Dr. Warkiani is a Senior Lecturer in the School of Biomedical Engineering, UTS, Sydney, Australia. He received his Ph.D. in Mechanical Engineering from Nanyang Technological University (NTU, Singapore), and undertook postdoctoral training at Massachusetts Institute of Technology (MIT, USA). He is also a member of Institute for Biomedical Materials & Devices (IBMD) and Center for Health Technologies (CHT) at UTS, a visiting scientist at the Garvan Institute for Biomedical Research as well as Translational Cancer Research Network (TCRN).
Dr Warkiani’s current research activities focus on three key areas of (i) Microfluidics involving the design and development of novel microfluidic systems for particle and cell sorting (e.g., circulating tumor cells, fetal cells & stem cells) for diagnostic and therapeutic applications, (ii) Bio-MEMS involving the fabrication and characterization of novel 3D lab-on-a-chip systems to model physiological functions of tissues and organs, and (iii) 3D Printing involving the design and development of novel miniaturized systems (e.g., micromixers, micro-cyclones) for basic and applied research.
- Nanyang Outstanding Young Alumni Award (2016).
- MIT TR35 young investigator, Asia-Pacific Region (2016).
- Fresh Science Program, Australia (2015).
- Students’ Design Gold Award, Biomedical Engineering Society, Singapore, (2014).
- SMART prestigious fellowship (2013).
- Best student paper award at AMN-APLOC 2011, 5-7 January, Singapore.
- Best research proposal award at NanoMemCourse 2010 at Netherlands.
- A*STAR prestigious scholarship for PhD study at NTU (2009)
For more information, please visit my group homepage: http://www.warkianilab.com
Can supervise: YES
Introduction to BioMEMS
Engineering Biomedical Polymer
Microfluidic cell-separation technologies have been studied for almost two decades, but the limited throughput has restricted their impact and range of application. Recent advances in microfluidics enable high-throughput cell sorting and separation, and this has led to various novel diagnostic and therapeutic applications that previously had been impossible to implement using microfluidics technologies. In this review, we focus on recent progress made in engineering large-volume microfluidic cell-sorting methods and the new applications enabled by them.
Abbasnejad, B., Thorby, W., Razmjou, A., Jin, D., Asadnia, M. & Ebrahimi Warkiani, M. 2018, 'MEMS piezoresistive flow sensors for sleep apnea therapy', Sensors and Actuators, A: Physical, vol. 279, pp. 577-585.View/Download from: Publisher's site
© 2018 Elsevier B.V. A MEMS liquid crystal polymer (LCP), used in the membrane-based pressure sensor, has been found highly useful as a flow sensor. Here we conducted a set of elaborate experiments using an air flow generator to investigate the potential of our LCP flow sensor for sleep apnea therapy. Critical properties of the LCP flow sensor, including flow range, resolution (sensitivity), accuracy, and response time, have been systematically characterized. As a result, LCP flow sensor achieves a limit of detection of 8 LPM to measure flow rate, better than the commercial flow sensor (>10 LPM). Our LCP flow sensor shows a favourable response in a large flow range (8–160 LPM) with a sensitivity of detecting a linear voltage response of 0.004 V per 1 LPM flow rate. With minimum detectable flow, high sensitivity and resolution, we further demonstrated our LCP flow sensor for detecting human respiration. Moreover, using a two- dimensional simulation in COMSOL Multiphysics, we demonstrated the deformation of LCP membrane in response to different flow velocities which leads to resistance change in sensor's strain gauge.
Ghorbani, S., Eyni, H., Tiraihi, T., Salari Asl, L., Soleimani, M., Atashi, A., Pour Beiranvand, S. & Ebrahimi Warkiani, M. 2018, 'Combined effects of 3D bone marrow stem cell-seeded wet-electrospun poly lactic acid scaffolds on full-thickness skin wound healing', International Journal of Polymeric Materials and Polymeric Biomaterials, pp. 1-8.View/Download from: Publisher's site
© 2017 Taylor & Francis Tissue engineering has emerged as an alternative treatment to traditional grafts for skin wound healing. Three-dimensional nanofibers have been used extensively for this purpose due to their excellent biomedical-related properties. In this study, high porous 3D poly lactic acid nanofibrous scaffolds (PLA-S) were prepared by wet-electrospinning technique and seeded with rat bone-marrow stem cells (BMSCs) to characterize the biocompatibility and therapeutic efficacy of these fibers on the treating full-thickness dermal wounds. The results of in vitro andin vivo studies indicate that the 3D fibrous PLA-S can be a potential wound dressing for wound repair, particularly when seeded with BMSCs.
Hassanzadeh-Barforoushi, A., Law, A.M.K., Hejri, A., Asadnia, M., Ormandy, C.J., Gallego-Ortega, D. & Ebrahimi Warkiani, M. 2018, 'Static droplet array for culturing single live adherent cells in an isolated chemical microenvironment.', Lab on a chip, vol. 18, no. 15, pp. 2156-2166.View/Download from: Publisher's site
We present here a new method to easily and reliably generate an array of hundreds of dispersed nanoliter-volume semi-droplets for single-cells culture and analysis. The liquid segmentation step occurs directly in indexed traps by a tweezer-like mechanism and is stabilized by spatial confinement. Unlike common droplet-based techniques, the semi-droplet wets its surrounding trap walls thus supporting the culturing of both adherent and non-adherent cells. To eliminate cross-droplet cell migration and chemical cross-talk each semi-droplet is separated from a nearby trap by an 80 pL air plug. The overall setup and injection procedure takes less than 10 minutes, is insensitive to fabrication defects and supports cell recovery for downstream analysis. The method offers a new approach to easily capture, image and culture single cells in a chemically isolated microenvironment as a preliminary step towards high-throughput single-cell assays.
Khan, H., Razmjou, A., Ebrahimi Warkiani, M., Kottapalli, A. & Asadnia, M. 2018, 'Sensitive and Flexible Polymeric Strain Sensor for Accurate Human Motion Monitoring.', Sensors (Basel, Switzerland), vol. 18, no. 2.View/Download from: Publisher's site
Flexible electronic devices offer the capability to integrate and adapt with human body. These devices are mountable on surfaces with various shapes, which allow us to attach them to clothes or directly onto the body. This paper suggests a facile fabrication strategy via electrospinning to develop a stretchable, and sensitive poly (vinylidene fluoride) nanofibrous strain sensor for human motion monitoring. A complete characterization on the single PVDF nano fiber has been performed. The charge generated by PVDF electrospun strain sensor changes was employed as a parameter to control the finger motion of the robotic arm. As a proof of concept, we developed a smart glove with five sensors integrated into it to detect the fingers motion and transfer it to a robotic hand. Our results shows that the proposed strain sensors are able to detect tiny motion of fingers and successfully run the robotic hand.
Moloudi, R., Oh, S., Yang, C., Ebrahimi Warkiani, M. & Naing, M.W. 2018, 'Inertial particle focusing dynamics in a trapezoidal straight microchannel: application to particle filtration', Microfluidics and Nanofluidics, vol. 22, no. 3.View/Download from: Publisher's site
© 2018, Springer-Verlag GmbH Germany, part of Springer Nature. Inertial microfluidics has emerged recently as a promising tool for high-throughput manipulation of particles and cells for a wide range of flow cytometric tasks including cell separation/filtration, cell counting, and mechanical phenotyping. Inertial focusing is profoundly reliant on the cross-sectional shape of channel and its impacts on not only the shear field but also the wall-effect lift force near the wall region. In this study, particle focusing dynamics inside trapezoidal straight microchannels was first studied systematically for a broad range of channel Re number (20 OpenSPiltSPi Re OpenSPiltSPi 800). The altered axial velocity profile and consequently new shear force arrangement led to a cross-lateral movement of equilibration toward the longer side wall when the rectangular straight channel was changed to a trapezoid; however, the lateral focusing started to move backward toward the middle and the shorter side wall, depending on particle clogging ratio, channel aspect ratio, and slope of slanted wall, as the channel Reynolds number further increased (Re CloseSPigtSPi 50). Remarkably, an almost complete transition of major focusing from the longer side wall to the shorter side wall was found for large-sized particles of clogging ratio K ~ 0.9 (K = a/Hmin) when Re increased noticeably to ~ 650. Finally, based on our findings, a trapezoidal straight channel along with a bifurcation was designed and applied for continuous filtration of a broad range of particle size (0.3 OpenSPiltSPi K OpenSPiltSPi 1) exiting through the longer wall outlet with ~ 99% efficiency (Re OpenSPiltSPi 100).
