Shaping acoustic fields as a toolset for microfluidic manipulations in diagnostic technologies.
ABSTRACT: Ultrasonics offers the possibility of developing sophisticated fluid manipulation tools in lab-on-a-chip technologies. Here we demonstrate the ability to shape ultrasonic fields by using phononic lattices, patterned on a disposable chip, to carry out the complex sequence of fluidic manipulations required to detect the rodent malaria parasite Plasmodium berghei in blood. To illustrate the different tools that are available to us, we used acoustic fields to produce the required rotational vortices that mechanically lyse both the red blood cells and the parasitic cells present in a drop of blood. This procedure was followed by the amplification of parasitic genomic sequences using different acoustic fields and frequencies to heat the sample and perform a real-time PCR amplification. The system does not require the use of lytic reagents nor enrichment steps, making it suitable for further integration into lab-on-a-chip point-of-care devices. This acoustic sample preparation and PCR enables us to detect ca. 30 parasites in a microliter-sized blood sample, which is the same order of magnitude in sensitivity as lab-based PCR tests. Unlike other lab-on-a-chip methods, where the sample moves through channels, here we use our ability to shape the acoustic fields in a frequency-dependent manner to provide different analytical functions. The methods also provide a clear route toward the integration of PCR to detect pathogens in a single handheld system.
Project description:Cell/bead washing is an indispensable sample preparation procedure used in various cell studies and analytical processes. In this article, we report a standing surface acoustic wave (SSAW)-based microfluidic device for cell and bead washing in a continuous flow. In our approach, the acoustic radiation force generated in a SSAW field is utilized to actively extract cells or beads from their original medium. A unique configuration of tilted-angle standing surface acoustic wave (taSSAW) is employed in our device, enabling us to wash beads with >98% recovery rate and >97% washing efficiency. We also demonstrate the functionality of our device by preparing high-purity (>97%) white blood cells from lysed blood samples through cell washing. Our SSAW-based cell/bead washing device has the advantages of label-free manipulation, simplicity, high biocompatibility, high recovery rate, and high washing efficiency. It can be useful for many lab-on-a-chip applications.
Project description:The Schrödinger equation is a fundamental equation to describe the wave function of a quantum-mechanical system. The similar forms between the Schrödinger equation and the paraxial wave equation allow a paradigm shift from the quantum mechanics to classical fields, opening up a plethora of interesting phenomena including the optical super-oscillatory behavior. Here, we propose an ultrasonic meta-lens for generating super-oscillation acoustic wave-packets with different spatial momenta and then superimposing them to a diffraction-limit-broken spot, visually represented by the ring-shaped trapping of tiny particles. Moreover, based on the focused super-oscillation packets, we experimentally verify proof-of-concept super-resolution ultrasound imaging, opening up the arena of super-oscillation ultrasonics for advanced acoustic imaging, biomedical applications, and versatile far-field ultrasound control.
Project description:BACKGROUND: Access to timely and accurate diagnostic tests has a significant impact in the management of diseases of global concern such as malaria. While molecular diagnostics satisfy this need effectively in developed countries, barriers in technology, reagent storage, cost and expertise have hampered the introduction of these methods in developing countries. In this study a simple, lab-on-chip PCR diagnostic was created for malaria that overcomes these challenges. METHODS: The platform consists of a disposable plastic chip and a low-cost, portable, real-time PCR machine. The chip contains a desiccated hydrogel with reagents needed for Plasmodium specific PCR. Chips can be stored at room temperature and used on demand by rehydrating the gel with unprocessed blood, avoiding the need for sample preparation. These chips were run on a custom-built instrument containing a Peltier element for thermal cycling and a laser/camera setup for amplicon detection. RESULTS: This diagnostic was capable of detecting all Plasmodium species with a limit of detection for Plasmodium falciparum of 2 parasites/?L of blood. This exceeds the sensitivity of microscopy, the current standard for diagnosis in the field, by ten to fifty-fold. In a blind panel of 188 patient samples from a hyper-endemic region of malaria transmission in Uganda, the diagnostic had high sensitivity (97.4%) and specificity (93.8%) versus conventional real-time PCR. The test also distinguished the two most prevalent malaria species in mixed infections, P. falciparum and Plasmodium vivax. A second blind panel of 38 patient samples was tested on a streamlined instrument with LED-based excitation, achieving a sensitivity of 96.7% and a specificity of 100%. CONCLUSIONS: These results describe the development of a lab-on-chip PCR diagnostic from initial concept to ready-for-manufacture design. This platform will be useful in front-line malaria diagnosis, elimination programmes, and clinical trials. Furthermore, test chips can be adapted to detect other pathogens for a differential diagnosis in the field. The flexibility, reliability, and robustness of this technology hold much promise for its use as a novel molecular diagnostic platform in developing countries.
