Electroactive nanocarbon materials as signaling tags for electrochemical PCR.

Electrochemical polymerase chain reaction (PCR) represents a valid alternative to the optical-based PCR due to reduced costs of signaling labels, use of simpler instrumentation, and possibility of miniaturization and portability of the systems, which can facilitate decentralized detection. The high intrinsic electroactivity and strong linear relationship between the material concentration and its redox signal suggest a possible use of oxidized nanocarbon materials as electroactive tags for PCR. Herein, we compared three different nanographene oxide materials namely nGO-1, nGO-2 and nGO-3 as signaling tags for the detection of genetically modified organisms (GMO) by electrochemical PCR. The three materials differ in size, chemical composition as well as type and amount of oxygen functionalities verified by extensive characterization with X-ray photoelectron spectroscopy (XPS), attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR), transmission electron microscopy (TEM) and electrochemical methods. A sense primer sequence belonging to the Cauliflower Mosaic Virus 35S promoter (a common genetic marker for GMO screening) was used to conjugate to the nanocarbon materials by carbodiimide chemistry before PCR amplification with a biotinylated antisense strand. Finally, the amplified electroactive PCR product was detected, where the reduction signal derived from the electrochemically reducible oxygenated functionalities on the nanocarbon material surface was directly correlated to the presence of GMO. Overall, we were able to correlate the different material characteristics with their performance as electroactive labels and identify the nanocarbon material that exhibits the highest potential to be used as innovative electroactive label for PCR in the amplification and detection of the selected target sequence.

Exploring graphene oxide intrinsic electroactivity to elucidate the non-covalent interactions with DNA oligonucleotides.

We show here how the electrochemical reduction signal of graphene oxide nanocolloids is inhibited upon the formation of non-covalent interactions with single stranded DNA oligonucleotides. The drop in the reduction current intensity is strongly influenced by the nucleobase sequence, and can therefore be directly correlated to the specific DNA homo-oligonucleotide.

Effect of surface chemistry on bio-conjugation and bio-recognition abilities of 2D germanene materials.

The interest of the scientific community for 2D graphene analogues has been recently focused on 2D-Xene materials from Group 14. Among them, germanene and its derivatives have shown great potential because of their large bandgap and easily tuneable electronic and optical properties. With the latter having been already explored, the use of chemically modified germanenes for optical bio-recognition is yet to be investigated. Herein, we have synthesized two germanene materials with different surface ligands namely hydrogenated germanene (Ge-H) and methylated germanene (Ge-Me) and used them as an optical platform for the label-free biorecognition of Ochratoxin A (OTA), a highly carcinogenic food contaminant. It was discovered that firstly the surface ligands on chemically modified germanenes have strong influence on the intrinsic fluorescence of the material; secondly they also highly affect both the bio-conjugation ability and the bio-recognition efficiency of the material towards the detection of the analyte. An improved calibration sensitivity, together with superior reproducibility and linearity of response, was obtained with a methylated germanene (Ge-Me) material, indicating also the better suitability of the latter for real sample analysis. Such research is highly beneficial for the development and optimization of 2D material based optical platforms for fast and cost-effective bioassays.

Unravelling the Aptamer-Analyte Interaction Dynamics through Fluorescence Quenching in Graphene Quantum Dots (GQDs) Based Homogeneous Assays.

Graphene quantum dots (GQDs) are used here as a biosensing platform for the recognition of the major food contaminant ochratoxin A (OTA), with a fluorescently labelled DNA aptamer (FAM OTA aptamer) functioning as the biorecognition element. The detection principle lies in the formation of noncovalent interactions between the FAM OTA aptamer and the GQD surface, and the consequent fluorescence quenching. The further change in the fluorescence signal, induced by the formation of the FAM OTA Aptamer/OTA conjugate during the detection step, could then be correlated to the presence and concentration of the target analyte. Upon tuning the concentration of GQDs, a switch in the biorecognition mechanism occurred. Specifically, while a lower GQD concentration (0.060 mg/mL) resulted in a restoration of the fluorescence intensity upon incubation with OTA, a higher GQD concentration (0.150 mg/mL) provided a further quenching of the final fluorescence intensity. Upon further calibration study, it was discovered that the latter mechanism provided a better option in terms of linearity of response, detection limit and selectivity.