Moshksayan, K., Kashaninejad, N., Warkiani, M.E., Lock, J.G., Moghadas, H., Firoozabadi, B., Saidi, M.S. & Nguyen, N.T. 2018, 'Spheroids-on-a-chip: Recent advances and design considerations in microfluidic platforms for spheroid formation and culture', Sensors and Actuators, B: Chemical, vol. 263, pp. 151-176.View/Download from: Publisher's site
© 2018 Elsevier B.V. A cell spheroid is a three-dimensional (3D) aggregation of cells. Synthetic, in-vitro spheroids provide similar metabolism, proliferation, and species concentration gradients to those found in-vivo. For instance, cancer cell spheroids have been demonstrated to mimic in-vivo tumor microenvironments, and are thus suitable for in-vitro drug screening. The first part of this paper discusses the latest microfluidic designs for spheroid formation and culture, comparing their strategies and efficacy. The most recent microfluidic techniques for spheroid formation utilize emulsion, microwells, U-shaped microstructures, or digital microfluidics. The engineering aspects underpinning spheroid formation in these microfluidic devices are therefore considered. In the second part of this paper, design considerations for microfluidic spheroid formation chips and microfluidic spheroid culture chips (SFCs and SCCs) are evaluated with regard to key parameters affecting spheroid formation, including shear stress, spheroid diameter, culture medium delivery and flow rate. This review is intended to benefit the microfluidics community by contributing to improved design and engineering of microfluidic chips capable of forming and/or culturing three-dimensional cell spheroids.
Sofela, S., Sahloul, S., Rafeie, M., Kwon, T., Han, J., Warkiani, M.E. & Song, Y.-.A. 2018, 'High-throughput sorting of eggs for synchronization of C. elegans in a microfluidic spiral chip.', Lab on a chip, vol. 18, no. 4, pp. 679-687.View/Download from: Publisher's site
In this study, we report the use of a high-throughput microfluidic spiral chip to screen out eggs from a mixed age nematode population, which can subsequently be cultured to a desired developmental stage. For the sorting of a mixture containing three different developmental stages, eggs, L1 and L4, we utilized a microfluidic spiral chip with a trapezoidal channel to obtain a sorting efficiency of above 97% and a sample purity (SP) of above 80% for eggs at different flow rates up to 10 mL min-1. The result demonstrated a cost-effective, simple, and highly efficient method for synchronizing C. elegans at a high throughput (4200 organisms per min at 6 mL min-1), while eliminating challenges such as clogging and non-reusability of membrane-based filtration. Due to its simplicity, our method can be easily adopted in the C. elegans research community.
Syed, M.S., Rafeie, M., Vandamme, D., Asadnia, M., Henderson, R., Taylor, R.A. & Warkiani, M.E. 2018, 'Selective separation of microalgae cells using inertial microfluidics.', Bioresource technology, vol. 252, pp. 91-99.View/Download from: Publisher's site
Microalgae represent the most promising new source of biomass for the world's growing demands. However, the biomass productivity and quality is significantly decreased by the presence of bacteria or other invading microalgae species in the cultures. We therefore report a low-cost spiral-microchannel that can effectively separate and purify Tetraselmis suecica (lipid-rich microalgae) cultures from Phaeodactylum tricornutum (invasive diatom). Fluorescent polystyrene-microbeads of 6m and 10m diameters were first used as surrogate particles to optimize the microchannel design by mimicking the microalgae cell behaviour. Using the optimum flowrate, up to 95% of the P. tricornutum cells were separated from the culture without affecting the cell viability. This study shows, for the first time, the potential of inertial microfluidics to sort microalgae species with minimal size difference. Additionally, this approach can also be applied as a pre-sorting technique for water quality analysis.
Alizadeh, A., Warkiani, M.E. & Wang, M. 2017, 'Manipulating electrokinetic conductance of nanofluidic channel by varying inlet pH of solution', Microfluidics and Nanofluidics, vol. 21, no. 3, pp. 1-15.View/Download from: UTS OPUS or Publisher's site
© 2017, Springer-Verlag Berlin Heidelberg. The electrokinetic conductivity of micro-/nanofluidic systems, which strongly depends on the local solution properties (e.g., pH and ionic strength), has wide applications in nanosystems to control the system performance and ion rectification. Accurate and active manipulation of this parameter is proven to be very challenging since, in nanoscale, the ion transport is particularly dominated by the acquired surface charge on the solid–liquid interfaces. In this study, we propose an approach to manipulate the nanochannel electrokinetic conductivity by changing the pH value of the solution at the inlet in order to impose asymmetrical conditions inside nanochannel. The variable surface charge of walls is determined by considering the chemical adsorption on the solid–liquid interface and the electrical double layer interaction. The presented numerical model, which couples Poisson–Nernst–Planck and Navier–Stokes equations, can fully consider the electro-chemo-mechanical transport phenomena and predict the electrokinetic conductivity of nanofluidic channels with good accuracy. Modeling results show that the electrokinetic conductivity of the nanofluidic systems can be regulated by varying the solution pH at the inlet. It is revealed that the stronger electric double layers interaction can enhance the sensitivity of the nanochannel electrokinetic conductance to the inlet pH. This unique behavior of the nanochannel electrokinetic conductivity could broaden potential applications in biomedical, energy, and environmental systems using nanofluidic devices.
Asadnia, M., Ehteshami, S.M.M., Chan, S.H. & Warkiani, M.E. 2017, 'Development of a fiber-based membraneless hydrogen peroxide fuel cell', RSC Advances, vol. 7, no. 65, pp. 40755-40760.View/Download from: UTS OPUS or Publisher's site
© 2017 The Royal Society of Chemistry. In this paper, polyvinylidene fluoride (PVDF) nanofibers have been suggested as a viable substrate for flexible and implantable electrochemical devices. PVDF electrospun nanofibers exhibit excellent mechanical properties, flexibility, chemical stability, and biocompatibility, making them a potential option in the development of implant fuel cells. This paper presents a membraneless hydrogen peroxide fuel cell that is fabricated to demonstrate the possibility of using these nanofibers as the substrate for electrochemical devices. An open circuit potential of 0.65 V was achieved for the cell fabricated using Prussian Blue (PB) as the cathode material and nickel and aluminium as the anode materials. The power produced by the cell was 1 mW cm-2at 0.32 V. The results presented compare favourably with available power generators reviewed in the literature. Based on the proof of concept demonstration; PVDF electrospun nanofibers can be successfully used for implantable electrochemical devices such as bio-fuel cells and self-sustained point-of-care diagnostic systems.
