Functionalization of silicon dioxide and silicon nitride surfaces with aminosilanes for optical biosensing applications (original) (raw)
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Light assisted antibody immobilization for bio-sensing
Biomedical Optics Express, 2011
Ultrashort UV pulses at 258 nm with repetition rate of 10 kHz have been used to irradiate buffer solution of antibody. The tryptophan residues strongly absorb this radiation thus becoming capable to disrupt the disulfide bridges located next to them. Due to their high reactivity the opened bridges can anchor a gold plate more efficiently than other sites of the macromolecule giving rise to preferential orientations of the variable part of the antibody. UV irradiation has been applied to anchor antiIgG antibody to the electrode of a Quartz Crystal Microbalance (QCM) that lends itself as a sensor, the antibody acting as the bio-receptor. An increase of the QCM sensitivity and of the linear range has been measured when the antibody is irradiated with UV laser pulses. The photo-induced reactions leading to disulfide bridge breakage have been analyzed by means of a chemical assay that confirms our explanation. The control of disulfide bridges by UV light paves the way to important applications for sensing purpose since cysteine in combination with tryptophan can act as a hook to link refractory bio-receptors to surfaces.
Label-free monitoring of antibody–antigen interactions using optical microchip biosensors
Journal of Immunological Methods, 2010
A rapid, label-free optical biosensor system for sensitive monitoring of bio-molecular interactions in real-time is presented. SpectroSens ™ sensor chips are based on integrated planar Bragg gratings sensitive to localised changes in refractive index. Bio-molecule recognition is imparted by functionalisation of the sensing surface with antibodies against targets of interest. In this study, antibodies against selected proteins were oriented with recombinant Protein A/G, which was covalently immobilised to the sensor chip via amine coupling to a glutaraldehyde-activated silane layer. Immunoassays for the detection of rabbit IgG and ovalbumin proteins as model antibody-antigen interaction systems were performed. Binding of complementary antigens to respective antibody-functionalised sensors manifested as changes in wavelength of light reflected from the optical sensors. Quantitative binding kinetics with detection sensitivities in the mid ng/ml range were obtained for both antigens using this planar, two-dimensional surface coating. Data presented demonstrate the suitability of SpectroSens ™ sensors as a valuable tool in life science research and development for monitoring bio-specific interactions, protein concentration determination and antibody selection; the optical integration and analytical characteristics of these sensors suggest that they may find numerous applications in bio-pharmaceutical development and clinical diagnostics.
Biosensors and Bioelectronics, 1998
In the present paper surface studies for the development of a direct optical immunosensor for fast diagnosis of a myocardial infarction are presented. A fatty acid binding protein was detected by monoclonal antibodies. The applied measuring system was the grating coupler BIOS-1. Based on commercially available transducer materials protein immobilisation techniques have been developed and characterised by TOF-SIMS, AFM and EM. Three different label-free assay types were investigated. Only one assay leads to a sensitive and regenerable sensor set-up. It was possible to detect concentrations of the fatty acid binding protein down to 330 ng/ml. The general applicability of a direct optical immunosensor in the field of myocardial infarction diagnosis was demonstrated by this.
Applied Surface Science, 2014
In the present work, we developed and optimized a technique to produce a thin, stable silane layer on silicon substrate in a controlled environment using (3-aminopropyl)triethoxysilane (APTES). The effect of APTES concentration and silanization time on the formation of silane layer is studied using spectroscopic ellipsometry and Fourier transform infrared spectroscopy (FTIR). Biomolecules of interest are immobilized on optimized silane layer formed silicon substrates using glutaraldehyde linker. Surface analytical techniques such as ellipsometry, FTIR, contact angle measurement system, and atomic force microscopy are employed to characterize the biochemically modified silicon surfaces at each step of the biomolecule immobilization process. It is observed that a uniform, homogenous and highly dense layer of biomolecules are immobilized with optimized silane layer on the silicon substrate. The developed immobilization method is successfully implemented on different silicon substrates (flat and pillar). Also, different types of biomolecules such as anti-human IgG (rabbit monoclonal to human IgG), Listeria monocytogenes, myoglobin and dengue capture antibodies were successfully immobilized. Further, standard sandwich immunoassay (antibody-antigen-antibody) is employed on respective capture antibody coated silicon substrates. Fluorescence microscopy is used to detect the respective FITC tagged detection antibodies bound to the surface after immunoassay.
Development of optical immunosensors for detection of proteins in serum
Talanta, 2013
The detection of proteins in biological samples such as blood, serum or plasma by biosensors is very challenging due to the complex nature of the matrix, which contains a high level of many interfering compounds. Here we show the application of a novel polymeric immobilisation matrix that helps in the detection of specific protein analytes in biological samples by surface plasmon resonance (SPR) immunosensors. This polymer matrix contains thioacetal functional groups included in the network, and these groups do not require any further activation in order to react with proteins, making it attractive for sensor fabrication. The protein prostate specific antigen (PSA) was selected as a model target analyte. A sandwich format with two primary antibodies recognising different parts (epitopes) of the analyte was used for the detection of PSA in serum. The efficiency of the reduction of non-specific binding achieved with novel polymer was compared with those of other techniques such as coating of sensor surface with polyethylene glycol (PEG), use of charged hydrophilic aspartic acid and surfactants such as Tween20. The detection limit of the polymer based immunosensor was 0.1 ng ml À 1 for free form PSA (f-PSA) in buffer and 5 ng ml À 1 in 20% serum. This is an improvement compared with similar devices reported on literature, indicating the potential of the immunosensor developed here for the analysis of real samples.
