Ratnesh Lal

We aimed to develop a scalable, low-power graphene field-effect transistor (GFET) crossbar array integrated with a neural-interface system-on-chip (NISoC), termed the GFET-NISoC platform, for multiplexed, real-time biomolecular sensing, including ultra-sensitive detection of Pb²⁺ using a G-quadruplex ssDNA aptamer. A 12×12 GFET crossbar array was fabricated using a multilayer dielectric stack and optimized graphene transfer. Characterization was performed using both benchtop instrumentation and a low-power NISoC front-end supporting current- and voltage-clamp modes. Ionic strength, pH, and Pb²⁺ dose-response measurements were obtained from functionalized arrays. The platform demonstrated uniform Dirac behavior, reversible pH-dependent conductance, predictable ionic-strength responses, and femtomolar-level Pb²⁺ detection with strong selectivity over Ca²⁺ and Co²⁺. The GFET-NISoC system enables multiplexed, multi-modal sensing at sub-μW power levels and provides a promising foundation for wearable or point-of-care chemical and biomolecular monitoring.

Amyloid β (Aβ) ion channels destabilize cellular ionic homeostasis, which contributes to neurotoxicity in Alzheimer’s disease. The relative roles of various Aβ isoforms are poorly understood. We use bilayer electrophysiology, AFM imaging, circular dichroism, FTIR and fluorescence spectroscopy to characterize channel activities of four most prevalent Aβ peptides, Aβ_1-42, Aβ_1-40, and their pyroglutamylated forms (AβpE_3-42, AβpE_3-40) and correlate them with the peptides’ structural features. Solvent-induced fluorescence splitting of tyrosine-10 is discovered and used to assess the sequestration from the solvent and membrane insertion. Aβ_1-42 effectively embeds in lipid membranes, contains large fraction of β-sheet in a β-barrel-like structure, forms multi-subunit pores in membranes, and displays well-defined ion channel features. In contrast, the other peptides are partially solvent-exposed, contain minimal β-sheet structure, form less-ordered assemblies, and produce irregular ionic currents. These findings illuminate the structural basis of Aβ neurotoxicity through membrane permeabilization and may help develop therapies that target Aβ-membrane interactions.

Dementia is a brain disease which results in irreversible and progressive loss of cognition and motor activity. Despite global efforts, there is no simple and reliable diagnosis or treatment option. Current diagnosis involves indirect testing of commonly inaccessible biofluids and low-resolution brain imaging. We have developed a portable, wireless readout-based Graphene field-effect transistor (GFET) biosensor platform that can detect viruses, proteins, and small molecules with single-molecule sensitivity and specificity. We report the detection of three important amyloids, namely, Amyloid beta (Aβ), Tau (τ), and α-Synuclein (αS) using DNA aptamer nanoprobes. These amyloids were isolated, purified, and characterized from the autopsied brain tissues of Alzheimer’s Disease (AD) and Parkinson’s Disease (PD) patients. The limit of detection (LoD) of the sensor is 10 fM, 1–10 pM, 10–100 fM for Aβ, τ, and αS, respectively. Synthetic as well as autopsied brain-derived amyloids showed a statistically significant sensor response with respect to derived thresholds, confirming the ability to define diseased vs. nondiseased states. The detection of each amyloid was specific to their aptamers; Aβ, τ, and αS peptides when tested, respectively, with aptamers nonspecific to them showed statistically insignificant cross-reactivity. Thus, the aptamer-based GFET biosensor has high sensitivity and precision across a range of epidemiologically significant AD and PD variants. This portable diagnostic system would allow at-home and POC testing for neurodegenerative diseases globally.

We have developed a DNA aptamer-conjugated graphene field-effect transistor (GFET) biosensor platform to detect receptor-binding domain (RBD), nucleocapsid (N), and spike (S) proteins, as well as viral particles of original Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) coronavirus and its variants in saliva samples. The GFET biosensor is a label-free, rapid (≤20 min), ultrasensitive handheld wireless readout device. The limit of detection (LoD) and the limit of quantitation (LoQ) of the sensor are 1.28 and 3.89 plaque-forming units (PFU)/mL for S protein and 1.45 and 4.39 PFU/mL for N protein, respectively. Cognate spike proteins of major variants of concern (N501Y, D614G, Y453F, Omicron-B1.1.529) showed sensor response ≥40 mV from the control (aptamer alone) for fM to nM concentration range. The sensor response was significantly lower for viral particles and cognate proteins of Middle East Respiratory Syndrome (MERS) compared to SARS-CoV-2, indicating the specificity of the diagnostic platform for SARS-CoV-2 vs. MERS viral proteins. During the early phase of the pandemic, the GFET sensor response agreed with RT-PCR data for oral human samples, as determined by the negative percent agreement (NPA) and positive percent agreement (PPA). During the recent Delta/Omicron wave, the GFET sensor also reliably distinguished positive and negative clinical saliva samples. Although the sensitivity is lower during the later pandemic phase, the GFET-defined positivity rate is in statistically close alignment with the epidemiological population-scale data. Thus, the aptamer-based GFET biosensor has a high level of precision in clinically and epidemiologically significant SARS-CoV-2 variant detection. This universal pathogen-sensing platform is amenable for a broad range of public health applications and real-time environmental monitoring.

