interaction between it and its inhibitors KLVFFK6 by SPRi. As a striking result, the specific adsorption of KLVFFK6 solution at the concentration of 352 μM on Aβ40-PRX was 700 RU, whereas PEG SAM surface gave no significant binding. Interaction between other lower molecular weight peptides was detected via PRX surface, and the relatively weak interactions (K D ¼1.73 Â 10 À 4 M) between LPFFD (Mw ¼ 0.6 kDa) and amylin20-29 (Mw¼ 1.0 kDa) are successfully detected.
Surface plasmon resonance imaging (SPRi) has emerged as a versatile biosensor to detect a wide ra... more Surface plasmon resonance imaging (SPRi) has emerged as a versatile biosensor to detect a wide range of biomolecular interactions with divergent potential applications. However, the use of this advanced-level technology for stem cell lysate study is still not much explored. Cell lysates are significant biological analytes used for disease diagnostics and proteomic studies, but their complex nature limits their use as an analyte for SPRi biosensors. Here, we review the problems associated with the use of SPRi for stem cell lysate study and examine the role of surface chemistry, running buffer, and blocking solution in order to minimize nonspecific adsorption (NSA). We detect the expression of Oct4, Sox2, Nanog, Rex1, and Lin28 biomarkers present in mouse embryonic stem cell (mESC) lysate against their corresponding antibodies immobilized on the sensor surface with reduced NSA. The current study shows that the conjunction of SPRi and microarray can be used as a label-free, high-throughput, and rapid technique for detection of biomarkers and their relative abundance in stem cell lysate study.
In recent years, in situ protein synthesis microarray technologies have enabled protein microarra... more In recent years, in situ protein synthesis microarray technologies have enabled protein microarrays to be created on demand just before they are needed. In this paper, we utilized the TUS-TER immobilization technology to allow label-free detection with real-time kinetics of protein–protein interactions using surface plasmon resonance imaging (SPRi). We constructed an expression-ready plasmid DNA with a C-terminal TUS fusion tag to directionally immobilize the in situ synthesized recombinant proteins onto the surface of the biosensor. The expression plasmid was immobilized on the polyethylene imine-modified gold surface, which was then coupled with a cell-free expression system on the flow cell of the SPRi instrument. The expressed TUS fusion proteins bind on the surface via the immobilized TER DNA sequence with high affinity (3–7 ×10_13 M). The expression and immobilization of the recombinant in situ expressed proteins were confirmed by probing with specific antibodies. The present study shows a new low cost method for in situ protein expression microarrays that has the potential to study the kinetics of protein–protein interactions. These protein microarrays can be created on demand without the problems of stability associated with protein arrays used in the drug discovery and biomarker discovery fields.
The silver surface plasmon resonance (SPR) sensor has long been explored due to its intrinsic sen... more The silver surface plasmon resonance (SPR) sensor has long been explored due to its intrinsic sensitivity enhancement over the conventional single-layered gold SPR sensor. However, the silver SPR sensor has not been exploited for practical applications because of pronounced instability problems. We propose a novel gold−silver−gold trilayered SPR sensor chip, in which an extra buffer layer of gold is added between the silver and substrate adhesion layer (i.e., chromium) compared to the previously reported silver−gold bilayered SPR sensors. Subjected to prolonged agitation in phosphate-buffered saline (PBS) solution, the new chip exhibited high integrity according to both optical and atomic force microscopy (AFM) analysis. Having undergone repeated cycles of calibration, binding, and regeneration in various chemical solutions, 25 regions of interest (ROIs) over a 14 mm ×14 mm area were chosen and monitored by large detection area SPR microscopy; the new sensor chip exhibited stability comparable to the single gold layered SPR chip. In terms of sensing performances, over 50% increases in sensitivity and signal-to-noise ratio (S/N) than those of the single gold layered SPR chip were determined by SPR microscopy at 660 nm. Protein arrays of protein A and bovine serum albumin (BSA) on both the new chip and single-layered gold SPR chip were fabricated and underwent biomolecular interactions with human IgG, for the purpose of consistency, comparison on kinetics parameters, values from the microarray trilayered chip showed reasonable consistency with those from the single gold layered SPR chip. This study suggests that the new chip is a viable alternative to the conventional single gold layered SPR chip with improved sensing performances.