Analytical and Bioanalytical Chemistry, 2015
A 3-channel biosensor based on spectral correlation interferometry (SCI) has been adapted for direct optical detection of antigens by measuring changes in thickness of a biolayer on functionalized glass slips employed as affordable single-use sensor chips. The instrument is insensitive to the bulk refractive index of a solution under test and provides signals in metrological units (pm or nm). Using real-time monitoring with the SCI, protocols for fabrication of sensor chips with different functional (epoxylated, carboxylated, and biotinylated) surfaces for antibody immobilization have been developed and optimized to minimize chip-to-chip variations and achieve better limit of detection (LOD), shorter assay time, and longer shelf life. The optimized coupling surfaces have been compared for detection of human serum albumin (HSA) used as a model agent of medical significance. The dynamic ranges for measuring the HSA concentration were 0.07-20, 0.12-30, and 0.25-10 μg/ml, and the assay durations were less than 20, 15, and 30 min for the epoxylated, carboxylated, and biotinylated chips, respectively. The advantages of each type of sensor chip have been shown, namely, the carboxylated chips feature the shortest assay time, the epoxylated ones demonstrate the best LOD, and the biotinylated chips exhibit the longest shelf life in an unprotected environment. The developed protocols of antibody immobilization can be used in different biosensors and assay techniques including those based on fluorescent, magnetic or plasmonic labels, etc. The SCI is well compatible with various partially transparent layers used in biosensing and with microarrays for multi-analyte detection.
Silicon-based biosensors for rapid detection of protein or nucleic acid targets
Clinical chemistry, 2001
We developed a silicon-based biosensor that generates visual, qualitative results or quantitative results for the detection of protein or nucleic acid targets in a multiplex format. Capture probes were immobilized either passively or covalently on the optically coated surface of the biosensor. Intermolecular interactions of the immobilized capture probe with specific target molecules were transduced into a molecular thin film. Thin films were generated by enzyme-catalyzed deposition in the vicinity of the surface-bound target. The increased thickness on the surface changed the apparent color of the biosensor by altering the interference pattern of reflected light. Cytokine detection was achieved in a 40-min multiplex assay. Detection limits were 4 ng/L for interleukin (IL)-6, 31 ng/L for IL1-beta, and 437 ng/L for interferon-gamma. In multianalyte experiments, cytokines were specifically detected with signal-to-noise ratios ranging from 15 to 80. With a modified optical surface, spe...
Sensors
Optical immunosensors represent a research field of continuously increasing interest due to their unique features, which can mainly be attributed to the high-affinity and specific antibodies they use as biorecognition elements, combined with the advantageous characteristics of the optical transducing systems these sensors employ. The present work describes new developments in the field, focusing on recent bioanalytical applications (2021–2022) of labeled and label-free optical immunosensors. Special attention is paid to a specific immunosensing platform based on White Light Reflectance Spectroscopy, in which our labs have gained specific expertise; this platform is presented in detail so as to include developments, improvements, and bioanalytical applications since the mid-2000s. Perspectives on the field are been briefly discussed as well, highlighting the potential of optical immunosensors to eventually reach the state of a reliable, highly versatile, and widely applicable analyti...
Characterization of anhydrous silanization and antibody immobilization on silicon dioxide surface
Formation of uniform and cluster free silane The thicker silane layer on the sensor surface alters the monolayer is one of the fundamental prerequisite for affinity mechanical properties of the biosensor. It also induces cantflever based biosensors. We report anhydrous silanization stresses on the sensor surface which may leads to difficulties protocol for uniform silane monolayer on silicon dioxide (SiO2) s n the s ensor suracewicayi ed o difficulTi surface using [3-(2-aminoethyl) aminopropyll-trimethoxysiane (APTES) and characterized by AFM, spectroscopic anticipate these problems silane monolayer with controlled ellipsometry and FTIR. Silanization coverage is controled by thickness on biosensor is essential. availability of surface water. The roughness of the resulting In the conventional protocol of aqueous phase silanized surface following our protocol is in the range of SiO2 silanization, the aminosilanes undergo hydrolysis and surface roughness. Silanized SiO2surface is used to immobilize polymerization in the bulk phase before depositing and human immunoglobulin (HlgG) on it. FITC tagged goat forming bonds with the silicon dioxide surface [1]. However, antihuman IgG is allowed to react with HIgG. The immobilized this results in the formation of polysilane networks prior to surface is further characterized using Fluorescent spectroscopy deposition. Thus polymerization of multifunctional silane and Fluorescent Microscope. Characterization results obtained molecules is observed both parallel and perpendicular to the from anhydrous silanization protocol are compared with the solfces leadtoforatoofiane me cule te conventional aqueous silanization protocol.