Project description:Point-of-care (POC) molecular diagnostics play a crucial role in the prevention and treatment of infectious diseases. It is necessary to develop portable, easy-to-use, inexpensive and rapid molecular diagnostic tools. In this study, we proposed a lab-on-a-chip device that integrated DNA extraction, solid-phase PCR and genotyping detection. The ingenious design of the pneumatic microvalves enabled the fluid mixing and reagent storage to be organically combined, significantly reducing the size of the chip. The solid oligonucleotide array incorporated into the chip allowed the spatial separation of the primers and minimized undesirable interactions in multiplex amplification. As a proof-of-concept for POC molecular diagnostics on the device, five genotypes of high-risk human papillomavirus (HPV) (HPV16/HPV18/HPV31/HPV33/HPV58) were examined. Positive quality control samples and HPV patient cervical swab specimens were analyzed on the integrated microdevice. The platform was capable of detection approximately 50 copies of HPV virus per reaction during a single step, including DNA extraction, solid-phase PCR and genotype detection, in 1 h from samples being added to the chip. This simple and inexpensive microdevice provided great utility for the screening and monitoring of HPV genotypes. The sample-to-result platform will pave the way for wider application of POC molecular testing in the fields of clinical diagnostics, food safety, and environmental monitoring.
Project description:The use of electric fields for signalling and control in liquids is widespread, spanning bioelectric activity in cells to electrical manipulation of microstructures in lab-on-a-chip devices. However, an appropriate tool to resolve the spatio-temporal distribution of electric fields over a large dynamic range has yet to be developed. Here we present a label-free method to image local electric fields in real time and under ambient conditions. Our technique combines the unique gate-variable optical transitions of graphene with a critically coupled planar waveguide platform that enables highly sensitive detection of local electric fields with a voltage sensitivity of a few microvolts, a spatial resolution of tens of micrometres and a frequency response over tens of kilohertz. Our imaging platform enables parallel detection of electric fields over a large field of view and can be tailored to broad applications spanning lab-on-a-chip device engineering to analysis of bioelectric phenomena.
Project description:Directed transport of biological species across the surface of a substrate is essential for realizing lab-on-chip technologies. Approaches that utilize localized magnetic fields to manipulate magnetic particles carrying biological entities are attractive owing to their sensitivity, selectivity, and minimally disruptive impact on biomaterials. Magnetic domain walls in magnetic tracks produce strong localized fields and can be used to capture, transport, and detect individual superparamagnetic microbeads. The dynamics of magnetic microbead transport by domain walls has been well studied. However, demonstration of more complex functions such as selective motion and sorting using continuously driven domain walls in contiguous magnetic tracks is lacking. Here, a junction architecture is introduced that allows for branching networks in which superparamagnetic microbeads can be routed along dynamically-selected paths by a combination of rotating in-plane field for translation, and a pulsed out-of-plane field for path selection. Moreover, experiments and modeling show that the select-field amplitude is bead-size dependent, which allows for digital sorting of multiple bead populations using automated field sequences. This work provides a simple means to implement complex routing networks and selective transport functionalities in chip-based devices using magnetic domain wall conduits.
