WAVE kinetics

Grating-coupled interferometry with the Creoptix WAVEdelta
ApplicationsTechnology

The new Wave in Kinetics

The Creoptix™ WAVEsystem: High-throughput, automated, clog-free, label-free kinetics. Get superior affinity and kinetic data, even for the most challenging samples.

Confidently detect and quantify biological interactions in real-time. Works especially well with small molecule fragments and compounds. Suited for high-throughput screening campaings.

Superior resolution than traditional SPR due to the waveguide technology.
Read more below!

Affinity

LIGAND BINDING


KD
Equilibrium dissociation constant. Can be obtained by kinetic or classical equilibrium binding analysis. Provides information about the strength but not the dynamics of an interaction.


ΔH 
Binding enthalpy. KD values at different temperatures can be used to obtain the binding enthalpy of an interaction via vant-Hoff-plots.

 

Kinetics

BINDING KINETICS


ka (kon)
Association rate constant. Provides information on how fast complexes form; can be used for KD determination.


kd (koff)
Dissociation rate constant. Provides information on how fast complexes dissociate; can be used for KD determination.


KD
Equilibrium dissociation constant. Can be obtained by kinetic or classical equilibrium binding analysis. Provides information about the strength but not the dynamics of an interaction.

The WAVEsystem excels in small molecule analysis

For the WAVEsytem, all sizes and affinities are welcome: Screen, rank and characterize fragments, including weak binders with fast dissociation rates and very strong binders, to large particles. Profile kinetics closer to real life. Assess drug performance in undiluted human blood serum and plasma samples. Also, the WAVEsystem cannot just do buffers: Explore solubilization and purification conditions for membrane proteins, including viscous detergents, less common solvents, and different additives. The sophisticated chip technology ensures no clogging, regardless of the sample and solution type.

Screening

The WAVEsystem features 4 parallel lines and rapid kinetics analysis so that screening through large libraries is a breeze!

Ligand binding

Characterize ligand binding for almost any type of sample: Proteins, peptides, challenging membrane proteins, DNA, RNA, aptamers, small molecules, fragments, lipids, particles.

Biologics development

Characterize and quantify your development antibodies at low limits of detection. Not only in buffer, but also in complex biofluids or cell culture media.

Pharmacology

Measure binding kinetics in biofluids like human blood serum or plasma for drug potency assessment. No clogging due to highly sophisticated chip technology.

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  • Compound screening
  • Fragment screening
  • Off-rate screening
  • Full affinity and single-dose
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  • Kinetic binding analysis
  • Steady-state binding analysis
  • Single-dose
  • Competition
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  • Antibody off-rate ranking
  • Ligand screening
  • Avidity testing
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  • Ligand binding in biofluids
  • E,g, human serum, plasma, liquord
  • E.g. cell lysates
  • Quantitation in crude extracts

why WAVE and not SPR: the technology

SPR

SPR sensors are usually glass slides coated with a chemically inert metal (black and blue bar). When plane-polarized light hits the metal film at a defined angle (red lines), so-called surface plasmons are excited. Their electromagnetic field (orange bars) is called an evanescent wave and extends beyond the metal surface. Changes in refractive index above the surface, e.g. from ligand-analyte binding, are sensed. This has several limitations:

  • Complicated fluidics and detection arrangements necessary
  • Detection area limited to evanescent wave field
  • Ligand-analyte interaction outside of evancescent field are “invisible”

Wave-guide interferometry

GCI uses a similar metal-coated sensor surface. However, below the sensor, a laser beam (red lines) is coupled into a waveguide via a grating. This creates an evanescent field (horizontal bars) across the full lenght of the sensor surface. A reference beam is similarly coupled into the waveguide, leading to interferences between sample and reference beams within the waveguide. Once an analyte binds to the immobilized ligand, the refractive index change translates to a phase shift in the sample beam. This generates a high-resolution, time-dependent, and robust phase shift in the interference pattern. This setup brings several advantages:

