User:Whomtao/Bio-layer interferometry

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Bio-layer interferometry (BLI) is a optical biosensing technology that analyzes biomolecular interactions in real-time without the need for fluorescent labeling. Alongside Surface Plasmon Resonance(SPR), BLI is one of few widely available label-free biosensing technologies, a detection style that allows for a higher volume of information to be obtained in a quicker amount of time compared to traditional processes. The technology relies on the phase shift-wavelength correlation created between interference patterns off of two unique surfaces on the tip of a biosensor. BLI has significant applications in quantifying binding strength, measuring protein interactions, and identifying properties of reaction kinetics, such as rate constants and reaction rates.

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Explanation

Applications

Differences between BLI and SPR

Biosensor Type and Selection
Bio-layer interferometry utilizes the interference of two waves of differing intensities to determine the level of interaction between two molecules.

BLI monitors interactions between a ligand, immobilized on either the tip of a biosensor or the assay, and an analyte in solution.

The binding between a ligand immobilized on the biosensor tip surface and an analyte in solution produces an increase in optical thickness at the biosensor tip, which results in a wavelength shift, Δλ (Figure 3), which is a direct measure of the change in thickness of the biological layer. Interactions are measured in real time, providing the ability to monitor binding specificity, rates of association and dissociation, or concentration, with high precision and accuracy.

Only molecules binding to or dissociating from the biosensor can shift the interference pattern and generate a response profile. Unbound molecules, changes in the refractive index of the surrounding medium, or changes in flow rate do not affect the interference pattern. This is a unique characteristic of bio-layer interferometry and extends its capability to perform in crude samples used in applications for protein-protein interactions, quantitation, affinity, and kinetics.

don't forget to mention "dip-and-read)

Analyzing Biomolecular Interactions
A key use of Bio-layer interferometry is to analyze and quantify interactions between sets of biomolecules. This is extremely useful in pharmaceutical research, in which biomolecule-membrane interaction determines characteristics of a given drug. Due to its ability to achieve high-resolution data and high throughput, BLI has been used to identify biophysical properties of lipid bilayers, allowing for an alternative method of study than the traditional in vitro methods currently used (microscopy, electrophoresis). In addition, BLI can be used to study effector complex-target interactions. Where the traditional Electrophoretic Mobility Shift Assay (EMSA) method can be used, BLI can act as a suitable substitute if the provided benefits (label-free, real-time measurements) are desired.

Measuring Biomolecular Kinetics

Bio-layer interferometry can be used to analyze kinetics in biomolecular systems. Interference patterns found in BLI experiments can be used to calculate rate constants and other kinetic data in biomolecular interactions. The (relatively) lower sensitivity of the BLI sensor results in less response to changes in sample composition. As a result, BLI can also be used to investigate allosteric effects on enzyme conformational changes.

Distinguishing Characteristics
BLI and SPR are both dominant technologies in the label-free instruments market. Despite sharing some similarities in concept, there are significant differences between the two techniques. Micro-fluidic SPR relies on a closed architecture to transport samples to a stationary sensor chip. BLI instead utilizes an open system, shaking multiple wells on a plate to transport the sensors to the samples without need for micro-fluidics. Being a closed system, SPR's association and dissociation phases are limited by the technology's design. BLI's open plate design results in association and dissociation length limits determined by sample evaporation instead. SPR is easily reproducible due to its continuous flow microfluidics. BLI's multi well plate design allows for extremely high throughput in one batch. Assay configuration in BLI can, in stable conditions, allow for recovery of samples. Assay configuration in SPR allows for higher sensitivity. As a result, BLI results are often compared to SPR results for validation.