The Octet R8: a fast, sensitive, label-free protein analysis platform for characterising biomolecular interactions

Interactions between molecules are central to all processes of life. These interactions are diverse in nature and include protein-protein interactions, protein-oligonucleotide interactions, protein-cofactor interactions and interactions involving complex bilayer assemblies. Because of their importance in terms of furthering our biological understanding and accelerating pharmaceutical development, all of these interactions are at the heart of innovative research programs across the globe. Life science research at the University of Leicester has an outstanding track record in conducting groundbreaking research in this important field. To allow the world-leading researchers to drive innovation and address the most important biological questions of our time requires highly sensitive and accurate quantitative analytical tools. The heavily used instrumentation at UoL is ageing and has limitations compared to the currently available techniques. We, therefore, seek to purchase the Sartorius Octet R8.

The Octet R8 is a state-of-the-art in bio-layer interferometry (BLI), an optical technique for detecting biomolecular interactions by measuring the interference pattern of white light reflected from the surface of a biosensor. BLI compares the interference pattern of white light reflected from an internal reference layer with that reflected from the immobilised protein layer present on the biosensor tip. The binding of biomolecules (analytes) at the tip surface results in a shift in interference pattern allowing the quantification of binding events in real-time and with high precision. Only interactions of bound biomolecules are measured, making bio-layer interferometry a powerful analytical tool for a wide range of applications. The Octet R8 system measures binding events in small volumes of 200 microliters in a non-destructive manner to determine association rates (kon), dissociation rates (koff), and affinity constants (KD). Arguably one of the most powerful features of the Octet systems is the unique 'dip and read' biosensor format which permits the analysis of complex samples e.g. cell culture supernatants and lysates enabling biomolecular interactions to be studied in biologically relevant environments. Due to the multitude of different biosensors and surface chemistries BLI provides enormous flexibility for a range of multidisciplinary projects.

The label-free Octet system negates the need for intrinsic/external chromophores or the addition of fluorescent labels/probes: a prerequisite of solution optical spectroscopies. Although a competing technology like isothermal titration calorimetry (ITC) is a label-free technology that is very useful for providing thermodynamic data, it does not allow for real-time kinetic analyses and is less sensitive than the Octet system. Surface Plasmon Resonance (SPR, a comparable label-free, surface-based technology) generates highly accurate, reproducible data but does not have the high throughput or flexibility offered by the Octet platform and poses significant challenges associated with its microfluidics-based nature.

The Sartorius Octet R8 platform we seek here will provide continuity to our biolayer interferometry platform by replacing a 10-year-old Octet QKe, which will no longer be supported by the manufacturer by the end of 2022. The R8 will also constitute a significant step-change in our capabilities to measure biomolecular interactions due to the following improvements: (1) The Octet QKe provides enough sensitivity to measure interactions involving molecules with molecular weight as low as 10 kD, but not below. The R8 is advertised to measure down to 150 Da, and our own research suggests that it can will measure reliably to 300 Da. This corresponds to a more than 10-fold improvement in sensitivity, which provides access to most small molecules involved in biomolecular interactions. (2) The Octet QKe operates at 0.3 and 0.6 Hz using one spectrophotometer for all 8 channels. In contrast, each channel in the R8 has its dedicated spectrometer, which allows it to operate at 2, 5 or 10 Hz, yielding a more than 10-fold higher sampling rate, which is critical for measuring fast on and off rates. This permits the quantification of a wider range of affinities from 1 mM to 10 pM (compared with 0.1 mM-0.1 nM for the Octet QKe). (3) While the QKe is limited to operation above 28C, the Octet R8 features a cooling system that allows controlling the temperature down to 15C. This feature is critical for successfully measuring temperature-sensitive systems. (4) The R8 comes with a microplate evaporation cover that extends the observation time from 2.5 h to 12 h, which is instrumental for accurate measurements of very slow off rates. It also allows for measuring a full 96-well plate with minimal user intervention (6-12h run time). In summary, the Octet R8 will not only serve the current level of need for measuring binding kinetics but expand our capabilities in BLI to small molecules, drugs and more challenging biomolecular interactions.