  • Significantly enhanced detection sensitivity
  • Detection across the whole surface
  • More shallow field penetration in the solution
  • Less artifacts by bulk refractive index changes
  • Better signal/noise ratio

Evanescent field in SPR

Waveguide setup 

GCI: Grating-coupled interferometry

The technology behind the Creoptix™ WAVEsystem is called Grating-Coupled Interferometry (GCI). Similar to surface plasmon resonance (SPR), an evanescent field is used to measure changes in the refractive index close to a sensor surface. Upon binding of an analyte (in solution) to the ligand molecule (immobilized on the sensor surface), the local refractive index increases, proportional to the number of bound analytes. Thus, the refractive index shift is equivalent to the change in mass on the sensor surface.

In contrast to SPR, however, GCI uses a sophisticated detection system to significantly enhance detection sensitivity. Below the sensor surface, a laser beam is coupled into a waveguide via a grating. This creates an evanescent field across the full lenght of the sensor surface. A reference beam is similarly coupled into the waveguide, leading to interferences between sample and reference beams within the waveguide. Once an analyte binds to the immobilized ligand, the refractive index change translates to a phase shift in the sample beam. This generates a high-resolution, time-dependent and robust phase shift in the interference pattern. This interference shift is then converted to the classical kinetic sensorgrams as in SPR. A further advantage of GCI is that the evanescent field penetrates less deep into the sample. This minimizes disturbances caused by bulk refractive index changes and increases the signal-to-noise ratio.

Detection setup of GCI

WAVEsystem Advantages

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Broadest kinetic range

Quantify binding affinities (KD) from low pM to high µM with confident kinetic analysis.

Tight binders welcome

Measure kinetics of high-affinity binders at low pM affinity (KD) values.
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All sizes welcome

Screen, rank and characterize small analytes and fragments, including weak binders with fast dissociation rates (koff as high as 10/sec).

Kinetic profiling closer to real life

Assess drug performance in undiluted human blood serum and plasma samples.

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More than just buffers

Explore more solubilization and purification conditions for membrane proteins, including viscous detergents, less common solvents, and different additives.

No clogging, regardless of size

Analyze and characterize larger molecules (VLPs, liposomes) and aggregates (fibrils) without clogging-risk.

Frequently asked questions

FAQ – General

What is the difference between SPR and GCI?

Both technologies are based on evanescent field sensing. The major difference between GCI and SPR is that in GCI the light travels in a waveguide underneath the sensor surface, pretty much like in an optic fibre. Because of this, the evanescent wave in GCI travels along the entire surface of the chip and is not static like the SPR, in which only around 100 nm along the chip are “sensed”. This means that in GCI the evanescent waves “senses” a lot more interactions than SPR plasmons do. Thus, GCI is inherently more sensitive than SPR.

What surface chemistries can be used for ligand immobilization?

All common immobilization strategies used also in SPR can be directly transferred to Creoptix™WAVE sensors chips: Amine-coupling, Ni-NTA-coupliung, Streptavidin-Biotin capture, and others.

What are k-on and k-off ranges that can be measured with the Creoptix™ WAVEsystem?

Association rates constants range from 103 to 5·107 M-1s-1 for small molecules and 103 to 3·109 M-1s-1 for large molecules. Dissociation rate constants can be measured in the range between 10-5 and 10 per second.

What is the molecular weight (MW) limit for analytes?

 Analyte:ligand molecular weight ratios up to 1:1000 are possible.

Is the WAVE system compatible with crude sample material?

Absolutely. By combining the microfluidics and the biosensor in a single disposable cartridge, the Creoptix™ WAVE system offers crude-sample robustness normally only achieved with plate-based assays. Compatible biofluids include 100% serum or plasma, cell lysates, crude membrane preparations, large drug targets, virus-like particles (VLPs), liposomes, and aggregates (fibrils); all without the risk of clogging the system.

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