Asadnia, M., Khorasani, A.M. & Warkiani, M.E. 2017, 'An Accurate PSO-GA Based Neural Network to Model Growth of Carbon Nanotubes', Journal of Nanomaterials, vol. 2017.View/Download from: UTS OPUS or Publisher's site
© 2017 Mohsen Asadnia et al. By combining particle swarm optimization (PSO) and genetic algorithms (GA) this paper offers an innovative algorithm to train artificial neural networks (ANNs) for the purpose of calculating the experimental growth parameters of CNTs. The paper explores experimentally obtaining data to train ANNs, as a method to reduce simulation time while ensuring the precision of formal physics models. The results are compared with conventional particle swarm optimization based neural network (CPSONN) and Levenberg-Marquardt (LM) techniques. The results show that PSOGANN can be successfully utilized for modeling the experimental parameters that are critical for the growth of CNTs.
Gerami, A., Armstrong, R.T., Johnston, B., Warkiani, M.E., Mosavat, N. & Mostaghimi, P. 2017, 'Coal-on-a-Chip: Visualizing Flow in Coal Fractures', Energy and Fuels, vol. 31, no. 10, pp. 10393-10403.View/Download from: UTS OPUS or Publisher's site
© 2017 American Chemical Society. Geomaterial microfluidics are the next generation of tools necessary for studying fluid flows related to subsurface engineering technologies. Traditional microfluidic devices do not capture surface wettability and roughness parameters that can have a significant influence on porous media flows. This is particularly important for coal seam gas reservoirs in which methane gas is transported through a well-developed system of natural fractures that display unique wettability and roughness characteristics. A coal geomaterial microfluidic device can be generated by etching a fracture pattern on a coal surface by using three-dimensional laser micromachining; however, it is unclear if the resulting surface properties are representative of real coal. In an effort to generate a realistic coal microfluidic device, we characterize coal surface roughness properties from real coal cleats. We then compare these results to the roughness of the patterns, generated from laser etching. Roughness measurements in real coal fractures show that cleats and microfractures are mostly oriented parallel to the coal beddings rather than perpendicular to the bedding, which is important when selecting coal for fabrication of a microfluidic device since we find that the natural microfractures influence the resulting roughness of etched fractures. We also compare resulting coal/brine/gas contact angles under static and dynamics conditions. The contact angle for coal is highly heterogeneous. Surface roughness and pore pressure may influence the contact angle. With the aid of the coal geomaterial device, the effect of these parameters on coal wettability can be explored and a range of possible coal contact angles can be visualized and represented. The geomaterial fabrication, as outlined herein, provides a tool to capture more realistic coal surface properties in microfluidics experiments.
Kulasinghe, A., Perry, C., Kenny, L., Warkiani, M.E., Nelson, C. & Punyadeera, C. 2017, 'PD-L1 expressing circulating tumour cells in head and neck cancers.', BMC Cancer, vol. 17, no. 1, pp. 1-6.View/Download from: UTS OPUS or Publisher's site
Blockade of the PD-1/PD-L1 immune checkpoint pathway is emerging as a promising immunotherapeutic approach for the management and treatment of head and neck cancer patients who do not respond to 1st/2nd line therapy. However, as checkpoint inhibitors are cost intensive, identifying patients who would most likely benefit from anti PD-L1 therapy is required. Developing a non-invasive technique would be of major benefit to the patient and to the health care system.We report the case of a 56 year old man affected by a supraglottic squamous cell carcinoma (SCC). A CT scan showed a 20 mm right jugulodigastric node and suspicious lung lesions. The lung lesion was biopsied and confirmed to be consistent with SCC. The patient was offered palliative chemotherapy. At the time of presentation, a blood sample was taken for circulating tumour cell (CTC) analysis. The dissemination of cancer was confirmed by the detection of CTCs in the peripheral blood of the patient, measured by the CellSearch System (Janssen Diagnostics). Using marker-independent, low-shear spiral microfluidic technology combined with immunocytochemistry, CTC clusters were found in this patient at the same time point, expressing PD-L1.This report highlights the potential use of CTCs to identify patients which might respond to anti PD-L1 therapy.
Kulasinghe, A., Tran, T.H.P., Blick, T., O'Byrne, K., Thompson, E.W., Warkiani, M.E., Nelson, C., Kenny, L. & Punyadeera, C. 2017, 'Enrichment of circulating head and neck tumour cells using spiral microfluidic technology', Scientific Reports, vol. 7, pp. 1-10.View/Download from: UTS OPUS or Publisher's site
© 2017 The Author(s). Whilst locoregional control of head and neck cancers (HNCs) has improved over the last four decades, long-term survival has remained largely unchanged. A possible reason for this is that the rate of distant metastasis has not changed. Such disseminated disease is reflected in measurable levels of cancer cells in the blood of HNC patients, referred to as circulating tumour cells (CTCs). Numerous marker-independent techniques have been developed for CTC isolation and detection. Recently, microfluidics-based platforms have come to the fore to avoid molecular bias. In this pilot, proof of concept study, we evaluated the use of the spiral microfluidic chip for CTC enrichment and subsequent detection in HNC patients. CTCs were detected in 13/24 (54%) HNC patients, representing both early to late stages of disease. Importantly, in 7/13 CTC-positive patients, CTC clusters were observed. This is the first study to use spiral microfluidics technology for CTC enrichment in HNC.
Kwon, T., Prentice, H., Oliveira, J.D., Madziva, N., Warkiani, M.E., Hamel, J.-.F.P. & Han, J. 2017, 'Microfluidic Cell Retention Device for Perfusion of Mammalian Suspension Culture.', Scientific Reports, vol. 7, no. 1, pp. 1-11.View/Download from: UTS OPUS or Publisher's site
Continuous production of biologics, a growing trend in the biopharmaceutical industry, requires a reliable and efficient cell retention device that also maintains cell viability. Current filtration methods, such as tangential flow filtration using hollow-fiber membranes, suffer from membrane fouling, leading to significant reliability and productivity issues such as low cell viability, product retention, and an increased contamination risk associated with filter replacement. We introduce a novel cell retention device based on inertial sorting for perfusion culture of suspended mammalian cells. The device was characterized in terms of cell retention capacity, biocompatibility, scalability, and long-term reliability. This technology was demonstrated using a high concentration (>20 million cells/mL) perfusion culture of an IgG1-producing Chinese hamster ovary (CHO) cell line for 18-25 days. The device demonstrated reliable and clog-free cell retention, high IgG1 recovery (>99%) and cell viability (>97%). Lab-scale perfusion cultures (350mL) were used to demonstrate the technology, which can be scaled-out with parallel devices to enable larger scale operation. The new cell retention device is thus ideal for rapid perfusion process development in a biomanufacturing workflow.
Rafeie, M., Welleweerd, M., Hassanzadeh-Barforoushi, A., Asadnia, M., Olthuis, W. & Warkiani, M.E. 2017, 'An easily fabricated three-dimensional threaded lemniscate-shaped micromixer for a wide range of flow rates', Biomicrofluidics, vol. 11, no. 1, pp. 1-15.View/Download from: UTS OPUS or Publisher's site
Mixing fluid samples or reactants is a paramount function in the fields of micro total analysis system (TAS) and microchemical processing. However, rapid and efficient fluid mixing is difficult to achieve inside microchannels because of the difficulty of diffusive mass transfer in the laminar regime of the typical microfluidic flows. It has been well recorded that the mixing efficiency can be boosted by migrating from two-dimensional (2D) to three-dimensional (3D) geometries. Although several 3D chaotic mixers have been designed, most of them offer a high mixing efficiency only in a very limited range of Reynolds numbers (Re). In this work, we developed a 3D fine-threaded lemniscate-shaped micromixer whose maximum numerical and empirical efficiency is around 97% and 93%, respectively, and maintains its high performance (i.e., >90%) over a wide range of 1
Ramalingam, N., Warkiani, M.E. & Hai-Qing Gong, T. 2017, 'Acetylated bovine serum albumin differentially inhibits polymerase chain reaction in microdevices', Biomicrofluidics, vol. 11, no. 3, pp. 1-7.View/Download from: UTS OPUS or Publisher's site
Bovine serum albumin (BSA) is widely used as an additive in polymerase chain reaction (PCR)-based microfluidic devices to passivate reactors and alleviate nucleic-acid amplification. BSA is available commercially in two types: either acetylated or non-acetylated. A survey of literature indicates that both types of BSA are used in PCR-based microfluidic devices. Our study results reveal that the use of acetylated BSA in PCR micro-devices leads to differential inhibition of PCR, compared to non-acetylated BSA. This result is noticed for the first time, and the differential inhibition generally goes un-noticed, as compared to complete PCR inhibition.
