It is currently difficult to measure small dynamics of molecules in the brain with high spatial and temporal resolution while connecting them to the bigger picture of brain function. A step towards understanding the underlying neural networks of the brain is the ability to sense discrete changes of acetylcholine within a synapse. Here we show an efficient method for generating acetylcholine-detecting nanosensors based on DNA dendrimer scaffolds that incorporate butyrylcholinesterase and fluorescein in a nanoscale arrangement. These nanosensors are selective for acetylcholine and reversibly respond to levels of acetylcholine in the neurophysiological range. This DNA dendrimer architecture has the potential to overcome current obstacles to sensing in the synaptic environment, including the nanoscale size constraints of the synapse and the ability to quantify the spatio-temporal fluctuations of neurotransmitter release. By combining the control of nanosensor architecture with the strategic placement of fluorescent reporters and enzymes, this novel nanosensor platform can facilitate the development of new selective imaging tools for neuroscience.

Tracking protein levels in the body is vital in both research and medicine, where understanding their physiological roles provides insight into their regulation in homeostasis and diseases. In medicine, protein levels are actively sampled since they continuously fluctuate, reflecting the status of biological systems and provide insight into patient health. One such protein is interferon gamma, a clinically relevant protein with immunoregulatory functions that play critical roles against infection. New tools for continuously monitoring protein levels in vivo are invaluable in monitoring real-time conditions of patients to allow better care. Here, we developed a DNA-based nanosensor for the photoacoustic detection of interferon gamma. This work demonstrates how we transformed a simple DNA motif, receptors, and a novel phthalocyanine dye into a proof-of-concept photoacoustic nanosensor for protein detection. Surface plasmon resonance kinetic analysis demonstrated that the nanosensor is responsive and reversible to interferon gamma with an affinity in the nanomolar range, K D1 = 167 nM and K D2 = 316 nM. As a reporter, our design includes a novel phthalocyanine-based photoacoustic dye that stacks in a J-aggregate, causing a 22.5% increase in signal. Upon receptor binding, the DNA structure bends to induce phthalocyanine dye stacking, resulting in a 55% increase in photoacoustic signal in the presence of 10 μM interferon gamma. This proof-of-concept nanosensor is a novel approach to the development of a photoacoustic sensor and may be adapted for other proteins of interest in the future for in vivo tracking.

Silicon photonic (SiP) sensors offer a promising platform for robust and low-cost decentralized diagnostics due to their high scalability, low limit of detection, and ability to integrate multiple sensors for multiplexed analyte detection. Their CMOS-compatible fabrication enables chip-scale miniaturization, high scalability, and low-cost mass production. Sensitive, specific detection with silicon photonic sensors is afforded through biofunctionalization of the sensor surface; consequently, this functionalization chemistry is inextricably linked to sensor performance. In this review, we first highlight the biofunctionalization needs for SiP biosensors, including sensitivity, specificity, cost, shelf-stability, and replicability and establish a set of performance criteria. We then benchmark biofunctionalization strategies for SiP biosensors against these criteria, organizing the review around three key aspects: bioreceptor selection, immobilization strategies, and patterning techniques. First, we evaluate bioreceptors, including antibodies, aptamers, nucleic acid probes, molecularly imprinted polymers, peptides, glycans, and lectins. We then compare adsorption, bioaffinity, and covalent chemistries for immobilizing bioreceptors on SiP surfaces. Finally, we compare biopatterning techniques for spatially controlling and multiplexing the biofunctionalization of SiP sensors, including microcontact printing, pin- and pipette-based spotting, microfluidic patterning in channels, inkjet printing, and microfluidic probes.

Dopamine is widely innervated throughout the brain and critical for many cognitive and motor functions. Imbalances or loss in dopamine transmission underlie various psychiatric disorders and degenerative diseases. Research involving cellular studies and disease states would benefit from a tool for measuring dopamine transmission. Here we show a Quadruplex Integrated DNA (QuID) nanosensor platform for selective and dynamic detection of dopamine. This nanosensor exploits DNA technology and enzyme recognition systems to optically image dopamine levels. The DNA quadruplex architecture is designed to be compatible in physically constrained environments (110 nm) with high flexibility, homogeneity, and a lower detection limit of 110 µM.