Biolayer Interferometry (BLI)

Bio-Layer Interferometry (BLI) is an optical technique for measuring macromolecular interactions by analyzing interference patterns of white light reflected from the surface of a biosensor tip.  BLI experiments are used to determine the kinetics and affinity of molecular interactions. In a BLI experiment, one molecule is immobilized to a Dip and Read Biosensor and binding to a second molecule is measured. A change in the number of molecules bound to the end of the biosensor tip causes a shift in the interference pattern that is measured in real-time. 

The CMI has two ForteBio instruments for measuring BLI: the Octet RED384 and the BLItz.  The Octet RED384 is more sensitive and higher throughput and can be used for macromolecular and small molecule binding experiments.  The BLItz uses a very small sample volume, but is more time consuming and cannot measure molecules smaller than 10 KDa.

Applications

• Kinetic binding: k_a, k_d
• Equilibrium binding: K_D
• Macromolecular and small molecule binding (Octet only)

Octet RED384 Key Features

• Simultaneous real-time binding on up to 16 biosensors
• Disposable biosensors (sensor regeneration not required)
• 96 or 384 well reagent plate format
• Small molecule sensitivity

BLItz Key Features

• Low sample volume: 4 μl sample
• Disposable biosensors (sensor regeneration not required)

 In a BLI experiment, one molecule is immobilized to a Dip and Read Biosensor and binding to a second molecule is measured. A change in the number of molecules bound to the end of the biosensor tip causes a shift in the interference pattern that is measured in real-time. Sensors are moved from one well of a microplate to another to change solution from samples (load sample and analyte sample) to buffer for baselines and dissocation.

Figure adapted from ForteBio. 

There are two major phases in a BLI experiment and each will need to be optimized for you experiment. Regeneration is an optional third phase that may be used when a single set of sensors is reused for multiple measurements.

Immobilization

• the Load molecule is attached irreversibly or reversibly to the sensor chip surface

Interaction analysis

• In the association phase, the Analyte is injected over the sensor surface
• In the dissociation phase, the Analyte is washed off the surface

Regeneration

• the surface is regenerated by removing remaining bound analyte or by removing ligand (which will also remove remaining analyte)