Razmjou, A., Asadnia, M., Ghaebi, O., Yang, H.C., Ebrahimi Warkiani, M., Hou, J. & Chen, V. 2017, 'Preparation of Iridescent 2D Photonic Crystals by Using a Mussel-Inspired Spatial Patterning of ZIF-8 with Potential Applications in Optical Switch and Chemical Sensor', ACS Applied Materials and Interfaces, vol. 9, no. 43, pp. 38076-38080.View/Download from: UTS OPUS or Publisher's site
© 2017 American Chemical Society. In this work, spatial patterning of a thin, dense, zeolitic imidazolate framework (ZIF-8) pattern was generated using photolithography and nanoscale (60 nm) dopamine coating. A bioinspired, unique, reversible, two-color iridescent pattern can be easily obtained for potential applications in sensing and photonics.
Sengupta, D., Kottapalli, A.G.P., Chen, S.H., Miao, J.M., Kwok, C.Y., Triantafyllou, M.S., Warkiani, M.E. & Asadnia, M. 2017, 'Characterization of single polyvinylidene fluoride (PVDF) nanofiber for flow sensing applications', AIP Advances, vol. 7, no. 10, pp. 1-8.View/Download from: UTS OPUS or Publisher's site
© 2017 Author(s). The use of Polyvinylidene Fluoride (PVDF) based piezoelectric nanofibers for sensing and actuation has been reported widely in the past. However, in most cases, PVDF piezoelectric nanofiber mats have been used for sensing applications. This work fundamentally characterizes a single electrospun PVDF nanofiber and demonstrates its application as a sensing element for nanoelectromechanical sensors (NEMS). PVDF nanofiber mats were spun by far field electrospinning (FFES) process and complete material characterization was conducted by means of scanning electron microscope (SEM) imaging, Raman Spectroscopy and FTIR spectroscopy. An optimized recipe was developed for spinning a single suspended nanofiber on a specially designed MEMS substrate which allows the nano-mechanical and electrical characterization of a single PVDF nanofiber. Electrical characterization is conducted using a single suspended nanofiber to determine the piezoelectric coefficient (d33) of the nanofiber to be -58.77 pm/V. Also the mechanical characterization conducted using a nanoindenter revealed a Young's Modulus and hardness of 2.2 GPa and 0.1 GPa respectively. Finally, an application that utilizes the single PVDF nanofiber as a sensing element to form a NEMS flow sensor is demonstrated. The single nanofiber flow sensor is tested in presence of various oscillatory flow conditions.
Shakeel Syed, M., Rafeie, M., Henderson, R., Vandamme, D., Asadnia, M. & Ebrahimi Warkiani, M. 2017, 'A 3D-printed mini-hydrocyclone for high throughput particle separation: Application to primary harvesting of microalgae', Lab on a Chip, vol. 17, no. 14, pp. 2459-2469.View/Download from: UTS OPUS or Publisher's site
© The Royal Society of Chemistry 2017. The separation of micro-sized particles in a continuous flow is crucial part of many industrial processes, from biopharmaceutical manufacturing to water treatment. Conventional separation techniques such as centrifugation and membrane filtration are largely limited by factors such as clogging, processing time and operation efficiency. Microfluidic based techniques have been gaining great attention in recent years as efficient and powerful approaches for particle-liquid separation. Yet the production of such systems using standard micro-fabrication techniques is proven to be tedious, costly and have cumbersome user interfaces, which all render commercialization difficult. Here, we demonstrate the design, fabrication and evaluation based on CFD simulation as well as experimentation of 3D-printed miniaturized hydrocyclones with smaller cut-size for high-throughput particle/cell sorting. The characteristics of the mini-cyclones were numerically investigated using computational fluid dynamics (CFD) techniques previously revealing that reduction in the size of the cyclone results in smaller cut-size of the particles. To showcase its utility, high-throughput algae harvesting from the medium with low energy input is demonstrated for the marine microalgae Tetraselmis suecica. Final microalgal biomass concentration was increased by 7.13 times in 11 minutes of operation time using our designed hydrocyclone (HC-1). We expect that this elegant approach can surmount the shortcomings of other microfluidic technologies such as clogging, low-throughput, cost and difficulty in operation. By moving away from production of planar microfluidic systems using conventional microfabrication techniques and embracing 3D-printing technology for construction of discrete elements, we envision 3D-printed mini-cyclones can be part of a library of standardized active and passive microfluidic components, suitable for particle-liquid separation.
Shirani, E., Razmjou, A., Tavassoli, H., Landarani-Isfahani, A., Rezaei, S., Abbasi Kajani, A., Asadnia, M., Hou, J. & Ebrahimi Warkiani, M. 2017, 'Strategically Designing a Pumpless Microfluidic Device on an "inert" Polypropylene Substrate with Potential Application in Biosensing and Diagnostics', Langmuir, vol. 33, no. 22, pp. 5565-5576.View/Download from: UTS OPUS or Publisher's site
© 2017 American Chemical Society. This study is an attempt to make a step forward to implement the very immature concept of pumpless transportation of liquid into a real miniaturized device or lab-on-chip (LOC) on a plastic substrate. "Inert" plastic materials such as polypropylene (PP) are used in a variety of biomedical applications but their surface engineering is very challenging. Here, it was demonstrated that with a facile innovative wettability patterning route using fluorosilanized UV-independent TiO 2 nanoparticle coating it is possible to create wedge-shaped open microfluidic tracks on inert solid surfaces for low-cost biomedical devices (lab-on-plastic). For the future miniaturization and integration of the tracks into a device, a variety of characterization techniques were used to not only systematically study the surface patterning chemistry and topography but also to have a clear knowledge of its biological interactions and performance. The effect of such surface architecture on the biological performance was studied in terms of static/dynamic protein (bovine serum albumin) adsorption, bacterial (Staphylococcus aureus and Staphylococcus epidermidis) adhesion, cell viability (using HeLa and MCF-7 cancer cell lines as well as noncancerous human fibroblast cells), and cell patterning (Murine embryonic fibroblasts). Strategies are discussed for incorporating such a confined track into a diagnostic device in which its sensing portion is based on protein, microorganism, or cells. Finally, for the proof-of-principle of biosensing application, the well-known high-affinity molecular couple of BSA-antiBSA as a biological model was employed.
Hassanzadeh-Barforoushi, A., Shemesh, J., Farbehi, N., Asadnia, M., Yeoh, G.H., Harvey, R.P., Nordon, R.E. & Warkiani, M.E. 2016, 'A rapid co-culture stamping device for studying intercellular communication.', Scientific reports, vol. 6, p. 35618.View/Download from: Publisher's site
Regulation of tissue development and repair depends on communication between neighbouring cells. Recent advances in cell micro-contact printing and microfluidics have facilitated the in-vitro study of homotypic and heterotypic cell-cell interaction. Nonetheless, these techniques are still complicated to perform and as a result, are seldom used by biologists. We report here development of a temporarily sealed microfluidic stamping device which utilizes a novel valve design for patterning two adherent cell lines with well-defined interlacing configurations to study cell-cell interactions. We demonstrate post-stamping cell viability of >95%, the stamping of multiple adherent cell types, and the ability to control the seeded cell density. We also show viability, proliferation and migration of cultured cells, enabling analysis of co-culture boundary conditions on cell fate. We also developed an in-vitro model of endothelial and cardiac stem cell interactions, which are thought to regulate coronary repair after myocardial injury. The stamp is fabricated using microfabrication techniques, is operated with a lab pipettor and uses very low reagent volumes of 20l with cell injection efficiency of >70%. This easy-to-use device provides a general strategy for micro-patterning of multiple cell types and will be important for studying cell-cell interactions in a multitude of applications.
