Over the last two decades, Surface Plasmon Resonance (SPR) has evolved into a crucial label-free detection approach, revolutionising biomolecular interaction studies in clinical settings. This technology enables high-sensitivity real-time measurement of interactions without the use of labels. Label-free detection eliminates the requirement for specialised tags or dyes, allowing for more sensitive measurement of target analytes and the use of native biomolecules appropriate for biologically relevant methods.
Since its introduction in the early 1990s, SPR has proven to be one of the most powerful technologies for determining specificity, affinity, and kinetic parameters during macromolecule binding in a variety of bonds, including protein-protein, protein-DNA, enzyme-substrate or inhibitor, receptor-drug, lipid membrane-protein, protein-polysaccharide, cell or virus-protein, and others.
In this blog, we will delve into the principles of SPR, explore its applications, and highlight recent advances that underscore its importance in bioassays.
Principles of Surface Plasmon Resonance (SPR)
SPR operates by generating electron charge density waves at the interface between two media when light passes through a prism and strikes a metal surface, typically gold or silver. The resulting resonant angle, known as the SPR angle, leads to a reduction in reflected light intensity, creating a distinctive sensorgram.
When proteins are immobilized on the metal surface, potential ligands injected over the surface cause changes in the resonant angle, allowing real-time recording of biomolecular interactions. The association constant (Ka) and dissociation constant (Kd) are key parameters captured in the SPR sensorgram.
Biacore™ SPR Systems
Biacore™ SPR systems, which are widely used in pharmaceutical development and life science research, enable real-time label-free tracking of molecular interactions. Target molecules, commonly proteins, are immobilised on a sensor surface in these tests, and a sample containing a possible interaction partner is pumped across the surface using flow cells. The resultant sensorgram offers information on binding kinetics, specificity, concentration, and affinity.
(Fig – Scientist working on Surface Plasmon Resonance in Biology Lab at o2h discovery)
Advantages of SPR in Drug Discovery
- Real-Time Insights: SPR enables us to observe binding events between small molecules (fragments) and target proteins in real time. This means we can track the interaction as it happens, providing critical insights into kinetics, affinity, and specificity.
- Label-Free Analysis: Unlike many other screening techniques, SPR does not require labelling of the molecules involved. This not only simplifies the experimental setup but also ensures that the interactions are studied in their native state, without any alterations.
- High Sensitivity: SPR is incredibly sensitive, allowing us to detect even weak interactions. This is particularly valuable in FBDD, where we’re working with fragments that may have lower binding affinities compared to larger compounds.
- Quantitative Data: SPR provides quantitative data on binding kinetics, association and dissociation rates, equilibrium constants, and more. This wealth of information is crucial in making informed decisions about lead optimization.
- Fragment Screening Efficiency: SPR excels in fragment screening due to its ability to work with low molecular weight compounds. It allows us to rapidly screen a diverse set of fragments against a target, narrowing down the list of potential leads efficiently.
- Versatility in Target Classes: SPR is applicable to a wide range of target classes, including proteins, nucleic acids, and small molecules. This versatility makes it an indispensable tool in our pursuit of diverse therapeutic solutions.
Applications of SPR-Based Biosensors
- Biomedical Applications: SPR biosensing is one of the most effective approaches for monitoring biomolecule affinity binding and screening for druggable compounds. They are used in various research to analyse a wide range of biological substances, including DNAs, RNAs, proteins, carbohydrates, lipids, and cells.
- High-Throughput Screening (HTS): SPR are extensively used for drug discovery and development, especially in high-throughput screening, due to its label-free nature.
- Proteomics Research: SPR has proved effective in biomarker diagnostics because of its sensitivity, mobility, elimination of huge sample quantities, and multiplexed detection capacity. SPR biosensors have been used effectively to identify biomarkers for a variety of illnesses, including breast, ovarian, and pancreatic cancer, as well as cardiac and neurological ailments.
- Cellular Analysis and Cell-Based Detection: The SPR approach has advantages not only because of its real-time and label-free imaging capabilities for dynamic changes at the surface, but also for cellular changes such as physiology, cell-surface interactions, and cell identification. The SPR monitors cellular response, adhesion, and products, playing a significant role in cancer cell detection.
Recent Advances in Surface Plasmon Resonance Technology
Researchers have successfully used this high-spatial resolution SPR in imaging and detection of single DNA molecules, viruses, and cells. This unique approach also enabled the mapping of single cell-substrate interactions and direct binding kinetics of proteins to cell membrane proteins. Recent advances in plasmonic imaging techniques using high-resolution surface plasmon resonance microscopy (SPRM) would undoubtedly have a significant impact on the quantitative analysis of intracellular dynamics in live cells, single molecule analysis, studies of the biological activities of membrane proteins, and the discovery of new drugs that target membrane proteins.
Surface Plasmon Resonance has become an essential element of biosensing instruments, steering advancements in determining association and dissociation kinetics, nucleic acid hybridization, and protein-ligand and protein-protein interactions.
At o2h discovery, we have harnessed cutting-edge SPR technology to advance Fragment-Based Drug Discovery (FBDD) screening. With expertise in medicinal chemistry, biology, and biophysics, our team ensures the identification and optimization of the most promising fragments for further development.
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