Glossary

6.3 Surface plasmon resonance (SPR) biosensors

5 min read•august 14, 2024

Surface plasmon resonance biosensors are game-changers in detecting tiny molecular interactions. They use light and metal surfaces to spot when molecules stick together, without needing labels. This tech is super sensitive and can track binding in real-time.

SPR biosensors are used in , food safety, and medical testing. They're great for studying how proteins, antibodies, and DNA interact. The best part? They give us detailed info on how fast and how strongly molecules bind to each other.

Surface Plasmon Resonance: Principles and Applications

Physical Principles of SPR

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  • Surface plasmon resonance (SPR) is an optical phenomenon that occurs when polarized light strikes a metal film under total internal reflection conditions, causing a reduction in the intensity of reflected light at a specific angle known as the resonance angle
  • The excitation of surface plasmons depends on factors such as the wavelength of the incident light, the type and thickness of the metal film, and the dielectric properties of the surrounding medium
  • The generated during SPR penetrates a short distance into the medium adjacent to the metal surface, allowing the detection of biomolecular interactions within a few hundred nanometers from the surface
  • The resonance angle is sensitive to changes in the refractive index of the medium near the metal surface, making SPR suitable for detecting biomolecular interactions without the need for labeling

Applications of SPR in Biosensing

  • SPR biosensors exploit the relationship between the resonance angle and the refractive index changes caused by biomolecular interactions to quantify the kinetics and affinity of these interactions in real-time
  • SPR biosensors are used to study protein-protein interactions, antibody-antigen binding, enzyme-substrate interactions, and DNA hybridization
  • SPR biosensors find applications in drug discovery, food safety analysis, environmental monitoring, and clinical diagnostics
  • SPR imaging (SPRi) allows the simultaneous detection of multiple biomolecular interactions on a single sensor surface, enabling high-throughput screening and multiplexing

SPR Biosensor Components and Instrumentation

Key Components of an SPR Biosensor

  • A typical SPR biosensor consists of a light source, a prism, a thin metal film (usually gold), a flow cell, and a detector
  • The light source is typically a polarized laser or LED that emits monochromatic light at a specific wavelength, which is coupled to the prism to achieve total internal reflection at the prism-metal interface
  • The thin metal film, usually around 50 nm thick, is deposited on the prism surface and serves as the sensor surface where biomolecular interactions occur
  • The flow cell is positioned above the metal film and allows the controlled delivery of sample solutions containing the analyte of interest over the sensor surface

Instrumentation and Data Acquisition

  • The detector, often a CCD camera or photodiode array, measures the intensity of the reflected light at various angles, enabling the determination of the resonance angle and the monitoring of biomolecular interactions
  • Additional components may include a temperature control system to maintain a stable environment and a data acquisition and processing unit to analyze the SPR response in real-time
  • Advances in instrumentation, such as the development of portable and miniaturized SPR devices, have expanded the range of applications and improved the ease of use of SPR biosensors
  • Integration of SPR biosensors with microfluidic systems allows automated sample handling, reduced sample volumes, and improved assay throughput

Biomolecular Interaction Analysis with SPR

Ligand Immobilization Strategies

  • SPR biosensing involves immobilizing one of the interacting partners (ligand) on the sensor surface and introducing the other partner (analyte) in solution over the surface
  • Ligand immobilization can be achieved through various methods, such as covalent coupling, affinity capture, or hydrophobic adsorption, depending on the nature of the ligand and the surface chemistry of the sensor
  • Covalent coupling involves the formation of stable chemical bonds between the ligand and the sensor surface, often using amine, thiol, or carboxyl groups present on the ligand
  • Affinity capture relies on the specific interaction between the ligand and a capturing molecule immobilized on the sensor surface (streptavidin-biotin, antibody-antigen, or His-tag-metal ion interactions)

Kinetic and Affinity Analysis

  • As the analyte solution flows over the ligand-coated surface, the analyte binds to the ligand, causing an increase in the refractive index near the surface and a shift in the SPR resonance angle
  • The shift in the resonance angle is proportional to the change in mass on the sensor surface, allowing the quantification of the analyte-ligand interaction in real-time
  • By monitoring the SPR response during the association and dissociation phases of the interaction, kinetic parameters such as association and dissociation rate constants (ka and kd) can be determined, providing insights into the affinity and specificity of the interaction
  • Regeneration of the sensor surface by disrupting the analyte-ligand complex allows multiple interaction analyses to be performed on the same surface, enhancing the throughput and cost-effectiveness of the technique

