Yong-Hong Meng

Detection of hepatitis B virus markers using a biosensor based on imaging ellipsometry

Summary.  A biosensor based on imaging ellipsometry (BIE) has been developed and validated in 169 patients for detecting five markers of hepatitis B virus (HBV) infection. The methodology has been established to pave the way for clinical diagnosis, including ligand screening, determination of the sensitivity, set‐up of cut‐off values (CoVs) and comparison with other clinical methods. A matrix assay method was established for ligand screening. The CoVs of HBV markers were derived with the help of receiver operating characteristic curves. Enzyme‐linked immunosorbent assay (ELISA) was the reference method. Ligands with high bioactivity were selected and sensitivities of 1 ng/mL and 1 IU/mL for hepatitis B surface antigen (HBsAg) and surface antibody (anti‐HBs) were obtained respectively. The CoVs of HBsAg, anti‐HBs, hepatitis B e antigen, hepatitis B e antibody and core antibody were as follows: 15%, 18%, 15%, 20% and 15%, respectively, which were the percentages over the values of corresponding ligand controls. BIE can simultaneously detect up to five markers within 1 h with results in acceptable agreement with ELISA, and thus shows a potential for diagnosing hepatitis B with high throughput.

Phage M13Ko7 Detection With Biosensor Based On Imaging Ellipsometry And Afm Microscopic Confirmation

Abstract A rapid detection and identification of pathogens is important for minimizing transfer and spread of disease. A label-free and multiplex biosensor based on imaging ellipsometry (BIE) had been developed for the detection of phage M13KO7. The surface of silicon wafer is modified with aldehyde, and proteins can be patterned homogeneously and simultaneously on the surface of silicon wafer in an array format by a microfluidic system. Avidin is immobilized on the surface for biotin-anti-M13 immobilization by means of interaction between avidin and biotin, which will serve as ligand against phage M13KO7. Phages M13KO7 are specifically captured by the ligand when phage M13KO7 solution passes over the surface, resulting in a significant increase of mass surface concentration of the anti-M13 binding phage M13KO7 layer, which could be detected by imaging ellipsometry with a sensitivity of 109 pfu/ml. Moreover, atomic force microscopy is also used to confirm the fact that phage M13KO7 has been directly captured by ligands on the surface. It indicates that BIE is competent for direct detection of phage M13KO7 and has potential in the field of virus detection.

Optimization of off-null ellipsometry for air/solid interfaces.

The optimization of off-null ellipsometry is described with emphasis on the improvement of sample thickness sensitivity. Optimal conditions are dependent on azimuth angle settings of the polarizer, compensator, and analyzer in a polarizer-compensator-sample-analyzer ellipsometer arrangement. Numerical simulation utilized offers an approach to present the dependence of the sensitivity on the azimuth angle settings, from which optimal settings corresponding to the best sensitivity are derived. For a series of samples of SiO2 layer (thickness in the range of 1.8-6.5 nm) on silicon substrate, the theory analysis proves that sensitivity at the optimal settings is increased 20 times compared to that at null settings used in most works, and the relationship between intensity and thickness is simplified as a linear type instead of the original nonlinear type, with the relative error reduced to approximately 1/100 at the optimal settings. Furthermore the discussion has been extended toward other factors affecting the sensitivity of the practical system, such as the linear dynamic range of the detector, the signal-to-noise ratio and the intensity from the light source, etc. Experimental results from the investigation of SiO2 layer on silicon substrate are chosen to verify the optimization.

Biosensor with total internal reflection imaging ellipsometry

In order to monitor multiple protein reaction processes simultaneously, a biosensor based on imaging ellipsometry operated in the total internal reflection mode is proposed. It could be realised as an automatic analysis for protein interaction processes with real-time label-free method. Its principle and methodology as well as a demonstration for its applications are presented.

The development of biosensor with imaging ellipsometry

The concept of biosensor with imaging ellipsometry was proposed about ten years ago. It has become an automatic analysis technique for protein detection with merits of label-free, multi-protein analysis, and real-time analysis for protein interaction process, etc. Its principle, and related technique units, such as micro-array, micro-fluidic and bio-molecule interaction cell, sampling unit and calibration for quantitative detection as well as its applications in biomedicine field are presented here.