Project description:A specific double-stranded DNA sensing system is of great interest for diagnostic and other biomedical applications. Zinc finger domains, which recognize double-stranded DNA, can be engineered to form custom DNA-binding proteins for the recognition of specific DNA sequences. As a proof of concept, a sequence-enabled reassembly of a TEM-1 ?-lactamase system (SEER-LAC) was previously demonstrated to develop zinc finger protein (ZFP) arrays for the detection of a double-stranded bacterial DNA sequence. Here, we implemented the SEER-LAC system to demonstrate the direct detection of pathogen-specific DNA sequences present in E. coli O157:H7 on a lab-on-a-chip. ZFPs custom-designed to detect Shiga toxin in E. coli O157:H7 were immobilized on a cyclic olefin copolymer (COC) chip, which can function as a non-PCR based molecular diagnostic device. Pathogen-specific double-stranded DNA was directly detected by using engineered ZFPs immobilized on the COC chip with high specificity, providing a detection limit of 10 fmol of target DNA in a colorimetric assay. Therefore, in this study, we demonstrated the great potential of ZFP arrays on the COC chip for further development of a simple and novel lab-on-a-chip technology for the detection of pathogens.
Project description:Micro-particle operations in many lab-on-a-chip devices require active-type techniques that are accompanied by complex fabrication and operation. The present study describes an alternative method using a passive microfluidic scheme that allows for simpler operation and, therefore, potentially less expensive devices. We present three practical micro-particle operations using our previously developed passive mechanical trap, the asymmetric trap, in a non-acoustic oscillatory flow field. First, we demonstrate size-based segregation of both binary and ternary micro-particle mixtures using size-dependent trap-particle interactions to induce different transport speeds for each particle type. The degree of segregation, yield, and purity of the binary segregations are 0.97?±?0.02, 0.96?±?0.06, and 0.95?±?0.05, respectively. Next, we perform a solution exchange by displacing particles from one solution into another in a trap array. Lastly, we focus and split groups of micro-particles by exploiting the transport polarity of asymmetric traps. These operations can be implemented in any closed fluidic circuit containing asymmetric traps using non-acoustic oscillatory flow, and they open new opportunities to flexibly control micro-particles in integrated lab-on-a-chip platforms with minimal external equipment.
Project description:Two-dimensional acoustofluidic fields in an ultrasonic chamber actuated by segmented ring-shaped vibration sources with different excitation phases are simulated by COMSOL Multiphysics. Diverse acoustic streaming patterns, including aggregation and rotational modes, can be feasibly generated by the excitation of several sessile ultrasonic sources which only vibrate along radial direction. Numerical simulation of particle trajectory driven by acoustic radiation force and streaming-induced drag force also demonstrates that micro-scale particles suspended in the acoustofluidic chamber can be trapped in the velocity potential well of fluid flow or can rotate around the cavity center with the circumferential acoustic streaming field. Preliminary investigation of simple Russian doll- or Matryoshka-type configurations (double-layer vibration sources) provide a novel method of multifarious structure design in future researches on the combination of phononic crystals and acoustic streaming fields. The implementation of multiple segmented ring-shaped vibration sources offers flexibility for the control of acoustic streaming fields in microfluidic devices for various applications. We believe that this kind of acoustofluidic design is expected to be a promising tool for the investigation of rapid microfluidic mixing on a chip and contactless rotational manipulation of biosamples, such as cells or nematodes.
Project description:Portable, low-cost, and quantitative nucleic acid detection is desirable for point-of-care diagnostics; however, current polymerase chain reaction testing often requires time-consuming multiple steps and costly equipment. We report an integrated microfluidic diagnostic device capable of on-site quantitative nucleic acid detection directly from the blood without separate sample preparation steps. First, we prepatterned the amplification initiator [magnesium acetate (MgOAc)] on the chip to enable digital nucleic acid amplification. Second, a simplified sample preparation step is demonstrated, where the plasma is separated autonomously into 224 microwells (100 nl per well) without any hemolysis. Furthermore, self-powered microfluidic pumping without any external pumps, controllers, or power sources is accomplished by an integrated vacuum battery on the chip. This simple chip allows rapid quantitative digital nucleic acid detection directly from human blood samples (10 to 10<sup>5</sup> copies of methicillin-resistant <i>Staphylococcus aureus</i> DNA per microliter, ~30 min, via isothermal recombinase polymerase amplification). These autonomous, portable, lab-on-chip technologies provide promising foundations for future low-cost molecular diagnostic assays.