Sensitivity, Specificity, and Real-Time Monitoring of SPR Biosensors

Sensitivity and Detection Limits

  • SPR biosensors offer high sensitivity, with detection limits in the nanomolar to picomolar range, making them suitable for detecting low-abundance analytes in complex biological samples
  • The sensitivity of SPR biosensors depends on factors such as the affinity of the analyte-ligand interaction, the surface coverage of the ligand, and the noise level of the instrument
  • Strategies to enhance sensitivity include the use of nanostructured sensor surfaces (nanoparticles or nanogratings), signal amplification methods (enzymatic or nanoparticle-based), and the optimization of surface chemistry and assay conditions

Specificity and Selectivity

  • SPR biosensors provide high specificity, as the detection is based on the specific interaction between the analyte and the immobilized ligand, minimizing the interference from non-specific binding events
  • The specificity can be further enhanced by optimizing the surface chemistry and the buffer conditions to reduce non-specific adsorption and by including appropriate controls to account for background signals
  • Selectivity can be improved by using highly specific ligands (monoclonal antibodies or aptamers) or by incorporating additional recognition elements (secondary antibodies or sandwich assays) to increase the specificity of the detection

Real-Time Monitoring Capabilities

  • One of the key advantages of SPR biosensors is their ability to monitor biomolecular interactions in real-time, providing kinetic information that is not available from end-point assays
  • allows the determination of association and dissociation rates, equilibrium constants, and the detection of transient or weak interactions that may be missed by other techniques
  • The real-time nature of SPR biosensing also enables the study of complex multi-step interactions, such as conformational changes or sequential binding events, providing a more comprehensive understanding of the underlying biological processes
  • Real-time monitoring facilitates the optimization of assay conditions, the evaluation of the effect of inhibitors or competitors, and the screening of compound libraries for drug discovery applications

Key Terms to Review (18)