Khoo, B.L., Chaudhuri, P.K., Ramalingam, N., Tan, D.S.W., Lim, C.T. & Warkiani, M.E. 2016, 'Single-cell profiling approaches to probing tumor heterogeneity.', International journal of cancer, vol. 139, no. 2, pp. 243-255.View/Download from: Publisher's site
Tumor heterogeneity is a major hindrance in cancer classification, diagnosis and treatment. Recent technological advances have begun to reveal the true extent of its heterogeneity. Single-cell analysis (SCA) is emerging as an important approach to detect variations in morphology, genetic or proteomic expression. In this review, we revisit the issue of inter- and intra-tumor heterogeneity, and list various modes of SCA techniques (cell-based, nucleic acid-based, protein-based, metabolite-based and lipid-based) presently used for cancer characterization. We further discuss the advantages of SCA over pooled cell analysis, as well as the limitations of conventional techniques. Emerging trends, such as high-throughput sequencing, are also mentioned as improved means for cancer profiling. Collectively, these applications have the potential for breakthroughs in cancer treatment.
Kulasinghe, A., Perry, C., Warkiani, M.E., Blick, T., Davies, A., O'Byrne, K., Thompson, E.W., Nelson, C.C., Vela, I. & Punyadeera, C. 2016, 'Short term ex-vivo expansion of circulating head and neck tumour cells.', Oncotarget, vol. 7, no. 37, pp. 60101-60109.View/Download from: Publisher's site
Minimally invasive techniques are required for the identification of head and neck cancer (HNC) patients who are at an increased risk of metastasis, or are not responding to therapy. An approach utilised in other solid cancers is the identification and enumeration of circulating tumour cells (CTCs) in the peripheral blood of patients. Low numbers of CTCs has been a limiting factor in the HNC field to date. Here we present a methodology to expand HNC patient derived CTCs ex-vivo. As a proof of principle study, 25 advanced stage HNC patient bloods were enriched for circulating tumour cells through negative selection and cultured in 2D and 3D culture environments under hypoxic conditions (2% O2, 5% CO2). CTCs were detected in 14/25 (56%) of patients (ranging from 1-15 CTCs/5 mL blood). Short term CTC cultures were successfully generated in 7/25 advanced stage HNC patients (5/7 of these cultures were from HPV+ patients). Blood samples from which CTC culture was successful had higher CTC counts (p = 0.0002), and were predominantly from HPV+ patients (p = 0.007). This is, to our knowledge, the first pilot study to culture HNC CTCs ex-vivo. Further studies are warranted to determine the use of short term expansion in HNC and the role of HPV in promoting culture success.
Rafeie, M., Zhang, J., Asadnia, M., Li, W. & Warkiani, M.E. 2016, 'Multiplexing slanted spiral microchannels for ultra-fast blood plasma separation.', Lab on a chip, vol. 16, no. 15, pp. 2791-2802.View/Download from: Publisher's site
Blood and blood products are critical components of health care. Blood components perform distinct functions in the human body and thus the ability to efficiently fractionate blood into its individual components (i.e., plasma and cellular components) is of utmost importance for therapeutic and diagnostic purposes. Although conventional approaches like centrifugation and membrane filtration for blood processing have been successful in generating relatively pure fractions, they are largely limited by factors such as the required blood sample volume, component purity, clogging, processing time and operation efficiency. In this work, we developed a high-throughput inertial microfluidic system for cell focusing and blood plasma separation from small to large volume blood samples (1-100 mL). Initially, polystyrene beads and blood cells were used to investigate the inertial focusing performance of a single slanted spiral microchannel as a function of particle size, flow rate, and blood cell concentration. Afterwards, blood plasma separation was conducted using an optimised spiral microchannel with relatively large dimensions. It was found that the reject ratio of the slanted spiral channel is close to 100% for blood samples with haematocrit (HCT) values of 0.5% and 1% under an optimal flow rate of 1.5 mL min(-1). Finally, through a unique multiplexing approach, we built a high-throughput system consisting of 16 spiral channels connected together, which can process diluted samples with a total flow rate as high as 24 mL min(-1). The proposed multiplexed system can surmount the shortcomings of previously reported microfluidic systems for plasma separation and cell sorting in terms of throughput, yield and operation efficiency.
Ramalingam, N., Warkiani, M.E., Ramalingam, N., Keshavarzi, G., Hao-Bing, L. & Hai-Qing, T.G. 2016, 'Numerical and experimental study of capillary-driven flow of PCR solution in hybrid hydrophobic microfluidic networks.', Biomedical microdevices, vol. 18, no. 4, p. 68.View/Download from: Publisher's site
Capillary-driven microfluidics is essential for development of point-of-care diagnostic micro-devices. Polymerase chain reaction (PCR)-based micro-devices are widely developed and used in such point-of-care settings. It is imperative to characterize the fluid parameters of PCR solution for designing efficient capillary-driven microfluidic networks. Generally, for numeric modelling, the fluid parameters of PCR solution are approximated to that of water. This procedure leads to inaccurate results, which are discrepant to experimental data. This paper describes mathematical modeling and experimental validation of capillary-driven flow inside Poly-(dimethyl) siloxane (PDMS)-glass hybrid micro-channels. Using experimentally measured PCR fluid parameters, the capillary meniscus displacement in PDMS-glass microfluidic ladder network is simulated using computational fluid dynamic (CFD), and experimentally verified to match with the simulated data.
Warkiani, M.E., Khoo, B.L., Wu, L., Tay, A.K.P., Bhagat, A.A.S., Han, J. & Lim, C.T. 2016, 'Ultra-fast, label-free isolation of circulating tumor cells from blood using spiral microfluidics.', Nature protocols, vol. 11, no. 1, pp. 134-148.View/Download from: Publisher's site
Circulating tumor cells (CTCs) are rare cancer cells that are shed from primary or metastatic tumors into the peripheral blood circulation. Phenotypic and genetic characterization of these rare cells can provide important information to guide cancer staging and treatment, and thus further research into their characteristics and properties is an area of considerable interest. In this protocol, we describe detailed procedures for the production and use of a label-free spiral microfluidic device to allow size-based isolation of viable CTCs using hydrodynamic forces that are present in curvilinear microchannels. This spiral system enables us to achieve 85% recovery of spiked cells across multiple cancer cell lines and 99.99% depletion of white blood cells in whole blood. The described spiral microfluidic devices can be produced at an extremely low cost using standard microfabrication and soft lithography techniques (2-3 d), and they can be operated using two syringe pumps for lysed blood samples (7.5 ml in 12.5 min for a three-layered multiplexed chip). The fast processing time and the ability to collect CTCs from a large patient blood volume allows this technique to be used experimentally in a broad range of potential genomic and transcriptomic applications.
Chaudhuri, P.K., Ebrahimi Warkiani, M., Jing, T., Kenry & Lim, C.T. 2016, 'Microfluidics for research and applications in oncology.', The Analyst, vol. 141, no. 2, pp. 504-524.View/Download from: Publisher's site
Cancer is currently one of the top non-communicable human diseases, and continual research and developmental efforts are being made to better understand and manage this disease. More recently, with the improved understanding in cancer biology as well as the advancements made in microtechnology and rapid prototyping, microfluidics is increasingly being explored and even validated for use in the detection, diagnosis and treatment of cancer. With inherent advantages such as small sample volume, high sensitivity and fast processing time, microfluidics is well-positioned to serve as a promising platform for applications in oncology. In this review, we look at the recent advances in the use of microfluidics, from basic research such as understanding cancer cell phenotypes as well as metastatic behaviors to applications such as the detection, diagnosis, prognosis and drug screening. We then conclude with a future outlook on this promising technology.
