Long-lasting and responsive DNA/enzyme-based programs in serum-supplemented extracellular media

DNA molecular programs are emerging as promising pharmaceutical approaches due to their versatility for biomolecular sensing and actuation. However, the implementation of DNA programs has been mainly limited to serum-deprived in vitro assays due to the fast deterioration of the DNA reaction networks by the nucleases present in the serum. Here, we show that DNA/enzyme programs are functional in serum for 24h but are latter disrupted by nucleases that give rise to parasitic amplification. To overcome this, we implement 3-letter code networks that suppress autocatalytic parasites while still conserving the functionality of DNA/enzyme programs for at least 3 days in the presence of 10% serum. In addition, we define a new buffer that further increases the biocompatibility and conserves responsiveness to changes in molecular composition across time. Finally, we demonstrate how serum-supplemented extracellular DNA molecular programs remain responsive to molecular inputs in the presence of living cells, having responses 6-fold faster than cellular division rate and are sustainable for at least 3 cellular divisions. This demonstrates the possibility of implementing in situ biomolecular characterization tools for serum-demanding in vitro models. We foresee that the coupling of chemical reactivity to our DNA programs by aptamers or oligonucleotide conjugations will allow the implementation of extracellular synthetic biology tools, which will offer new biomolecular pharmaceutical approaches and the emergence of complex and autonomous in vitro models.

Fabrication robustness in BIC metasurfaces

All-dielectric metasurfaces supporting photonic bound states in the continuum (BICs) are an exciting toolkit for achieving resonances with ultranarrow linewidths. However, the transition from theory to experimental realization can significantly reduce the optical performance of BIC-based nanophotonic systems, severely limiting their application potential. Here, we introduce a combined numerical/experimental methodology for predicting how unavoidable tolerances in nanofabrication such as random geometrical variations affect the performance of different BIC metasurface designs. We compare several established all-dielectric BIC unit cell geometries with broken in-plane inversion symmetry including tilted ellipses, asymmetric double rods, and split rings. Significantly, even for low fabrication-induced geometrical changes, both the BIC resonance amplitude and its quality factor (Q-factor) are significantly reduced. We find that the all-dielectric ellipses maintain the highest Q-factors throughout the geometrical variation range, whereas the rod and split ring geometries fall off more quickly. The same behavior is confirmed experimentally, where geometrical variation values are derived from automated processing of sets of scanning electron microscopy (SEM) images. Our methodology provides crucial insights into the performance degradation of BIC metasurfaces when moving from simulations to fabricated samples and will enable the development of robust, high-Q, and easy to manufacture nanophotonic platforms for applications ranging from biomolecular sensing to higher harmonic generation.

Hybridizing nickel platform using ultra-short pulsed laser: a new paradigm towards biomedical sensing and theranostics

A self-assembled 3D nanonetwork of Nickel and Nickel Oxide is synthesized by ultrashort pulsed laser through multiphoton ionization. The synthesized nanonetwork with tunable physiochemical property was investigated for cancer therapeutic and biomolecular sensing applications. In this thesis, the developed 3D nickel nanomatrix effectively regulated HeLa cancer cell adhesion and proliferation mimicking Extracellular Matrix (ECM). This behaviour explicitly demonstrated that the initial incubation period was devoted to baiting fibroblast and HeLa cells to proliferate upon the nanomatrix and subsequently the same nanomatrix exhibited cell trapping behaviour upon HeLa cells after an increased incubation period thereby controlling proliferation. The results brought new insight as to how HeLa cells behaved differently when compared to NIH3T3 fibroblast cells opening pioneering application in drug-free cancer therapy. To delve deeper into nickel nanonetwork for cancer therapy the laser ionization was manipulated to induce two distinct quantum theranosomes. Presently, quantum materials are limited due to 0D & 1D materials lacking biocompatibility resulting in coated materials with labelled tags for fluorescence excitation. The theranosomes mimicked tumor microenvironment by selectively accelerating the proliferation of mammalian fibroblasts cells while inducing cancer therapy. Furthermore, the theranosomes opened up label-free bioimaging probe for differentiating (HeLa & MDAMB-231) from mammalian fibroblast cells for cancer diagnostics. In-addition to label-free bioimaging, the development of an ultrasensitive biosensor for targeted biomolecule sensing was developed addressing the drawback faced with fluorescence imaging using Surface Enhanced Raman Scattering (SERS). We developed a SERS active nano-biosensor to detect chemical dye Crystal Violet (CV) and biomolecule glutathione(GSH). The Raman detection of crystal violet (CV) and glutathione (GSH) molecules was noted with 1 pM (1×10-12M) concentrations at (532 & 785nm) excitation wavelengths with an enhancement factor of 109, not been observed even in plasmonic materials. This extends the limit of detection (LOD), confirming suitability for chemical and biomolecular sensing. Additionally, the quantum confinement effect will result in an ultrasensitive sensor diagnosing and differentiating cancer cells from fibroblast cells. Based on the results in this thesis, the multifunctional feasibility of nano and quantum scale nickel structures arranged in 3D assembly for its direct application in cancer therapeutics, encompassing cancer bioimaging and diagnostics.

