Surface plasmon resonance enables the detailed study of biomolecular function in real time and without the need for fluorescent tags or radioactive markers, and often without the need for prior purification. Elab reports.
Biacore's surface plasmon resonance (SPR) technology is a sensor chip-based analytical system where interactions between two or more biomolecules are measured by imobilising one type of molecule on the surface of a sensor chip and passing a solution containing the other molecule over the surface under controlled conditions.
By monitoring the interaction between the immobilised compound, and one or more compounds in the solution, the technology provides a level of functional data that is difficult or even impossible to obtain by any other single technique.
This technology contrasts with many of the traditional aend-point' test methods which measure a single point in the progress of an interaction and require large numbers of measurements at different time intervals to approximate the types of kinetic measurements achieved through Biacore's approach.
Biacore's SPR biosensors have a wide application in various areas of basic and applied research such as biochemisty, protein engineering, proteomics, cancer research and the discovery, design and evaluation of new therapeutic agents.
Biacore International AB (Biacore) is the world leader in the development of surface plasmon resonance (SPR) technology. The company's SPR technology enables the measurement of interactions between two or more biomolecules by immobilising one type of molecule (the chosen ainteractant') on the surface of a sensor chip and passing a solution containing the other molecule(s) (the aanalyte') over the surface under controlled flow conditions. For example, a particular type of antibody may be chosen as the interactant and immobilised on the sensor chip surface to study its interaction with a target antigen.
Similarly, a particular cell surface receptor may be immobilised to study its interaction with certain ligands.
The sensor chip consists of a glass surface coated with a thin layer of gold that provides the physical conditions required for SPR the phenomena on which the Biacore detection principle is based.
SPR is an electron charge density wave phenomenon that arises at the surface of a metallic film when light is reflected at the film under specific conditions. An evanescent wave extends beyond the sensor surface and detects mass changes on the surface.
At no time in the measurement process does light enter the sample, thereby eliminating the quenching or absorbance problems that beset all spectrophotometric and fluorescent techniques.
To prevent non-specific absorption of biomolecules onto the gold layer of the sensor surface, the gold is covered by a layer of adextran' on which the chosen interactant is immobilised by the user.
The user selects the chosen interactant and the method of immobilisation, thereby determining the specificity of the interaction being studied.
The dextran layer is designed to provide a favourable environment for interactions between typical biomolecules and to minimise non-specific binding.
Once the chosen interactant has been immobilised, a microfluidic system injects the analyte solution over the sensor surface, allowing binding to occur and enabling the quantitation of association rates.
When the analyte solution completes its pass over the sensor surface, measurements can be made of the dissociation rates.
Flow conditions are controlled through a liquid handling system made up of precision pumps and an integrated fluidic cartridge (IFC) that is responsible for both forming the flow cells on the sensor chip surface and for directing the flow of liquid over the surface.
Molecular binding information
Application of Biacore's SPR technology provides detailed information about several parameters that characterise molecular binding events: u How specific is the binding between two molecules (specificity)? u How much of a given molecule is present and active (concentration)? u How fast does the binding proceed (kinetics)? u How strong is the binding (affinity)?
In this way, Biacore's SPR technology has made a unique contribution to research. It enables the continuous monitoring of biomolecular interactions, thereby providing real-time kinetic information on molecular binding.
This is in contrast with many of the traditional aend-point' test methods which measure a single point in the progress of an interaction and require large numbers of measurements at different time intervals to approximate the types of kinetic measurements achieved through the Biacore approach.
Biacore's SPR biosensors are a truly non-label chip-based technology. Around 2300 peer reviewed scientific papers have been published citing Biacore instrumentation in basic and applied research by leading university groups, national research centres and major pharmaceutical companies.
Key advantages
In summary, Biacore's technology provides a number of key advantages: u Real-time kinetic analysis of molecular binding interaction giving an unparalleled level of valuable information on events as they occur u Non-label technology ensuring that biomolecules are not subject to alteration prior to investigation. u aNon-contact' detection in which the SPR measurement light does not enter the sample thereby eliminating the quenching or absorbance problems that beset all spectrophotometric and fluorescent techniques. u Minimum sample size and preparation required, saving both research costs and time.
Application in cancer research
Cancer is one of the major causes of death in Western world, killing one in five of the population. One in three people are affected by cancer at some stage in their lives.
The major challenges in cancer research focus on identifying cancer-causing agents, their mode of action, the biochemical events that underlie cancer development and new medical interventions that can impede or halt the proliferation of abnormal cells.
A recurrent theme in cancer research is the investigation of biomolecular binding events. Many processes underlying cancer development involve abnormal biomolecular interactions, such as the binding of carcinogenic compounds and/or their metabolites to DNA, or the abnormal activation or suppression of cell signaling pathways by abnormal receptor ligand interactions.
A key approach in cancer research is to study molecular interactions in normal and cancerous cells so that key differences can be identified. These differences may offer new targets for drug intervention.
Biacore's SPR technology, designed specifically for the study of biomolecular binding events is, therefore, a powerful and key tool in cancer research and is already bringing major benefits to areas, such as, DNA research, cell cycle control, apoptosis, signal transduction, angiogenesis, clinical studies and drug discovery.
As well as isolating specific ligands to a target molecule, Biacore's SPR technology also provides quantitative, real-time information about the binding event itself kinetics, affinity, specificity and the active concentration of biomolecules in solution.
Application in cancer research
The term proteomics describes the study of the entire protein complement of the genome.
A key aim in proteomics is to discover new targets for drug discovery but the eventual objective is to identify, characterise and assign function to all proteins.
Gene sequence data alone does not provide information about protein function or disease relevance. Understanding protein function is the key to gaining insights into the biochemical mechanisms that underlie disease and to the identification of new drug targets.
Determining protein function is therefore vital to the overall utility of proteomics research and bridges the gap between the information available from the sequencing of the human genome and the information needed for drug discovery.
As Biacore's SPR technology has been specifically designed for the study of biomolecular binding events, and the isolation and micropurification of native molecules to a target, it is a powerful and key tool for the functional analysis of the proteome in proteomics research. It is already bringing major benefits to areas such as ligand fishing, protein-protein interactions, binding site analysis and protein complex analysis.
It is important that proteins, as far as possible, are studied in their native state and environment. When investigating protein function, it is also important to conserve their native three-dimensional structure as this may be essential for in vivo function.
Biacore's sensor chips are designed to maintain the integrity and function of bound target molecules, as far as possible, and also allow membrane associated proteins to be studied. This enables the target to be studied in conditions close to those in vivo.
A variety of Biacore's SPR instruments are available to suit the needs of every proteomics and cancer research laboratory from the high throughput labs that require unattended, fully automated sample analysis to the smaller laboratory, where a large range of researchers may wish to study a variety of different molecules.
Biacore's SPR technology can also be linked to mass spectrometry (MS) to combine identification with mass determination and protein characterisation.
For further information, contact Biacore at www.biacore.com