Tay, A., Pavesi, A., Yazdi, S.R., Lim, C.T. & Warkiani, M.E. 2016, 'Advances in microfluidics in combating infectious diseases.', Biotechnology advances, vol. 34, no. 4, pp. 404-421.View/Download from: Publisher's site
One of the important pursuits in science and engineering research today is to develop low-cost and user-friendly technologies to improve the health of people. Over the past decade, research efforts in microfluidics have been made to develop methods that can facilitate low-cost diagnosis of infectious diseases, especially in resource-poor settings. Here, we provide an overview of the recent advances in microfluidic devices for point-of-care (POC) diagnostics for infectious diseases and emphasis is placed on malaria, sepsis and AIDS/HIV. Other infectious diseases such as SARS, tuberculosis, and dengue are also briefly discussed. These infectious diseases are chosen as they contribute the most to disability-adjusted life-years (DALYs) lost according to the World Health Organization (WHO). The current state of research in this area is evaluated and projection toward future applications and accompanying challenges are also discussed.
Zarepour, E., Hassan, M., Chou, C.T. & Ebrahimi Warkiani, M. 2016, 'Characterizing terahertz channels for monitoring human lungs with wireless nanosensor networks', Nano Communication Networks, vol. 9, pp. 43-57.View/Download from: Publisher's site
© 2016 We characterize terahertz wireless channels for extracting data from nanoscale sensors deployed within human lungs. We discover that the inhalation and exhalation of oxygen and carbon dioxide causes periodic variation of the absorption coefficient of the terahertz channel. Channel absorption drops to its minimum near the end of inhalation, providing a window of opportunity to extract data with minimum transmission power. We propose an algorithm for nanosensors to estimate the periodic channel by observing signal-to-noise ratio of the beacons transmitted from the data sink. Using real respiration data from multiple subjects, we demonstrate that the proposed algorithm can estimate the minimum absorption interval of the periodic channel with 98.5% accuracy. Our analysis shows that by confining all data collections during the estimated low-absorption window of the periodic channel, nanosensors can reduce power consumption by six orders of magnitude. Finally, we demonstrate that for wireless communications within human lungs, 0.1–0.12 THz is the least absorbing spectrum within the terahertz band.
Zhang, J., Yan, S., Yuan, D., Alici, G., Nguyen, N.-.T., Ebrahimi Warkiani, M. & Li, W. 2016, 'Fundamentals and applications of inertial microfluidics: a review.', Lab on a chip, vol. 16, no. 1, pp. 10-34.View/Download from: Publisher's site
In the last decade, inertial microfluidics has attracted significant attention and a wide variety of channel designs that focus, concentrate and separate particles and fluids have been demonstrated. In contrast to conventional microfluidic technologies, where fluid inertia is negligible and flow remains almost within the Stokes flow region with very low Reynolds number (Re 1), inertial microfluidics works in the intermediate Reynolds number range (~1 < Re < ~100) between Stokes and turbulent regimes. In this intermediate range, both inertia and fluid viscosity are finite and bring about several intriguing effects that form the basis of inertial microfluidics including (i) inertial migration and (ii) secondary flow. Due to the superior features of high-throughput, simplicity, precise manipulation and low cost, inertial microfluidics is a very promising candidate for cellular sample processing, especially for samples with low abundant targets. In this review, we first discuss the fundamental kinematics of particles in microchannels to familiarise readers with the mechanisms and underlying physics in inertial microfluidic systems. We then present a comprehensive review of recent developments and key applications of inertial microfluidic systems according to their microchannel structures. Finally, we discuss the perspective of employing fluid inertia in microfluidics for particle manipulation. Due to the superior benefits of inertial microfluidics, this promising technology will still be an attractive topic in the near future, with more novel designs and further applications in biology, medicine and industry on the horizon.
Asadnia, M., Kottapalli, A.G.P., Miao, J., Warkiani, M.E. & Triantafyllou, M.S. 2015, 'Artificial fish skin of self-powered micro-electromechanical systems hair cells for sensing hydrodynamic flow phenomena.', Journal of the Royal Society, Interface, vol. 12, no. 111, p. 20150322.View/Download from: Publisher's site
Using biological sensors, aquatic animals like fishes are capable of performing impressive behaviours such as super-manoeuvrability, hydrodynamic flow 'vision' and object localization with a success unmatched by human-engineered technologies. Inspired by the multiple functionalities of the ubiquitous lateral-line sensors of fishes, we developed flexible and surface-mountable arrays of micro-electromechanical systems (MEMS) artificial hair cell flow sensors. This paper reports the development of the MEMS artificial versions of superficial and canal neuromasts and experimental characterization of their unique flow-sensing roles. Our MEMS flow sensors feature a stereolithographically fabricated polymer hair cell mounted on Pb(Zr(0.52)Ti(0.48))O3 micro-diaphragm with floating bottom electrode. Canal-inspired versions are developed by mounting a polymer canal with pores that guide external flows to the hair cells embedded in the canal. Experimental results conducted employing our MEMS artificial superficial neuromasts (SNs) demonstrated a high sensitivity and very low threshold detection limit of 22 mV/(mm s(-1)) and 8.2 µm s(-1), respectively, for an oscillating dipole stimulus vibrating at 35 Hz. Flexible arrays of such superficial sensors were demonstrated to localize an underwater dipole stimulus. Comparative experimental studies revealed a high-pass filtering nature of the canal encapsulated sensors with a cut-off frequency of 10 Hz and a flat frequency response of artificial SNs. Flexible arrays of self-powered, miniaturized, light-weight, low-cost and robust artificial lateral-line systems could enhance the capabilities of underwater vehicles.
Warkiani, M.E., Tay, A.K.P., Khoo, B.L., Xiaofeng, X., Han, J. & Lim, C.T. 2015, 'Malaria detection using inertial microfluidics.', Lab on a chip, vol. 15, no. 4, pp. 1101-1109.View/Download from: Publisher's site
Diagnosis of malaria at the early stage of infection is challenging due to the difficulty in detecting low abundance parasites from blood. Molecular methods such as real-time polymerase chain reaction (qPCR) can be especially useful for detecting low parasitemia levels due to their high sensitivity and their ability to recognize different malarial species and strains. Unfortunately, the accuracy of qPCR-based malaria detection can be compromised by many factors, including the limited specificity of primers, presence of PCR inhibitors in blood serum and DNA contamination from nucleated blood cells. Here, we use a label-free, shear-modulated inertial microfluidic system to enrich malaria parasites from blood so as to facilitate a more reliable and specific PCR-based malaria detection. This technique capitalizes on cell focusing behaviors in high aspect ratio microchannels coupled with pinched flow dynamics to isolate ring-stage malaria parasites from lysed blood containing white blood cells (WBCs). In this high aspect ratio (ratio of the channel height to the width) platform, the high shear rate along the channel width causes the dispersed WBCs at the inlet to migrate and align into two streams near the channel sidewalls while the malaria parasites remain unfocused. Sensitive detection of parasites at spiked densities ranging from 10(3) to 10(4)Plasmodium falciparum parasites per mL (~2-10 per L) has been demonstrated; they have also been quantified in whole blood using qPCR. This is approximately 100-fold more sensitive than the gold standard conventional microscopy analysis of thick blood smears. The simplicity of this device makes it ideal for integration with an automatic system for ultra-fast and accurate detection despite low levels of parasitemia. It can also help in malaria screening and elimination efforts.
