The most important organ in the human body is unquestionably the brain. Yet, despite its importance, it is one of the least well understood organs. One reason for this lack of understanding of the brain is the lack of data available to researchers from in vivo studies. Historically, collecting measurements from the brain has been difficult due to the high risk to the patient. Recently technology has been developed to allow electrical measurements to be taken from the brain directly, however most systems involve non-permanent sensors because of the requirement for transcranial wiring for power and data. Developments in the field of CMOS circuit design, wireless power transmission, and wireless data transmission have enabled the creation of implantable neural recording devices as a combination of these technologies. The implant designed in this paper is ~15 mm in diameter and 2 mm at its thickest point on a flexible polyamide PCB. The flexible nature of the implant allows for the implant to conform to the surface of the brain. The implant requires no transcranial orifice since it is powered wirelessly and transmits data wirelessly via Bluetooth low energy. The CMOS neural amplifier chip on the implant utilizes an enhanced form of delta modulation to remove the requirement for large ADCs to be present on the die, saving space and enabling 1024 amplifiers and electrodes to be present on the chip. The implant is capable of measuring, modulating, and wirelessly transmitting a millivolt order signal to a PC for demodulation and analysis.
Large-scale microelectrode arrays offers enhanced spatiotemporal resolution in electrophysiology studies. In this paper, we discuss the design and performance of an electrochemical detector array which is capable of 1024-ch parallel cyclic voltammetry (CV) as well as other electrochemical measurements. The electrochemical detector is fabricated using a custom-designed CMOS chip which integrates both the circuity and on-chip microelectrode array, to operate and record from electrochemical measurements. For parallel 1024-ch recordings, 1024 capacitor-based integrating transimpedance amplifiers (TIA) are designed and integrated. The TIA design features the bipolar capabilities for measuring both negative and positive electrochemical currents due to reduction and oxidation of molecules. The resulted dynamic range of this TIA is -700 pA to 1968 pA. CV can be used to examine the quality of electrochemical electrodes by measuring the double-layer capacitance. Double-layer capacitance forms at the electrode-electrolyte interface and is a function of the effective area of the electrode. Thus, a contaminated electrode can have smaller effective area resulting in smaller double-layer capacitance. Using the parallel CV capability of the monolithic CMOS device, the double layer capacitance of all 1024 electrodes are simultaneously measured to examine the status of the electrode surface in real time. The initial measurement of the electrode array showed a mean capacitance of 466 pF. After plasma treatment to remove contamination on the electrode surface, the increased capacitance was 1.36 nF nearly tripling the effective surface area. We have successfully developed of 1024-ch electrochemical detector array using the monolithic CMOS sensor. The CV functionality was validated by measuring the double-layer capacitance of the on-chip electrode array. This method can accelerate the characterization of a massive electrode array before analytical experiments to provide well-controlled electrochemical electrodes, which is crucial in conducting reliable electrochemical measurements.
Over the last decade, nucleic acid amplification tests (NAATs) including polymerase chain reaction (PCR) were an indispensable methodology for diagnosing cancers, viral and bacterial infections owing to their high sensitivity and specificity. Because the NAATs can recognize and discriminate even a few copies of nucleic acid (NA) and species-specific NA sequences, NAATs have become the gold standard in a wide range of applications. However, limitations of NAAT approaches have recently become more apparent by reason of their lengthy run time, large reaction volume, and complex protocol. To meet the current demands of clinicians and biomedical researchers, new NAATs have developed to achieve ultrafast sample-to-answer protocols for the point-of-care testing (POCT). In this review, ultrafast NA-POCT platforms are discussed, outlining their NA amplification principles as well as delineating recent advances in ultrafast NAAT applications. The main focus is to provide an overview of NA-POCT platforms in regard to sample preparation of NA, NA amplification, NA detection process, interpretation of the analysis, and evaluation of the platform design. Increasing importance will be given to innovative, ultrafast amplification methods and tools which incorporate artificial intelligence (AI)-associated data analysis processes and mobile-healthcare networks. The future prospects of NA POCT platforms are promising as they allow absolute quantitation of NA in individuals which is essential to precision medicine.
