Constructions Of Biosensors Based On Optics And Their Applications In Biomarkers Analysis

A biosensor is defined by the International Union of Pure and Applied Chemistry as a device that uses specific biochemical reactions mediated by isolated enzymes,immunosystems,tissues,organelles or whole cells to detect chemical compounds usually by electrical,thermal or optical signals.Therefore,a biosensor is an integrated biorecognition-transducer device,which can convert a biological response into a detectable signal.Over the past decades,the design and development of biosensors have taken a center stage for researchers or scientists owing to the wide range of biosensor applications,such as health care and disease diagnosis,environmental monitoring,water and food quality monitoring,and drug delivery.Since the invention of the Clark oxygen electrode sensor,there have been many improvements in sensitivity,selectivity,and multiplexing capacity of modern biosensors.Focusing on the construction and application of biosensors,the following several aspects of research work were carried out.（1）The semi-cylindrical prism to expand a point laser source to a long beam for illuminating multiple wells was used to enhance the sample throughput of microplate-based photothermal biosensor detection.Coupled with four epoxy-coated thermocouples in alignment with wells on a 96-well microplate,four parallel immunoassays of C-reaction protein（CRP）with antibody-conjugated gold nanoparticles can be simultaneously performed.The sample throughput further increased by mounting the styrofoam-enclosed microplate onto a translational/elevator stage so that immunoassays and thermocouple rinse/drying cycles can be implemented in a pro-grammed fashion.With careful calibration of the energy distribution of the expanded laser beam and controllable immersion of the thermocouples,excellent reproducibility can be attained.The temperature changes can be correlated with the CRP concentration by the Langmuir isotherm,and the low limit of detection,0.52 ng/m L,is well below the plasma CRP levels of both healthy people（<5μg/m L）and patients（10-500μg/m L）.The serum CRP concentrations quantified by the developed plate reader are in excellent agreement with the immunoturbidimetric results,demonstrating that this cost-effective,robust,and high-throughput mode for microplate-based photothermal biosensor is amenable to detecting biomarkers in many clinical samples.（2）MnO₂ nanosheets were synthesized in a one-pot synthesis via reducing KMnO₄ with sodium alginate（SA）,environmentally friendly materials,a natural product isolated from marine algae.A radiometric fluorescence immunoassay biosensor was designed via the fluorescence resonance energy transfer（FRET）in a system comprising MnO₂ nanosheets,o-phenylenediamine（OPD）,and fluorescent carbon nanodots（FCNs）.The MnO₂ nanosheets oxidizes OPD,producing a fluorescence peak at 580 nm,while FCNs exhibit a fluorescence peak at 490 nm.Under the effect of FRET,the fluorescence at 580 nm increases significantly while the fluorescence at 490 nm reduces.The FRET effect was confirmed by fluorescence lifetime measurements.By electrostatically coating the MnO₂ nanosheets with antibody molecules and using the resultant materials as a detection probe,the ratiometric fluorescence sensing was extended to the detection of alpha fetoprotein（AFP）in human serum samples on a 96-well titer plate.The ratiometric fluorescence immunoassay biosensor possesses a good dynamic range（from 0.01 to 9.00 ng/m L）.The detection limit was estimated to be0.009 ng/m L,which is 7.6-fold lower than that of the single-wavelength-based immunoassay biosensor.The measurement results of serum samples from healthy and patient donors are highly comparable to those measured by chemiluminescence for the same donors,demonstrating the viability for serological assays of important cancer markers.（3）Surface plasmon resonance microscopy（SPRM）biosensor is a powerful technique for measuring the binding kinetics of biomolecular interactions in a real-time and label-free manner.But the non-specific adsorption of proteins on gold sensor chip surfaces that leads to adverse problems and is prevented by a biocompatible surface coating.Poly（ethylene glycol）（PEG）is one of the current common material for biocompatible surface coating.Hence,a cell-based PEG chip prepared by zoning method was designed to minimize nonspecific interactions and maximize the response from the analyte of interest.A combination of the cell-based PEG chip and surface plasmon resonance microscope（SPRM）technique was applied to study the interaction between wheat germ lectin and cell membrane glycoproteins,as well as the interaction between Spike S1 protein of SARS-Co V-2 and angiotensin converting enzyme 2（ACE2）receptor.More accurate kinetic information were obtained.Importantly,this work is the first to use a cell-based PEG chips for SPRM technology as background deductions.