Attenuated Total Reflection: Attenuated Total Reflection (ATR) is an optical phenomenon that occurs when light traveling through a medium hits the boundary of another medium at an angle greater than the critical angle, causing some of the light to be reflected back while the rest penetrates into the second medium. This method is particularly important in biosensing applications, as it allows for the interrogation of thin films and molecular interactions at interfaces without extensive sample preparation.
Biomolecular interaction analysis: Biomolecular interaction analysis is the study of the interactions between biomolecules, such as proteins, nucleic acids, and small molecules, to understand their functions and roles in biological systems. This analysis is crucial for drug discovery, diagnostics, and understanding cellular mechanisms, as it provides insights into how molecules communicate and interact at a molecular level.
Drug discovery: Drug discovery is the complex process of identifying, developing, and bringing new pharmaceutical compounds to market. This process involves various stages, including target identification, compound screening, lead optimization, and clinical trials, all aimed at developing effective treatments for diseases. Innovative techniques in biophotonics and optical biosensors have become essential in enhancing the efficiency and accuracy of drug discovery.
Evanescent Wave: An evanescent wave is a near-field wave that occurs when light or other electromagnetic radiation travels through a medium and encounters a boundary with a different medium. Instead of propagating into the second medium, the wave decays exponentially with distance from the boundary, creating a field that can interact with nearby materials without significant loss of energy. This phenomenon is crucial in optical applications, particularly in biosensors where it can enhance sensitivity and enable the detection of biomolecules at very low concentrations.
Functionalization: Functionalization refers to the process of chemically modifying a surface or material to introduce specific functional groups that enable desired interactions with biological molecules. This technique enhances the sensitivity and specificity of sensors, particularly in biosensing applications, by allowing targeted binding events and improving overall performance.
Gold film: A gold film is a thin layer of gold that is used in various optical applications, including biosensors. In the context of biosensors, gold films are critical because they support the excitation of surface plasmon resonance (SPR), which allows for sensitive detection of biological interactions at the molecular level. The unique optical and electronic properties of gold make it an ideal material for enhancing the sensitivity and specificity of these biosensors.
Homola: Homola refers to a significant contribution in the field of surface plasmon resonance (SPR) biosensors, specifically regarding the development of optical sensing techniques that utilize surface plasmons to detect biomolecular interactions. This term is closely associated with advancements in sensor design and signal enhancement, emphasizing the importance of nanostructures and their role in improving the sensitivity and specificity of SPR biosensors.
Kinetic analysis: Kinetic analysis refers to the study of the rates of reactions or interactions over time, often used to assess binding kinetics in biosensors. This approach is essential for understanding how quickly and efficiently a molecule binds to its target and how long it remains bound, which are crucial factors in evaluating sensor performance and sensitivity.
Kreibig: Kreibig refers to a model that describes the spectral features of surface plasmon resonance (SPR) biosensors, particularly focusing on the absorption and scattering properties of metal nanoparticles. This model helps in understanding how these particles interact with light and how this interaction can be quantified for biosensing applications, leading to enhanced sensitivity and specificity in detecting biomolecular interactions.
Label-free biosensors: Label-free biosensors are analytical devices that detect biological interactions without the need for labeling molecules with tags or dyes. This approach allows for real-time monitoring of biomolecular interactions, making it a powerful tool in various fields like diagnostics and research. By eliminating the need for labels, these biosensors reduce potential interference and simplify the detection process.
Limit of Detection: The limit of detection (LOD) refers to the lowest concentration of an analyte that can be reliably detected but not necessarily quantified in a given sample. It is a critical parameter in analytical methods, as it determines the sensitivity and effectiveness of detection systems, especially in biophotonics and optical biosensing applications.
Nanoplasmonic sensors: Nanoplasmonic sensors are highly sensitive detection devices that exploit surface plasmon resonance (SPR) at the nanoscale to identify and quantify biological and chemical substances. These sensors utilize metallic nanoparticles to enhance electromagnetic fields at their surfaces, significantly improving sensitivity and enabling detection at low concentrations. They offer advantages in miniaturization and integration into various platforms for biosensing applications.
Real-time monitoring: Real-time monitoring refers to the continuous observation and analysis of data as it is generated, allowing for immediate feedback and decision-making. This capability is essential in various applications, including tracking biological processes, assessing disease states, and evaluating drug interactions, enabling timely interventions and enhancing overall understanding of dynamic systems.
Resonance wavelength: Resonance wavelength refers to the specific wavelength at which a system experiences maximum response or resonance, particularly in relation to surface plasmon resonance (SPR) phenomena. In SPR biosensors, this wavelength is crucial for detecting biomolecular interactions as it correlates with the surface plasmon wave excited at the interface between a metal and a dielectric material. Understanding this wavelength helps in optimizing sensor performance and enhancing sensitivity to analyte binding events.
Sensor chip: A sensor chip is a small, often silicon-based device that detects and measures physical or chemical properties in its environment, converting this data into signals for analysis. In the context of surface plasmon resonance (SPR) biosensors, sensor chips are crucial components that provide a surface for biomolecular interactions to occur, enabling the measurement of changes in refractive index associated with binding events.
Signal-to-Noise Ratio: Signal-to-noise ratio (SNR) is a measure that compares the level of a desired signal to the level of background noise, indicating how well a signal can be distinguished from noise. A higher SNR means clearer, more reliable signals which is critical for various imaging and biosensing techniques, as it directly influences the accuracy and sensitivity of the measurements being taken.
Surface Modification: Surface modification refers to the process of altering the surface properties of a material to enhance its performance or functionality. This technique is crucial in biosensors, particularly for optimizing interactions between the sensor's surface and biological molecules, improving sensitivity and specificity in detection applications.
Surface plasmon polaritons: Surface plasmon polaritons (SPPs) are electromagnetic waves that propagate along the interface between a dielectric and a conductor, resulting from the coupling of light with surface plasmons. These waves play a crucial role in the field of biosensing, particularly in surface plasmon resonance (SPR) biosensors, as they enhance light-matter interactions, enabling the detection of biomolecular events at very low concentrations.
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