Hybridizing nickel platform using ultra-short pulsed laser: a new paradigm towards biomedical sensing and theranostics

A self-assembled 3D nanonetwork of Nickel and Nickel Oxide is synthesized by ultrashort pulsed laser through multiphoton ionization. The synthesized nanonetwork with tunable physiochemical property was investigated for cancer therapeutic and biomolecular sensing applications. In this thesis, the developed 3D nickel nanomatrix effectively regulated HeLa cancer cell adhesion and proliferation mimicking Extracellular Matrix (ECM). This behaviour explicitly demonstrated that the initial incubation period was devoted to baiting fibroblast and HeLa cells to proliferate upon the nanomatrix and subsequently the same nanomatrix exhibited cell trapping behaviour upon HeLa cells after an increased incubation period thereby controlling proliferation. The results brought new insight as to how HeLa cells behaved differently when compared to NIH3T3 fibroblast cells opening pioneering application in drug-free cancer therapy. To delve deeper into nickel nanonetwork for cancer therapy the laser ionization was manipulated to induce two distinct quantum theranosomes. Presently, quantum materials are limited due to 0D & 1D materials lacking biocompatibility resulting in coated materials with labelled tags for fluorescence excitation. The theranosomes mimicked tumor microenvironment by selectively accelerating the proliferation of mammalian fibroblasts cells while inducing cancer therapy. Furthermore, the theranosomes opened up label-free bioimaging probe for differentiating (HeLa & MDAMB-231) from mammalian fibroblast cells for cancer diagnostics. In-addition to label-free bioimaging, the development of an ultrasensitive biosensor for targeted biomolecule sensing was developed addressing the drawback faced with fluorescence imaging using Surface Enhanced Raman Scattering (SERS). We developed a SERS active nano-biosensor to detect chemical dye Crystal Violet (CV) and biomolecule glutathione(GSH). The Raman detection of crystal violet (CV) and glutathione (GSH) molecules was noted with 1 pM (1×10-12M) concentrations at (532 & 785nm) excitation wavelengths with an enhancement factor of 109, not been observed even in plasmonic materials. This extends the limit of detection (LOD), confirming suitability for chemical and biomolecular sensing. Additionally, the quantum confinement effect will result in an ultrasensitive sensor diagnosing and differentiating cancer cells from fibroblast cells. Based on the results in this thesis, the multifunctional feasibility of nano and quantum scale nickel structures arranged in 3D assembly for its direct application in cancer therapeutics, encompassing cancer bioimaging and diagnostics.

NBD-based synthetic probes for sensing small molecules and proteins: design, sensing mechanisms and biological applications

Compounds with a nitrobenzoxadiazole (NBD) skeleton exhibit high reactivity toward biological nucleophilies accompanied by distinct colorimetric and fluorescent changes, environmental sensitivity, and small size, all of which facilitate biomolecular sensing and self-assembly.

Comprehensive Understanding of Silicon-Nanowire Field-Effect Transistor Impedimetric Readout for Biomolecular Sensing

Impedance sensing with silicon nanowire field-effect transistors (SiNW-FETs) shows considerable potential for label-free detection of biomolecules. With this technique, it might be possible to overcome the Debye-screening limitation, a major problem of the classical potentiometric readout. We employed an electronic circuit model in Simulation Program with Integrated Circuit Emphasis (SPICE) for SiNW-FETs to perform impedimetric measurements through SPICE simulations and quantitatively evaluate influences of various device parameters to the transfer function of the devices. Furthermore, we investigated how biomolecule binding to the surface of SiNW-FETs is influencing the impedance spectra. Based on mathematical analysis and simulation results, we proposed methods that could improve the impedimetric readout of SiNW-FET biosensors and make it more explicable.