switchSENSE^® Technology

DNA duplexes are forming, separating, and re-forming successfully since first life forms appeared on earth. 3.7 billion years later we are using DNA helix formation to zip molecules onto sensor spots. Sorry for taking so long, but we wanted to be sure it works just fine.

• heliX^® chip with DNA anchors
  The heliX^® chip surface is modified with covalently attached single stranded DNA anchor sequences, which have been optimized to bind adapters with ultimate specificity and stability.
• Ligand strand, exchangeable
  Your molecule of interest (the ligand) is immobilized onto the surface by the DNA nanolever, either by direct covalent coupling or by tag-capturing methods. Coupling of proteins to ligand-strands is readily performed using easy-to-use coupling kits.
• Adapter strand, exchangeable
  Adapters give you maximal experimental flexibility: use a standard adapter for binding applications or pick a long DNA-origami nanostructure to investigate conformational changes. Dyes are exchangeable, too. 

Hands-on sample preparation

Couple ligands covalently to ligand strands (e.g. via NHS) or use ready-to-use capture kits.

Fully automated workflow

Functionalization and regeneration are automated and DNA-based, so there is no need to scout for regeneration conditions.

The DNA-based functionalization offers precise control of the ligand density on the chip surface. By mixing nanolevers without ligands or nanolevers carrying different molecules of interest, the ligand density and the ratio of two ligands can be adjusted.

In static measurement mode, association and dissociation of molecules are detected in real-time using fluorescence. A constant negative potential is applied to the gold surface of the chip, keeping the DNA nanolever in an upright position.

The binding of molecules changes the local environment of the dye, thus causing a change in fluorescence (fluorescence proximity sensing). This is reverted when the molecule dissociates upon buffer flow. The chip surface can easily be regenerated to be used for further experiments.

By using the possibility of measuring two independent interactions simultaneously on the same sensor spot, four signals can be monitored in parallel, allowing for real-time referencing of two interactions at the same time or multiplexing of up to four interactions per run.

The two-color mode also enables the analysis of bispecific analytes like bispecific antibodies, yielding insights into the specific binary affinities as well as the avidity of bivalent binding. 

switchSENSE^® can be used to measure the binding, activity, as well as inhibition of nucleic acid-modifying enzymes like polymerases or reverse transcriptases. This assay format uses a mechanism called Surface Energy Transfer, by which fluorescence is quenched in close proximity to the gold surface of the chip.

The fluorescent dye is located at the end of the template strand. Upon injection of the enzyme and nucleotides, the complementary strand is synthesized. This results in the stiffening and stretching of the now double-stranded nucleic acid, increasing the distance of the dye from the quenching gold surface. The fluorescence change monitors the activity of the enzyme in real time. 

Two color assays additionally offer the possibility of Förster resonance energy transfer (FRET) experiments by using a donor-acceptor fluorochrome pair. This assay format can give unique insights into structural rearrangements of biomolecules, for example RNA secondary structures and loop formation.

The portfolio of dual-color assays is further expanded using DNA nanotechnology. Specialized Y-shaped DNA nanostructures bestowed with a fluorochrome FRET-pair are available for the detailed characterization of ternary interactions of bispecific binders. This assay format can be used for example to screen PROTACs and molecular glues or to investigate binding mechanisms of bispecific antibodies.

The target proteins can be functionalized on the end of the two FRET pair color-coded Y-arms of the structure. The Y-structure closes upon small molecule binding, and the subsequent ternary complex formation brings together the green donor and the red acceptor dye into a closer, FRET sensitive, distance. The change in red fluorescence signal intensity directly correlates with ternary complex formation kinetics.

With the heliX^® biosensor and its highly sensitive FRET read-out, it is possible to perform high-throughput screening and ranking of bifunctional small molecules and to differentiate between binary affinities and avidity of the ternary complex.

In the dynamic measurement mode, the DNA nanolevers are electrically actuated to oscillate at high frequencies. The fluorescence of the dye is quenched close to the gold surface. Monitoring the dye fluorescence thus allows the measurement of the speed of the nanolever movement.

The measured instantaneous velocity of the nanolever depends on the hydrodynamic friction of the attached molecules. Thus, changes in the upward motion of the nanolever can be used to determine ligand size and shape.

Larger molecules or conformational changes that cause an expansion of the molecule will slow down the movement, whereas smaller molecules or analyte-induced compaction of the molecule will increase the velocity of the motion.

Targeted protein degradation

Overcome the challenge of analyzing small molecules, which inhibit or stabilize protein-protein interactions (PPIs) using switchSENSE^® and the DNA Y-structure.

switchSENSE^® enables an effortless and precise control of target surface density and allows the adjustment of defined target ratios on the chip. This feature, together with the dual-color option and the advanced DNA Y-structure offer a diverse application range for multispecific binders from small molecules and peptides to therapeutic antibodies.

Its DNA-based chip design makes switchSENSE^® perfectly suited to investigate DNA- and RNA-binding molecules. The flexible assay design goes beyond simple binding kinetics and additionally allows the investigation of binding-induced structural changes like RNA-looping and aptamer folding. Furthermore, the contributions of different binding domains to the overall interaction can be differentiated. 

A unique application of switchSENSE^® is the measurement of catalytic rates of nucleic acid-modifying enzymes like polymerases or transcriptases. The influence of inhibitors on enzyme binding and catalytic activity can be differentiated. 

In dynamic measurement mode, switchSENSE^® enables the detection of binding-induced protein conformational changes and allows relative size comparisons of proteins of interest.