Imaging ellipsometry for the visualization of bio-molecular layers

Imaging ellipsometry is presented as a visualization technique to study bio-molecular layers. The layers have become more and more attractive in materials science, especially layers with thickness similar to cellular layers and with physiological activities which are very important in molecular biology and medicine. Normally bio-molecular layers are very thin, with thickness of between sub-nanometer and several tens nanometer. They are transparent in the visible range of light, so that they are recognized as a phase object in physics. Imaging ellipsometry is non-destructive and exhibits a high sensitivity to phase transitions within thin layers. It is capable of imaging local variations in the optical properties such as thickness due to the presence of different surface concentrations of bio-molecule or different deposited molecules. Imaging ellipsometry is based on conventional ellipsometry with charge coupled device (CCD) technique. The images are captured with a computer with image processing technique. It has high sensitivity to thickness variation (resolution in the order of angstrom), and high sampling speed (25 pictures with more than 10/sup 5/ pixels per second).

Application of Optical Protein-chip in Detecting Phage M13KO7

Abstract Avidin layer was bound on the substrate surface of Silicon wafer modified with aldehyde. The interaction between avidin and biotin was adopted for the immobilization of mouse monoclonal biotin-anti-M13 (antibody GP3)-labeled biotin. The surface was incubated in a solution containing phage M13KO7, which was trapped by the antibody GP3 with the interaction between phage M13KO7 and antibody GP3, resulting in a variation of layer thickness that was detected by imaging ellipsometry. The results showed a saturated layer of antibody GP3 with a thickness about 6.9 nm on the surface of the silicon wafer. The specific interaction between phage M13KO7 and antibody GP3 resulted in a variation of layer thickness. The layer of phage M13KO7 bound with antibody GP3 was 17.5 nm in the concentration of 1.1×1010 pfu/mL. Each variation of the layer thickness corresponded to a concentration of phage M13KO7 in the range of 0.1×1010–2.5×1010 pfu/mL, with the sensitivity of 109 pfu/mL. Compared with other methods, the optical protein-chip, requiring only short measurement time, label free, is a quantitative test, and can be visualized. This study could be significant on the interactions between the antibody and the virus, showing potential in the early diagnosis of virosis.

Optical protein chip as microarrays for protein interaction determination

One kind of optical protein chip and a ProteinChip analysis system have been introduced. The basic principle is that one kind of optical imaging technique - imaging ellipsometry is used to visualize optical biochips with the lateral thickness (or surface concentration) distribution of protein layers attached on a patterned surface. The chip is processed with surface patterning, surface modification and ligand immobilization to form a sensing surface with multi-bioactivity. The affinity between proteins is used to realize non-labeling micro-assays for protein identifications and protein interactions. A model of the chip and some demonstration results are presented.

CD146 detection with real-time total internal reflection imaging ellipsometry

Biosensor with the total internal reflection imaging ellipsometry (TIRIE) uses an evanescent wave as optical probe to monitor bio-molecular interaction with a high sensitivity due to its property of phase sensitive. Here, the biosensor is applied for a quantitative detection of CD146 with concentrations of 0.1 to 100 ng/mL in order to realize a high sensitive quantitative detection. Moreover, the regression curve between the signal of biosensor (y) and CD146 concentration (x.. lnC+2.4) is deduced as a linear y=1.0544x+0.7839.

Soluble Angiopoietin Receptor Tie-2 in Patients with Acute Myocardial Infarction and its Detection by Optical Protein-chip

The Tie-2 receptor has been shown to play a role in angiogenesis in atherosclerosis. The conventional method assaying the level of soluble Tie-2 (sTie-2) was ELISA. However, this method has some disadvantages. The aims of this research are to establish a more simple detection method, the optical protein-chip based on imaging ellipsomtry (OPC-IE) applying to Tie-2 assay. The sTie-2 biosensor surface on silicon wafer was prepared first, and then serum levels of sTie-2 in 38 patients with AMI were measured on admission (day 1), day 2, day 3 and day 7 after onset of chest pain and 41 healthy controls by ELISA and OPC-IE in parallel. Median level of sTie-2 increased significantly in the AMI patients when compared with the controls. Statistics showed there was a significant correlation in sTie-2 results between the two methods (r=0.923, P<0.01). The result of this study showed that the level of sTie-2 increased in AMI, and OPC-IE assay was a fast, reliable, and convenient technique to measure sTie-2 in serum.