Smartphone-Imaged HIV-1 Reverse-Transcription Loop-Mediated Isothermal Ampliflcation(RT-LAMP) on a Chip from Whole Blood

Viral load measurements are an essential tool for the long-term clinical care of human immunodeficiency virus （HIV）-positive individuals. The gold standards in viral load instrumentation, however, are still too limited by their size, cost, and sophisticated operation for these measurements to be ubiquitous in remote settings with poor healthcare infrastructure, including parts of the world that are disproportionately affected by HIV infection. The challenge of developing a point-of-care platform capable of making viral load more accessible has been frequently approached but no solution has yet emerged that meets the practical requirements of low cost, portability, and ease-of-use. In this paper, we perform reverse-transcription loop-mediated isothermal amplification （RT-LAMP） on minimally processed HIV-spiked whole blood samples with a microfluidic and silicon microchip platform, and perform fluorescence measurements with a consumer smartphone. Our integrated assay shows amplification from as few as three viruses in a - 60 nL RT- LAMP droplet, corresponding to a whole blood concentration of 670 viruses per μL of whole blood. The technology contains greater power in a digital RT-LAMP approach that could be scaled up for the determination of viral load from a finger prick of blood in the clinical care of HIV-positive individuals. We demonstrate that all aspects of this viral load approach, from a drop of blood to imaging the RT-LAMP reaction, are compatible with lab-on-a-chip components and mobile instrumentation.

Efficient and wideband acousto-optic modulation on thin-film lithium niobate for microwave-to-photonic conversion

Microwave photonics,a field that crosscuts microwave/millimeter-wave engineering with optoelectronics,has sparked great interest from research and commercial sectors. This multidisciplinary fusion can achieve ultrawide bandwidth and ultrafast speed that were considered impossible in conventional chip-scale microwave/millimeterwave systems. Conventional microwave-to-photonic converters,based on resonant acousto-optic modulation,produce highly efficient modulation but sacrifice bandwidth and limit their applicability for most real-world microwave signal-processing applications. In this paper,we build highly efficient and wideband microwaveto-photonic modulators using the acousto-optic effect on suspended lithium niobate thin films. A wideband microwave signal is first piezoelectrically transduced using interdigitated electrodes into Lamb acoustic waves,which directly propagates across an optical waveguide and causes refractive index pertp urbation through the photoelastic effect. This approach is power-efficient,with phase shifts up to 0.0166 rad∕m W over a 45μm modulation length and with a bandwidth up to 140 MHz at a center frequency of 1.9 GHz. Compared to the state-ofthe-art,a 9×more efficient modulation has been achieved by optimizing the acoustic and optical modes and their interactions.

Self-assembled microtubular electrodes for on-chip low-voltage electrophoretic manipulation of charged particles and macromolecules

On-chip manipulation of charged particles using electrophoresis or electroosmosis is widely used for many applications,including optofluidic sensing,bioanalysis and macromolecular data storage.We hereby demonstrate a technique for the capture,localization,and release of charged particles and DNA molecules in an aqueous solution using tubular structures enabled by a strain-induced self-rolled-up nanomembrane(S-RuM)platform.Cuffed-in 3D electrodes that are embedded in cylindrical S-RuM structures and biased by a constant DC voltage are used to provide a uniform electrical field inside the microtubular devices.Efficient charged-particle manipulation is achieved at a bias voltage of<2-4 V,which is~3 orders of magnitude lower than the required potential in traditional DC electrophoretic devices.Furthermore,Poisson-Boltzmann multiphysics simulation validates the feasibility and advantage of our microtubular charge manipulation devices over planar and other 3D variations of microfluidic devices.This work lays the foundation for on-chip DNA manipulation for data storage applications.

On the design and fabrication of nanoliter-volume hanging drop networks

Hanging drop cultures provide a favorable environment for the gentle,gel-free formation of highly uniform three-dimensional cell cultures often used in drug screening applications.Initial cell numbers can be limited,as with primary cells provided by minimally invasive biopsies.Therefore,it can be beneficial to divide cells into miniaturized arrays of hanging drops to supply a larger number of samples.Here,we present a framework for the miniaturization of hanging drop networks to nanoliter volumes.The principles of a single hanging drop are described and used to construct the fundamental equations for a microfluidic system composed of multiple connected drops.Constitutive equations for the hanging drop as a nonlinear capacitive element are derived for application in the electronic-hydraulic analogy,forming the basis for more complex,time-dependent numerical modeling of hanging drop networks.This is supplemented by traditional computational fluid dynamics simulation to provide further information about flow conditions within the wells.A fabrication protocol is presented and demonstrated for creating transparent,microscale arrays of pinned hanging drops.A custom interface,pressure-based fluidic system,and environmental chamber have been developed to support the device.Finally,fluid flow on the chip is demonstrated to align with expected behavior based on the principles derived for hanging drop networks.Challenges with the system and potential areas for improvement are discussed.This paper expands on the limited body of hanging drop network literature and provides a framework for designing,fabricating,and operating these systems at the microscale.