Nonmesoporous Janus silica nanobowls (NBs) are unique in that they possess two different nonporous surfaces per particle for loading biological molecules and can thus be designed with multifunctional properties. Although silica NBs have been successfully employed for both targeted therapeutic and diagnostic applications, their ability to deliver DNA has not yet been fully explored. The purpose of this study was to design and develop an in vitro transfection agent that would exploit the distinct characteristics of the silica NB. First, we determined that the NB surface can be linked to either supercoiled cDNA plasmids or vectorless, linear cDNA constructs. Additionally, the linearized cDNA can be functionalized and chemisorbed on NBs to obtain a controlled release. Second, the successful transfection of cells studied was dependent on lipid coating of the NB (LNBs). Although both NBs and LNBs were capable of undergoing endocytosis, NBs appeared to remain within vesicles as shown by transmission electron microscopy (TEM). Third, fluorescence microscopy and Western blotting assays revealed that transfection of four different cell lines and acutely isolated rat sensory neurons with LNBs loaded with either linear or supercoiled cDNA constructs coding for the fluorescent protein, clover and tdTomato, resulted in protein expression. Fourth, two separate opioid receptor–ion channel signaling pathways were functionally reconstituted in HEK cells transfected with LNBs loaded with three separate cDNA constructs. Overall, these results lay the foundation for the use and further development of LNBs as in vitro transfection agents.

The burgeoning field of nanotechnology aims to create and deploy nanoscale structures, devices, and systems with novel, size-dependent properties and functions. The nanotechnology revolution has sparked radically new technologies and strategies across all scientific disciplines, with nanotechnology now applied to virtually every area of research and development in the US and globally. NanoFlorida was founded to create a forum for scientific exchange, promote networking among nanoscientists, encourage collaborative research efforts across institutions, forge strong industry-academia partnerships in nanoscience, and showcase the contributions of students and trainees in nanotechnology fields. The 2019 NanoFlorida International Conference expanded this vision to emphasize national and international participation, with a focus on advances made in translating nanotechnology. This review highlights notable research in the areas of engineering especially in optics, photonics and plasmonics and electronics; biomedical devices, nano-biotechnology, nanotherapeutics including both experimental nanotherapies and nanovaccines; nano-diagnostics and -theranostics; nano-enabled drug discovery platforms; tissue engineering, bioprinting, and environmental nanotechnology, as well as challenges and directions for future research.

The past two decades have witnessed a dramatic progress in the development of novel super-resolution fluorescence microscopy technologies. Here we report a new fluorescence imaging method, called metamaterial assisted photobleaching microscopy (MAPM), which possesses a nanometer-scale axial resolution and suitable for broadband operation across the entire visible spectrum. The photobleaching kinetics of fluorophores can be greatly modified via a separation-dependent energy transfer process to a nearby metamaterial. The corresponding photobleaching rate is thus linked to the distance between the fluorophores and the metamaterial layer, leading to a reconstructed image with exceptionally high axial resolution. We apply the MAPM technology to image the HeLa cell membranes tagged with fluorescent proteins and demonstrate an axial resolution of ~2.4 nm with multiple colors. MAPM utilizes a metamaterial-coated substrate to achieve super-resolution without altering anything else in a conventional microscope, representing a simple solution for fluorescence imaging at nanometer axial resolution.

Non-invasive and label-free calorimetry could become a disruptive technique to study single cell metabolic heat production without altering the cell behavior, but it is currently limited by insufficient sensitivity. Here, we demonstrate microfluidic single-cell calorimetry with 0.2-nW sensitivity, representing more than ten-fold enhancement over previous record, which is enabled by (i) a low-noise thermometry platform with ultralow long-term (10-h) temperature noise (80 μK) and (ii) a microfluidic channel-in-vacuum design allowing cell flow and nutrient delivery while maintaining a low thermal conductance of 2.5 μW K⁻¹. Using Tetrahymena thermophila as an example, we demonstrate on-chip single-cell calorimetry measurement with metabolic heat rates ranging from 1 to 4 nW, which are found to correlate well with the cell size. Finally, we perform real-time monitoring of metabolic rate stimulation by introducing a mitochondrial uncoupling agent to the microchannel, enabling determination of the spare respiratory capacity of the cells.