Khoo, B.L., Lee, S.C., Kumar, P., Tan, T.Z., Warkiani, M.E., Ow, S.G.W., Nandi, S., Lim, C.T. & Thiery, J.P. 2015, 'Short-term expansion of breast circulating cancer cells predicts response to anti-cancer therapy.', Oncotarget, vol. 6, no. 17, pp. 15578-15593.View/Download from: Publisher's site
Circulating tumor cells (CTCs) are considered as surrogate markers for prognosticating and evaluating patient treatment responses. Here, 226 blood samples from 92 patients with breast cancer, including patients with newly diagnosed or metastatic refractory cancer, and 16 blood samples from healthy subjects were cultured in laser-ablated microwells. Clusters containing an increasing number of cytokeratin-positive (CK+) cells appeared after 2 weeks, while most blood cells disappeared with time. Cultures were heterogeneous and exhibited two distinct sub-populations of cells: 'Small' ( 25 m; high nuclear/cytoplasmic ratio; CD45-) cells, comprising CTCs, and 'Large' (> 25 m; low nuclear/cytoplasmic ratio; CD68+ or CD56+) cells, corresponding to macrophage and natural killer-like cells. The Small cell fraction also showed copy number increases in six target genes (FGFR1, Myc, CCND1, HER2, TOP2A and ZNF217) associated with breast cancer. These expanded CTCs exhibited different proportions of epithelial-mesenchymal phenotypes and were transferable for further expansion as spheroids in serum-free suspension or 3D cultures. Cluster formation was affected by the presence and duration of systemic therapy, and its persistence may reflect therapeutic resistance. This novel and advanced method estimates CTC clonal heterogeneity and can predict, within a relatively short time frame, patient responses to therapy.
Shemesh, J., Jalilian, I., Shi, A., Heng Yeoh, G., Knothe Tate, M.L. & Ebrahimi Warkiani, M. 2015, 'Flow-induced stress on adherent cells in microfluidic devices.', Lab on a chip, vol. 15, no. 21, pp. 4114-4127.View/Download from: Publisher's site
Transduction of mechanical forces and chemical signals affect every cell in the human body. Fluid flow in systems such as the lymphatic or circulatory systems modulates not only cell morphology, but also gene expression patterns, extracellular matrix protein secretion and cell-cell and cell-matrix adhesions. Similar to the role of mechanical forces in adaptation of tissues, shear fluid flow orchestrates collective behaviours of adherent cells found at the interface between tissues and their fluidic environments. These behaviours range from alignment of endothelial cells in the direction of flow to stem cell lineage commitment. Therefore, it is important to characterize quantitatively fluid interface-dependent cell activity. Common macro-scale techniques, such as the parallel plate flow chamber and vertical-step flow methods that apply fluid-induced stress on adherent cells, offer standardization, repeatability and ease of operation. However, in order to achieve improved control over a cell's microenvironment, additional microscale-based techniques are needed. The use of microfluidics for this has been recognized, but its true potential has emerged only recently with the advent of hybrid systems, offering increased throughput, multicellular interactions, substrate functionalization on 3D geometries, and simultaneous control over chemical and mechanical stimulation. In this review, we discuss recent advances in microfluidic flow systems for adherent cells and elaborate on their suitability to mimic physiologic micromechanical environments subjected to fluid flow. We describe device design considerations in light of ongoing discoveries in mechanobiology and point to future trends of this promising technology.
Khoo, B.L., Warkiani, M.E., Tan, D.S.-.W., Bhagat, A.A.S., Irwin, D., Lau, D.P., Lim, A.S.T., Lim, K.H., Krisna, S.S., Lim, W.-.T., Yap, Y.S., Lee, S.C., Soo, R.A., Han, J. & Lim, C.T. 2014, 'Clinical validation of an ultra high-throughput spiral microfluidics for the detection and enrichment of viable circulating tumor cells.', PloS one, vol. 9, no. 7, p. e99409.View/Download from: Publisher's site
Circulating tumor cells (CTCs) are cancer cells that can be isolated via liquid biopsy from blood and can be phenotypically and genetically characterized to provide critical information for guiding cancer treatment. Current analysis of CTCs is hindered by the throughput, selectivity and specificity of devices or assays used in CTC detection and isolation.Here, we enriched and characterized putative CTCs from blood samples of patients with both advanced stage metastatic breast and lung cancers using a novel multiplexed spiral microfluidic chip. This system detected putative CTCs under high sensitivity (100%, n=56) (Breast cancer samples: 12-1275 CTCs/ml; Lung cancer samples: 10-1535 CTCs/ml) rapidly from clinically relevant blood volumes (7.5 ml under 5 min). Blood samples were completely separated into plasma, CTCs and PBMCs components and each fraction were characterized with immunophenotyping (Pan-cytokeratin/CD45, CD44/CD24, EpCAM), fluorescence in-situ hybridization (FISH) (EML4-ALK) or targeted somatic mutation analysis. We used an ultra-sensitive mass spectrometry based system to highlight the presence of an EGFR-activating mutation in both isolated CTCs and plasma cell-free DNA (cf-DNA), and demonstrate concordance with the original tumor-biopsy samples.We have clinically validated our multiplexed microfluidic chip for the ultra high-throughput, low-cost and label-free enrichment of CTCs. Retrieved cells were unlabeled and viable, enabling potential propagation and real-time downstream analysis using next generation sequencing (NGS) or proteomic analysis.
Warkiani, M.E., Guan, G., Luan, K.B., Lee, W.C., Bhagat, A.A.S., Chaudhuri, P.K., Tan, D.S.-.W., Lim, W.T., Lee, S.C., Chen, P.C.Y., Lim, C.T. & Han, J. 2014, 'Slanted spiral microfluidics for the ultra-fast, label-free isolation of circulating tumor cells.', Lab on a chip, vol. 14, no. 1, pp. 128-137.View/Download from: Publisher's site
The enumeration and characterization of circulating tumor cells (CTCs), found in the peripheral blood of cancer patients, provide a potentially accessible source for cancer diagnosis and prognosis. This work reports on a novel spiral microfluidic device with a trapezoidal cross-section for ultra-fast, label-free enrichment of CTCs from clinically relevant blood volumes. The technique utilizes the inherent Dean vortex flows present in curvilinear microchannels under continuous flow, along with inertial lift forces which focus larger CTCs against the inner wall. Using a trapezoidal cross-section as opposed to a traditional rectangular cross-section, the position of the Dean vortex core can be altered to achieve separation. Smaller hematologic components are trapped in the Dean vortices skewed towards the outer channel walls and eventually removed at the outer outlet, while the larger CTCs equilibrate near the inner channel wall and are collected from the inner outlet. By using a single spiral microchannel with one inlet and two outlets, we have successfully isolated and recovered more than 80% of the tested cancer cell line cells (MCF-7, T24 and MDA-MB-231) spiked in 7.5 mL of blood within 8 min with extremely high purity (400-680 WBCs mL(-1); ~4 log depletion of WBCs). Putative CTCs were detected and isolated from 100% of the patient samples (n = 10) with advanced stage metastatic breast and lung cancer using standard biomarkers (CK, CD45 and DAPI) with the frequencies ranging from 3-125 CTCs mL(-1). We expect this simple and elegant approach can surmount the shortcomings of traditional affinity-based CTC isolation techniques as well as enable fundamental studies on CTCs to guide treatment and enhance patient care.
