Sensitive detection of analytes with NOVAswitch aptasensors (aptamer-based biosensors).

Aptamers are chemically synthesized oligonucleotides. They can be easily conjugated to various functional groups for grafting, immobilization or signaling. This includes (but is not restricted to) amino -NH2, carboxyl -COOH or thiol -SH functions, fluorescent (FAM, Alexa, Cyanine) or redox (methylene blue) reporters. A spacer can be introduced between these groups and either the 5’ or 3’ end of the aptamer. Through our privileged partner, Eurogentec , our customers have full access to such functionally modified aptamers.

Aptamers are easily integrated into biosensors, utilizing different signal transduction modalities. At Novaptech, we routinely use Surface Plasmon Resonance (SPR) and Bio-Layer Interferometry (BLI) to characterize aptamer binding (see Novaptech Custom aptamer selection & characterization process).

Additionally, Novaptech engineers aptamer-based fluorescent and colorimetric aptasensors for the detection of small molecules and proteins.

Our NOVAswitches simplify the design of aptasensors. The structural change induced by the binding of the target to its cognate aptamer is exploited for the emission of a signal. Therefore, our NOVAswitch aptasensors ensure the specific and quantitative detection of the molecule of interest.

NOVAswitch in its free form (left) and bound to its target (right).

Novaptech can engineer an aptasensor from your own aptamer. Please, contact us!

Fluorescent NOVAswitch Aptasensors

Fluorescence can be used in several ways in NOVAswitch aptasensors:

Molecular beacons
Energy transfer
Fluorescence anisotropy

Molecular beacon – Structure switching aptamers NOVAswitch

A bi-molecular beacon is a combination of a fluorescently labeled aptamer and a short complementary oligonucleotide conjugated to a quencher. The close proximity between the fluorophore and the quencher inhibits the emission of light. When the target molecule binds to the aptamer, it induces a structural change of the NOVAswitch and releases the quenching oligonucleotide, resulting in fluorescence emission that correlates with the target molecule concentration.

Scheme of a molecular beacon assay

Novaptech has designed molecular beacon-based aptasensors for the detection of various antibiotics .The example below shows a typical assay for the specific detection of streptomycine by a beacon made of a FAM-conjugated NOVAswitch combined with a BHQ1-conjugated complementary oligonucleotide.

Fluorescence intensity as a function of antibiotic concentration, demonstrating the specific detection of streptomycin using a molecular beacon-based NOVAswitch aptasensor.

Energy transfer with Lanthanide nanoparticles - Structure switching aptamers NOVAswitch

Lanthanide (La) nanoparticles (NPs) are characterized by bright, long-lived fluorescence emission. This fluorescence is quenched when energy is transferred to a dye. In collaboration with Poly-DTech , we designed a NOVAswitch-based aptasensor that combines La particles grafted with a short oligonucleotide complementary to part of an aptamer, which is conjugated to a dye.

The binding of the target molecule to the aptamer induces a structural change of the NOVAswitch and dissociates the oligonucleotide hybrid, thereby preventing energy transfer from the nanoparticle to the acceptor, hence allowing the fluorescence emission of the La particle.

Scheme of La-NP-based NOVAswitch aptasensor

Novaptech has developed La-nanoparticle-based aptasensors for the detection of various fungicides.

The example below shows a typical assay for the specific and quantitative detection of thiabendazole (TBZ) using a sensor that includes a dye-conjugated NOVAswitch. La NP-NOVAswitch aptasensors, which enable the measurement of delayed fluorescence, are particularly useful for the quantitative detection of analytes in complex matrices with high fluorescence backgrounds.

FRET response as a function of thiabendazole (TBZ) concentration, using La NP-based NOVAswitch aptasensors

Fluorescence anisotropy-based aptasensor - Structure switching aptamers NOVAswitch

In this particular format we take advantage of the change in the molecular weight, i.e. the mobility that accompanies the conformational modification of the NOVAswitch in response to the binding to its target.

An improved design of the kissing complex-based aptasensor for the detection of adenosine

Goux, E., Lisi, S., Ravelet, C., Durand, G., Fiore, E., Dausse, E., Toulmé, J. J., & Peyrin, E.

Keywords: aptamer biosensing, dual recognition, nucleic acid assembly, aptaswitch, loop-loop interactions, kissing complex, fluorescence anisotropy, adenosine-adenoswitch, fluorophore-labeled aptakiss, blocking DNA, detection limit, human serum, biosensor optimization.

Electronic conduction (BIOFET) NOVAswitch aptasensors

Novaptech, in collaboration with Grapheal is developing a BIOFET-based NOVAswitch sensor for the detection of biomolecules of interest in health sciences. Grapheal is a French spin-off from CNRS-Grenoble which markets a new sensitive material, graphene on polymer, capable of measuring variations in electronic conduction (BIOFET principle). It has developed “GraphLAB” a chemical and biological field analytical sensor capable of transforming this interaction measurement into a quantitative information for the user.

Colorimetric aptasensors – NOVAswitches

Gold nanoparticles (AuNPs) have the unique ability to change color depending on their aggregation or dispersion state due to the plasmon resonance property of colloidal gold. Novaptech has exploited this property to design NOVAswitch aptasensors. Two batches of AuNPs are functionalized: one with the aptamer and the other with a short oligonucleotide complementary to part of the aptamer. Duplex formation leads to AuNPs aggregation, resulting in a blue-colored solution. When the target molecule binds to the aptamer, it induces a structural change that disrupts the duplex, causing the GNP aggregates to dissociate and shifting the color to red.

Illustration of a gold nanoparticles-based aptasensor

Novaptech has designed AuNP-based aptasensors for the detection of fungicides. The example below shows a typical assay for the specific and quantitative detection of thiabendazole (TBZ) using a AuNP-based NOVAswitch sensor.

Colorimetric detection of TBZ. Upon TBZ binding to its cognate NOVAswitch aptamer immobilized on AuNPs, the absorption peak of AuNPs shifts to a lower wavelength.

Point of care/ in the field NOVAswitch aptasensors

Rapid, simple, accurate, and cost-effective tests are valuable for various applications in health, agri-food, and environmental sectors. To this end, Novaptech, in collaboration with Goyalab , has developed IndiGo, a light, user-friendly, handheld and smartphone-connected spectrophotometer for absorption and emission measurements. It is fully compatible with NOVAswitch.