We review some emerging trends in transduction, connectivity and data analytics for Point-of-Care Testing (POCT) of infectious and non-communicable diseases. The patient need for POCT is described along with developments in portable diagnostics, specifically in respect of Lab-on-chip and microfluidic systems. We describe some novel electrochemical and photonic systems and the use of mobile phones in terms of hardware components and device connectivity for POCT. Developments in data analytics that are applicable for POCT are described with an overview of data structures and recent AI/Machine learning trends. The most important methodologies of machine learning, including deep learning methods, are summarised. The potential value of trends within POCT systems for clinical diagnostics within Lower Middle Income Countries (LMICs) and the Least Developed Countries (LDCs) are highlighted.
High-throughput recordings of small current are becoming more common in biosensor applications, including in vivo dopamine measurements, single-cell electrophysiology, photoplethysmography, pulse oximetry, and nanopore recordings. Thus, a highly scalable transimpedance amplifier design is in demand. Half-Shared amplifier design is one way to improve the scalability by sharing the non-inverting side of the operational amplifier design for many inverting halves. This method reduces silicon area and power by nearly half compared to using independent operational amplifiers. In this paper, we analyze the scalability of a simple half-shared amplifier structure while investigating the trade-off of increasing the number of inverting half amplifiers sharing a single non-inverting half. A transimpedance amplifier is designed using the half-shared structure to minimize the size per amplifier. The transimpedance amplifier is based on a current integration of a capacitor. The noise analysis of the integration amplifier is a challenging task because it does not reach a steady-state, thus, being a non-stationary circuit. For frequency analysis, a conversion method is discussed to estimate the noise characteristic in the simulation. The array design of 1024 transimpedance amplifiers is fabricated using a standard 0.35-μm process and is tested to confirm the validity of above analysis. The amplifier array exhibits high linearity in transimpedance gain (7.00 mV/pA for high-gain and 0.86 mV/pA for low-gain), low mismatch of 1.65 mV across the entire 1024 amplifier array, and extremely low noise. The technique will be crucial in enabling the fabrication of larger arrays to enable higher-throughput measurement tools for biosensor applications.
Human neuroblastoma cells, SH-SY5Y, are often used as a neuronal model to study Parkinson's disease and dopamine release in the substantia nigra, a midbrain region that plays an important role in motor control. Using amperometric single-cell recordings of single vesicle release events, we can study molecular manipulations of dopamine release and gain a better understanding of the mechanisms of neurological diseases. However, single-cell analysis of neurotransmitter release using traditional techniques yields results with very low throughput. In this paper, we will discuss a monolithically-integrated CMOS sensor array that has the low-noise performance, fine temporal resolution, and 1024 parallel channels to observe dopamine release from many single cells with single-vesicle resolution. The measured noise levels of our transimpedance amplifier are 415, 622, and 1083 fARMS , at sampling rates of 10, 20, and 30 kS/s, respectively, without additional filtering. Post-CMOS processing is used to monolithically integrate 1024 on-chip gold electrodes, with an individual electrode size of 15 μm × 15 μm, directly on 1024 transimpedance amplifiers in the CMOS device. SU-8 traps are fabricated on individual electrodes to allow single cells to be interrogated and to reject multicellular clumps. Dopamine secretions from 76 cells are simultaneously recorded by loading the CMOS device with SH-SY5Y cells. In the 42-second measurement, a total of 7147 single vesicle release events are monitored. The study shows the CMOS device's capability of recording vesicle secretion at a single-cell level, with 1024 parallel channels, to provide detailed information on the dynamics of dopamine release at a single-vesicle resolution.
Sample preparation is an essential process that precedes nucleic acid detections which use quantitative polymerase chain reaction (qPCR). However, sample preparation is a labor-intensive process and requires skilled labor, thus limiting the public's access in low-resource settings to many high-quality nucleic acid-based detection mechanisms. In this paper, we present a simple, handheld, battery-operated sample preparation device to minimize user's involvement. The device uses a simple pouring method to process the DNA sample without pipetting or using disposable pipette tips. The developed device has a size of 12 × 8 × 8 cm3 and mass of only 364 g. The device is compared to gold standard methods, including magnetic bead-based and silica filter-based DNA extractions. For a short segment DNA target of 68 bp, the presented device captured 8.67× more DNA compared to that of the manual magnetic bead-based method. Because of automation, the measured capture efficiency is more consistent and has a smaller deviation between multiple repetitions than the manual method. To present a comprehensive, portable, battery-operated diagnostic system, the sample preparation device is tested in conjunction with a 3D-manufactured qPCR device. The test using three diluted target DNA samples, each spiked in whole blood (1×, 0.1×, and 0.01×), revealed a quantitative detection with ideal cycle threshold separations between the measurements. The combination of two devices will aid in resource-limited settings to promptly and accurately diagnose infections of patients.