Warkiani, M.E., Khoo, B.L., Tan, D.S.-.W., Bhagat, A.A.S., Lim, W.-.T., Yap, Y.S., Lee, S.C., Soo, R.A., Han, J. & Lim, C.T. 2014, 'An ultra-high-throughput spiral microfluidic biochip for the enrichment of circulating tumor cells.', The Analyst, vol. 139, no. 13, pp. 3245-3255.View/Download from: Publisher's site
The detection and characterization of rare circulating tumor cells (CTCs) from the blood of cancer patients can potentially provide critical insights into tumor biology and hold great promise for cancer management. The ability to collect a large number of viable CTCs for various downstream assays such as quantitative measurements of specific biomarkers or targeted somatic mutation analysis is increasingly important in medical oncology. Here, we present a simple yet reliable microfluidic device for the ultra-high-throughput, label-free, size-based isolation of CTCs from clinically relevant blood volumes. The fast processing time of the technique (7.5 mL blood in less than 10 min) and the ability to collect more CTCs from larger blood volumes lends itself to a broad range of potential genomic and transcriptomic applications. A critical advantage of this protocol is the ability to return all fractions of blood (i.e., plasma (centrifugation), CTCs and white blood cells (WBCs) (size-based sorting)) that can be utilized for diverse biomarker studies or time-sensitive molecular assays such as RT-PCR. The clinical use of this biochip was demonstrated by detecting CTCs from 100% (10/10) of blood samples collected from patients with advanced-stage metastatic breast and lung cancers. The CTC recovery rate ranged from 20 to 135 CTCs mL(-1) and obtained under high purity (of 1 CTC out of every 30-100 WBCs which gives 4 log depletion of WBCs). They were identified with immunofluorescence assays (pan-cytokeratin+/CD45-) and molecular probes such as HER2/neu.
Hou, H.W., Warkiani, M.E., Khoo, B.L., Li, Z.R., Soo, R.A., Tan, D.S.-.W., Lim, W.-.T., Han, J., Bhagat, A.A.S. & Lim, C.T. 2013, 'Isolation and retrieval of circulating tumor cells using centrifugal forces.', Scientific reports, vol. 3, p. 1259.View/Download from: Publisher's site
Presence and frequency of rare circulating tumor cells (CTCs) in bloodstreams of cancer patients are pivotal to early cancer detection and treatment monitoring. Here, we use a spiral microchannel with inherent centrifugal forces for continuous, size-based separation of CTCs from blood (Dean Flow Fractionation (DFF)) which facilitates easy coupling with conventional downstream biological assays. Device performance was optimized using cancer cell lines (> 85% recovery), followed by clinical validation with positive CTCs enumeration in all samples from patients with metastatic lung cancer (n = 20; 5-88CTCs per mL). The presence of CD133 cells, a phenotypic marker characteristic of stem-like behavior in lung cancer cells was also identified in the isolated subpopulation of CTCs. The spiral biochip identifies and addresses key challenges of the next generation CTCs isolation assay including antibody independent isolation, high sensitivity and throughput (3mL/hr); and single-step retrieval of viable CTCs.
Warkiani, M.E., Bhagat, A.A.S., Khoo, B.L., Han, J., Lim, C.T., Gong, H.Q. & Fane, A.G. 2013, 'Isoporous micro/nanoengineered membranes.', ACS nano, vol. 7, no. 3, pp. 1882-1904.View/Download from: Publisher's site
Isoporous membranes are versatile structures with numerous potential and realized applications in various fields of science such as micro/nanofiltration, cell separation and harvesting, controlled drug delivery, optics, gas separation, and chromatography. Recent advances in micro/nanofabrication techniques and material synthesis provide novel methods toward controlling the detailed microstructure of membrane materials, allowing fabrication of membranes with well-defined pore size and shape. This review summarizes the current state-of-the-art for isoporous membrane fabrication using different techniques, including microfabrication, anodization, and advanced material synthesis. Various applications of isoporous membranes, such as protein filtration, pathogen isolation, cell harvesting, biosensing, and drug delivery, are also presented.
Warkiani, M.E., Lou, C.-.P., Liu, H.-.B. & Gong, H.-.Q. 2012, 'A high-flux isopore micro-fabricated membrane for effective concentration and recovering of waterborne pathogens.', Biomedical microdevices, vol. 14, no. 4, pp. 669-677.View/Download from: Publisher's site
A high-flux metallic micro/nano-filtration membrane has been fabricated and validated for isolation of waterborne pathogens from drinking water. Obtained membrane with smooth surface and perfectly ordered pores was achieved by a high yield and cost effective multilevel lithography and electroplating technique. The micro-fabricated membrane was also strengthened with an integrated back-support, which can withstand a high pressure during filtration. The results of microfiltration tests with model particles revealed the superior performance of the micro-fabricated filter than current commercial filters in sample throughput, recovery ratio, and reusability. This study highlighted the potential application of micro-fabricated filer in rapid filtration and recovery of C. parvum oocysts for downstream analysis.
Ebrahimi Warkiani, M., Lou, C.P. & Gong, H.Q. 2011, 'Fabrication of multi-layer polymeric micro-sieve having narrow slot pores with conventional ultraviolet-lithography and micro-fabrication techniques', Biomicrofluidics, vol. 5, no. 3.View/Download from: Publisher's site
Fast detection of waterborne pathogens is important for securing the hygiene of drinking water. Detection of pathogens in water at low concentrations and minute quantities demands rapid and efficient enrichment methods in order to improve the signal-to-noise ratio of bio-sensors. We propose and demonstrate a low cost and rapid method to fabricate a multi-layer polymeric micro-sieve using conventional lithography techniques. The micro-fabricated micro-sieves are made of several layers of SU-8 photoresist using multiple coating and exposure steps and a single developing process. The obtained micro-sieves have good mechanical properties, smooth surfaces, high porosity (40%), and narrow pore size distribution (coefficient of variation < 3.33%). Sample loading and back-flushing using the multi-layer micro-sieve resulted in more than 90% recovery of pathogens, which showed improved performance than current commercial filters. © 2011 American Institute of Physics.
Teymourtash, A.R. & Ebrahimi Warkiani, M. 2009, 'Natural convection over a non-isothermal vertical flat plate in supercritical fluids', Scientia Iranica, vol. 16, no. 6 B, pp. 470-478.
In many applications, convection heat transfer is coupled with conduction and radiation heat transfer, which generate temperature gradients along the walls and may greatly affect natural convection heat transfer. The main objective of this study is to calculate the heat-transfer characteristics for natural convection from a non-isothermal vertical flat plate into a supercritical fluid. The influence of the non-uniformity of wall temperature on the heat transfer by natural convection along a vertical plate, having a linearly distributed temperature (characterized by the slope S) is also investigated. The thermal expansion coefficient is considered as a function of the temperature, the pressure, the van der Waals constants and the compressibility factor. The trends of the curves obtained with this equation and with values from tables of thermodynamic properties were similar and diverged at a critical point. These features confirmed the validity of this equation. Then, the governing systems of partial differential equations are solved numerically using the finite difference method. The local Nusselt number was then calculated and plotted as a function of the local Rayleigh number. It was observed that a positive slope of temperature distribution increases the heat transfer rate and a negative slope decreases it. © Sharif University of Technology, December 2009.
Polymeric micro-fabricated filters have excellent sieving properties. Their identical properties such as high surface porosity and perfectly patterned pore structure, which is combined with mechanical strength make them ideal for many applications such as microorganism removal, blood filtration and protein purification. To improve the performance of the micro-fabricated filters, we employed oxygen plasma treatment to increase the surface hydrophilicity and reduce the membrane fouling during microfiltration. Hydrophilization and integrity of the surfaces were analyzed by contact angle measurements and topographic imaging with an atomic force microscope (AFM). Treatment of polymeric membranes with oxygen plasma led to a stable hydrophilization and an increased surface roughness. The filtration properties of the modified and unmodified membranes were examined using clay particles. A significant increase in total collected volume of filtrate was observed for the treated membranes during filtration of simulated drinking water samples using clay suspension. © (2012) Trans Tech Publications, Switzerland.