A novel surface plasmon resonance imaging (SPRi) biosensor based on waveguide coupled surface pla... more A novel surface plasmon resonance imaging (SPRi) biosensor based on waveguide coupled surface plasmon resonance (WCSPR) measurements is reported. Simply fabricated by gold evaporation deposition and polymer spin-coating, the WCSPR chips can be easily adapted on an commercial SPRi monitoring instrument. The chips were designed by using Fresnel method and fabricated according to its optimized design, the calculation and experimental results fits well. Over an 8 mm × 8 mm rectangle imaging area, the WCSPR chips realized about 52% increase of sensitivity, 57% enhancement of resolution and 55% improvement of signal to noise ratio compared to conventional SPR chips. Furthermore, high throughput protein-protein interaction analyses based on WCSPR chips were successfully accomplished by applying Protein A and rabbit antibody interaction in an array format. The reflectivity responses observed from the WCSPR chips were about 50% higher than the conventional SPR chip while the binding kinetic parameters were within the same range as with conventional SPR chip. (J. Zhu). 1 Equal contribution, Zhiyou Wang designed and fabricated WCSPR chips, Lusheng Song performed the parameters analysis of the WCSPR chips and the applications for high-throughput bio-interaction measurements. 0925-4005/$ -see front matter His current study is focused on optical sensing, material fabrication process, research and application of new SPR structure to imaging technique. . His current study is focused on microfluidic devices and biosensors.
Accurate measurement of inter-peptide interactions is beneficial for in-depth understanding disea... more Accurate measurement of inter-peptide interactions is beneficial for in-depth understanding diseaserelated protein folding and peptide aggregation, and further for designing and selecting potential peptide drugs to the target antigen. Herein, we demonstrate a 3D polyrotaxane (PRX) surface for detecting peptides interactions by surface plasmon resonance imaging (SPRi). This surface is supramolecular selfassembly monolayer (SAM) structure fabricated by threading α-cyclodextrans (α-CD) through a linear polyethylene glycol (PEG) chain fixed on gold chip surface to form pseudopolyrotaxane, and further capping the pseudopolyrotaxane with bulky terminated group to form PRX film. The hydroxyl groups of α-CD can provide more active sites to increase molecules immobilization density, and PEG chain has unique protein non-fouling feature. We chose Alzheimer's disease marker β-amyloid 40 (Aβ40) as model peptide, and detected the interaction between it and its inhibitors KLVFFK6 by SPRi. As a striking result, the specific adsorption of KLVFFK6 solution at the concentration of 352 μM on Aβ40-PRX was 700 RU, whereas PEG SAM surface gave no significant binding. Interaction between other lower molecular weight peptides was detected via PRX surface, and the relatively weak interactions (K D ¼1.73 Â 10 À 4 M) between LPFFD (Mw ¼ 0.6 kDa) and amylin20-29 (Mw¼ 1.0 kDa) are successfully detected.
Surface plasmon resonance imaging (SPRi) has emerged as a versatile biosensor to detect a wide ra... more Surface plasmon resonance imaging (SPRi) has emerged as a versatile biosensor to detect a wide range of biomolecular interactions with divergent potential applications. However, the use of this advanced-level technology for stem cell lysate study is still not much explored. Cell lysates are significant biological analytes used for disease diagnostics and proteomic studies, but their complex nature limits their use as an analyte for SPRi biosensors. Here, we review the problems associated with the use of SPRi for stem cell lysate study and examine the role of surface chemistry, running buffer, and blocking solution in order to minimize nonspecific adsorption (NSA). We detect the expression of Oct4, Sox2, Nanog, Rex1, and Lin28 biomarkers present in mouse embryonic stem cell (mESC) lysate against their corresponding antibodies immobilized on the sensor surface with reduced NSA. The current study shows that the conjunction of SPRi and microarray can be used as a label-free, high-throughput, and rapid technique for detection of biomarkers and their relative abundance in stem cell lysate study.