Fast electrochemical imaging enables the dynamic study of electroactive molecule diffusion in neurotransmitter release from single cells and dopamine mapping in brain slices. In this paper, we discuss the design of an electrochemical imaging sensor using a monolithic complementary metal-oxide-semiconductor (CMOS) sensor array and a multifunctional data acquisition system. Using post-CMOS fabrication, the CMOS sensor integrates 1024 on-chip electrodes on the surface and contains 1024 low-noise amplifiers to simultaneous process parallel electrochemical recordings. Each electrochemical electrode and amplifier is optimized to operate at 10.38-kHz sampling rate. To support the operation of the high-throughput CMOS device, a multifunctional data acquisition device is developed to provide the required speed and accuracy. The high analog data rate of 10.63 MHz from all 1024 amplifiers is redundantly sampled by the custom-designed data acquisition system which can process up to 73.6 MHz with up to ~400 Mbytes/s data rate to a computer using universal serial bus 3.0 interface. To contain the liquid above the electrochemical sensors and prevent electronic and wire damage, we packaged the monolithic sensor using a 3-D printed well. Using the presented device, 32 pixel × 32 pixel electrochemical imaging of dopamine diffusion is successfully demonstrated at over 10,000 frames per second, the fastest reported to date.
Diagnosing infectious diseases using quantitative polymerase chain reaction (qPCR) offers a conclusive result in determining the infection, the strain or type of pathogen, and the level of infection. However, due to the high-cost instrumentation involved and the complexity in maintenance, it is rarely used in the field to make a quick turnaround diagnosis. In order to provide a higher level of accessibility than current qPCR devices, a set of 3D manufacturing methods is explored as a possible option to fabricate a low-cost and portable qPCR device. The key advantage of this approach is the ability to upload the digital format of the design files on the internet for wide distribution so that people at any location can simply download and feed into their 3D printers for quick manufacturing. The material and design are carefully selected to minimize the number of custom parts that depend on advanced manufacturing processes which lower accessibility. The presented 3D manufactured qPCR device is tested with 20-μL samples that contain various concentrations of lentivirus, the same type as HIV. A reverse-transcription step is a part of the device’s operation, which takes place prior to the qPCR step to reverse transcribe the target RNA from the lentivirus into complementary DNA (cDNA). This is immediately followed by qPCR which quantifies the target sequence molecules in the sample during the PCR amplification process. The entire process of thermal control and time-coordinated fluorescence reading is automated by closed-loop feedback and a microcontroller. The resulting device is portable and battery-operated, with a size of 12 × 7 × 6 cm3 and mass of only 214 g. By uploading and sharing the design files online, the presented low-cost qPCR device may provide easier access to a robust diagnosis protocol for various infectious diseases, such as HIV and malaria.
The resemblance of lipid membrane models to physiological membranes determines how well molecular dynamics (MD) simulations imitate the dynamic behavior of cell membranes and membrane proteins. Physiological lipid membranes are composed of multiple types of phospholipids, and the leaflet compositions are generally asymmetric. Here we describe an approach for self-assembly of a Coarse-Grained (CG) membrane model with physiological composition and leaflet asymmetry using the MARTINI force field. An initial set-up of two boxes with different types of lipids according to the leaflet asymmetry of mammalian cell membranes stacked with 0.5 nm overlap, reliably resulted in the self-assembly of bilayer membranes with leaflet asymmetry resembling that of physiological mammalian cell membranes. Self-assembly in the presence of a fragment of the plasma membrane protein syntaxin 1A led to spontaneous specific positioning of phosphatidylionositol(4,5)bisphosphate at a positively charged stretch of syntaxin consistent with experimental data. An analogous approach choosing an initial set-up with two concentric shells filled with different lipid types results in successful assembly of a spherical vesicle with asymmetric leaflet composition. Self-assembly of the vesicle in the presence of the synaptic vesicle protein synaptobrevin 2 revealed the correct position of the synaptobrevin transmembrane domain. This is the first CG MD method to form a membrane with physiological lipid composition as well as leaflet asymmetry by self-assembly and will enable unbiased studies of the incorporation and dynamics of membrane proteins in more realistic CG membrane models.