Label-free SARS-CoV-2 detection and classification using phase imaging with computational specificity

Efforts to mitigate the COVID-19 crisis revealed that fast,accurate,and scalable testing is crucial for curbing the current impact and that of future pandemics.We propose an optical method for directly imaging unlabeled viral particles and using deep learning for detection and classification.An ultrasensitive interferometric method was used to image four virus types with nanoscale optical path-length sensitivity.Pairing these data with fluorescence images for ground truth,we trained semantic segmentation models based on U-Net,a particular type of convolutional neural network.The trained network was applied to classify the viruses from the interferometric images only,containing simultaneously SARS-CoV-2,H1N1(influenza-A virus),HAdV(adenovirus),and ZIKV(Zika virus).Remarkably,due to the nanoscale sensitivity in the input data,the neural network was able to identify SARS-CoV-2 vs.the other viruses with 96%accuracy.The inference time for each image is 60 ms,on a common graphic-processing unit.This approach of directly imaging unlabeled viral particles may provide an extremely fast test,of less than a minute per patient.As the imaging instrument operates on regular glass slides,we envision this method as potentially testing on patient breath condensates.The necessary high throughput can be achieved by translating concepts from digital pathology,where a microscope can scan hundreds of slides automatically.

Quantitative analysis of focal adhesion dynamics using photonic resonator outcoupler microscopy (PROM)

Focal adhesions are critical cell membrane components that regulate adhesion and migration and have cluster dimensions that correlate closely with adhesion engagement and migration speed.We utilized a label-free approach for dynamic,long-term,quantitative imaging of cell–surface interactions called photonic resonator outcoupler microscopy(PROM)in which membrane-associated protein aggregates outcoupled photons from the resonant evanescent field of a photonic crystal biosensor,resulting in a highly localized reduction of the reflected light intensity.By mapping the changes in the resonant reflected peak intensity from the biosensor surface,we demonstrate the ability of PROM to detect focal adhesion dimensions.Similar spatial distributions can be observed between PROM images and fluorescence-labeled images of focal adhesion areas in dental epithelial stem cells.In particular,we demonstrate that cell–surface contacts and focal adhesion formation can be imaged by two orthogonal label-free modalities in PROM simultaneously,providing a general-purpose tool for kinetic,high axial-resolution monitoring of cell interactions with basement membranes.

Multiaxially-stretchable kirigami-patterned mesh design for graphene sensor devices

In wearable electronics,significant research has gone into imparting stretchability and flexibility to otherwise rigid electronic components while maintaining their electrical properties.Thus far,this has been achieved through various geometric modifications of the rigid conductive components themselves,such as with microcracked,buckled,or planar meander structures.Additionally,strategic placement of these resulting components within the overall devices,such as embedding them at the neutral plane,has been found to further enhance mechanical stability under deformation.However,these strategies are still limited in performance,failing to achieve fully strain-insensitive electrical performance under biaxial stretching,twisting,and mixed strain states.Here,we developed a new platform for wearable,motion artifact-free sensors using a graphene-based multiaxially stretchable kirigami-patterned mesh structure.The normalized resistance change of the electrodes and graphene embedded in the structure is smaller than 0.5%and 0.23%under 180°torsion and 100%biaxial strain,respectively.Moreover,the resistance change is limited to 5%under repeated stretching-releasing cycles from 0%to 100%biaxial strain.In addition,we investigated the deformation mechanisms of the structure with finite element analysis.Based on the simulation results,we derived a dimensionless geometric parameter that enables prediction of stretchability of the structure with high accuracy.Lastly,as a proof-of-concept,we demonstrated a biaxially-stretchable graphene-based sensor array capable of monitoring of temperature and glucose level with minimized motion-artifacts.

A 3D-printed platform for modular neuromuscular motor units

A complex and functional living cellular system requires the interaction of one or more cell types to perform specific tasks,such as sensing,processing,or force production.Modular and flexible platforms for fabrication of such multi-cellular modules and their characterization have been lacking.Here,we present a modular cellular system,made up of multi-layered tissue rings containing integrated skeletal muscle and motor neurons(MNs)embedded in an extracellular matrix.The MNs were differentiated from mouse embryonic stem cells through the formation of embryoid bodies(EBs),which are spherical aggregations of cells grown in a suspension culture.The EBs were integrated into a tissue ring with skeletal muscle,which was differentiated in parallel,to create a co-culture amenable to both cell types.The multi-layered rings were then sequentially placed on a stationary three-dimensionalprinted hydrogel structure resembling an anatomical muscle–tendon–bone organization.We demonstrate that the site-specific innervation of a group of muscle fibers in the multi-layered tissue rings allows for muscle contraction via chemical stimulation of MNs with glutamate,a major excitatory neurotransmitter in the mammalian nervous system,with the frequency of contraction increasing with glutamate concentration.The addition of tubocurarine chloride(a nicotinic receptor antagonist)halted the contractions,indicating that muscle contraction was MN induced.With a bio-fabricated system permitting controllable mechanical and geometric attributes in a range of length scales,our novel engineered cellular system can be utilized for easier integration of other modular“building blocks”in living cellular and biological machines.