In recent years, in situ protein synthesis microarray technologies have enabled protein microarra... more In recent years, in situ protein synthesis microarray technologies have enabled protein microarrays to be created on demand just before they are needed. In this paper, we utilized the TUS-TER immobilization technology to allow label-free detection with real-time kinetics of protein–protein interactions using surface plasmon resonance imaging (SPRi). We constructed an expression-ready plasmid DNA with a C-terminal TUS fusion tag to directionally immobilize the in situ synthesized recombinant proteins onto the surface of the biosensor. The expression plasmid was immobilized on the polyethylene imine-modified gold surface, which was then coupled with a cell-free expression system on the flow cell of the SPRi instrument. The expressed TUS fusion proteins bind on the surface via the immobilized TER DNA sequence with high affinity (3–7 ×10_13 M). The expression and immobilization of the recombinant in situ expressed proteins were confirmed by probing with specific antibodies. The present study shows a new low cost method for in situ protein expression microarrays that has the potential to study the kinetics of protein–protein interactions. These protein microarrays can be created on demand without the problems of stability associated with protein arrays used in the drug discovery and biomarker discovery fields.
The silver surface plasmon resonance (SPR) sensor has long been explored due to its intrinsic sen... more The silver surface plasmon resonance (SPR) sensor has long been explored due to its intrinsic sensitivity enhancement over the conventional single-layered gold SPR sensor. However, the silver SPR sensor has not been exploited for practical applications because of pronounced instability problems. We propose a novel gold−silver−gold trilayered SPR sensor chip, in which an extra buffer layer of gold is added between the silver and substrate adhesion layer (i.e., chromium) compared to the previously reported silver−gold bilayered SPR sensors. Subjected to prolonged agitation in phosphate-buffered saline (PBS) solution, the new chip exhibited high integrity according to both optical and atomic force microscopy (AFM) analysis. Having undergone repeated cycles of calibration, binding, and regeneration in various chemical solutions, 25 regions of interest (ROIs) over a 14 mm ×14 mm area were chosen and monitored by large detection area SPR microscopy; the new sensor chip exhibited stability comparable to the single gold layered SPR chip. In terms of sensing performances, over 50% increases in sensitivity and signal-to-noise ratio (S/N) than those of the single gold layered SPR chip were determined by SPR microscopy at 660 nm. Protein arrays of protein A and bovine serum albumin (BSA) on both the new chip and single-layered gold SPR chip were fabricated and underwent biomolecular interactions with human IgG, for the purpose of consistency, comparison on kinetics parameters, values from the microarray trilayered chip showed reasonable consistency with those from the single gold layered SPR chip. This study suggests that the new chip is a viable alternative to the conventional single gold layered SPR chip with improved sensing performances.
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Papers by Zhiqiang Cheng
biomolecular interactions with divergent potential applications. However, the use of this advanced-level
technology for stem cell lysate study is still not much explored. Cell lysates are significant biological analytes
used for disease diagnostics and proteomic studies, but their complex nature limits their use as an
analyte for SPRi biosensors. Here, we review the problems associated with the use of SPRi for stem cell
lysate study and examine the role of surface chemistry, running buffer, and blocking solution in order
to minimize nonspecific adsorption (NSA). We detect the expression of Oct4, Sox2, Nanog, Rex1, and
Lin28 biomarkers present in mouse embryonic stem cell (mESC) lysate against their corresponding antibodies
immobilized on the sensor surface with reduced NSA. The current study shows that the conjunction
of SPRi and microarray can be used as a label-free, high-throughput, and rapid technique for
detection of biomarkers and their relative abundance in stem cell lysate study.
biomolecular interactions with divergent potential applications. However, the use of this advanced-level
technology for stem cell lysate study is still not much explored. Cell lysates are significant biological analytes
used for disease diagnostics and proteomic studies, but their complex nature limits their use as an
analyte for SPRi biosensors. Here, we review the problems associated with the use of SPRi for stem cell
lysate study and examine the role of surface chemistry, running buffer, and blocking solution in order
to minimize nonspecific adsorption (NSA). We detect the expression of Oct4, Sox2, Nanog, Rex1, and
Lin28 biomarkers present in mouse embryonic stem cell (mESC) lysate against their corresponding antibodies
immobilized on the sensor surface with reduced NSA. The current study shows that the conjunction
of SPRi and microarray can be used as a label-free, high-throughput, and rapid technique for
detection of biomarkers and their relative abundance in stem cell lysate study.