SNAP-25 is a Q-SNARE protein mediating exocytosis of neurosecretory vesicles including chromaffin granules. Previous results with a SNAP-25 construct lacking the nine C terminal residues (SNAP-25Δ9) showed changed fusion pore properties (Fang et al., 2008), suggesting a model for fusion pore mechanics that couple C terminal zipping of the SNARE complex to the opening of the fusion pore. The deleted fragment contains the positively charged residues R198 and K201, adjacent to layers 7 and 8 of the SNARE complex. To determine how fusion pore conductance and dynamics depend on these residues, single exocytotic events in bovine chromaffin cells expressing R198Q, R198E, K201Q, or K201E mutants were investigated by carbon fiber amperometry and cell-attached patch capacitance measurements. Coarse grain molecular dynamics simulations revealed spontaneous transitions between a loose and tightly zippered state at the SNARE complex C terminus. The SNAP-25 K201Q mutant showed no changes compared with SNAP-25 wild-type. However, K201E, R198Q, and R198E displayed reduced release frequencies, slower release kinetics, and prolonged fusion pore duration that were correlated with reduced probability to engage in the tightly zippered state. The results show that the positively charged amino acids at the SNAP-25 C terminus promote tight SNARE complex zippering and are required for high release frequency and rapid release in individual fusion events.
Neurotransmitter release is modulated by many drugs and molecular manipulations. We present an active CMOS-based electrochemical biosensor array with high throughput capability (100 electrodes) for on-chip amperometric measurement of neurotransmitter release. The high-throughput of the biosensor array will accelerate the data collection needed to determine statistical significance of changes produced under varying conditions, from several weeks to a few hours. The biosensor is designed and fabricated using a combination of CMOS integrated circuit (IC) technology and a photolithography process to incorporate platinum working electrodes on-chip. We demonstrate the operation of an electrode array with integrated high-gain potentiostats and output time-division multiplexing with minimum dead time for readout. The on-chip working electrodes are patterned by conformal deposition of Pt and lift-off photolithography. The conformal deposition method protects the underlying electronic circuits from contact with the electrolyte that covers the electrode array during measurement. The biosensor was validated by simultaneous measurement of amperometric currents from 100 electrodes in response to dopamine injection, which revealed the time course of dopamine diffusion along the surface of the biosensor array. The biosensor simultaneously recorded neurotransmitter release successfully from multiple individual living chromaffin cells. The biosensor was capable of resolving small and fast amperometric spikes reporting release from individual vesicle secretions. We anticipate that this device will accelerate the characterization of the modulation of neurotransmitter secretion from neuronal and endocrine cells by pharmacological and molecular manipulations of the cells.
We have developed and tested transparent microelectrode arrays capable of simultaneous amperometric measurement of oxidizable molecules and fluorescence imaging through the electrodes. Surface patterned microelectrodes were fabricated from three different conducting materials: Indium-tin-oxide (ITO), nitrogen-doped diamond-like carbon (DLC) deposited on top of ITO, or very thin (12-17 nm) gold films on glass substrates. Chromaffin cells loaded with lysotracker green or acridine orange dye were placed atop the electrodes and vesicle fluorescence imaged with total internal reflection fluorescence (TIRF) microscopy while catecholamine release from single vesicles was measured as amperometric spikes with the surface patterned electrodes. Electrodes fabricated from all three materials were capable of detecting amperometric signals with high resolution. Unexpectedly, amperometric spikes recorded with ITO electrodes had only about half the amplitude and about half as much charge as those detected with DLC or gold electrodes, indicating that the ITO electrodes are not as sensitive as gold or DLC electrodes for measurement of quantal catecholamine release. The lower sensitivity of ITO electrodes was confirmed by chronoamperometry measurements comparing the currents in the presence of different analytes with the different electrode materials.
Microelectrodes based on poly(3,4‐ethylenedioxythiophene) doped with poly(styrene sulfonate) (PEDOT:PSS) are able to record the oxidation of catecholamines released from chromaffin cells during exocytosis with a high signal‐to‐noise ratio. This result represents a new capability for organic electronics that could lead to devices that interface with the nervous system in novel ways.