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Other Publications (46)
- Ultrasonics
- IEEE Transactions on Ultrasonics, Ferroelectrics, and Frequency Control
- IEEE Transactions on Ultrasonics, Ferroelectrics, and Frequency Control
- IEEE Transactions on Ultrasonics, Ferroelectrics, and Frequency Control
- Lab on a Chip
- Journal of Acquired Immune Deficiency Syndromes (1999)
- Lab on a Chip
- Lab on a Chip
- Lab on a Chip
- Lab on a Chip
- Lab on a Chip
- Lab on a Chip
- Biosensors & Bioelectronics
- Lab on a Chip
- Analytical and Bioanalytical Chemistry
- Biosensors & Bioelectronics
- Lab on a Chip
- Conference Proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference
- Integrative Biology : Quantitative Biosciences from Nano to Macro
- Tissue Engineering. Part C, Methods
- Advanced Drug Delivery Reviews
- Philosophical Transactions. Series A, Mathematical, Physical, and Engineering Sciences
- Biotechnology and Bioengineering
- Proceedings of the National Academy of Sciences of the United States of America
- Nanomedicine (London, England)
- Biotechnology Advances
- Physics of Fluids (Woodbury, N.Y. : 1994)
- Biotechnology Journal
- Biotechnology Journal
- Biotechnology Journal
- Tissue Engineering. Part C, Methods
- PloS One
- PloS One
- PloS One
- Lab on a Chip
- Biofabrication
- PloS One
- Biomicrofluidics
- PloS One
- Biomaterials
- Advanced Materials (Deerfield Beach, Fla.)
- Lab on a Chip
- Applied Physics Letters
- Nanomedicine (London, England)
- Lab on a Chip
- Nanomedicine (London, England)
Articles by Utkan Demirci in JoVE
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Droplets tarafından Başlığı Hücre Kapsülleme
Sangjun Moon1,2, Pei-Ann Lin1,2, Hasan Onur Keles1,2, Seung-Schick Yoo3, Utkan Demirci1,2,4
1Bio-Acoustic-MEMS Laboratory in Medicine (BAMM), HST-Center for Bioengineering, Brigham and Women's, Harvard Medical School, 2Bio-Acoustic-MEMS Laboratory in Medicine (BAMM), HST-Center for Bioengineering, Brigham and Women's Hospital, 3Brigham and Women's Hospital, Harvard Medical School, 4Harvard-MIT Division of Health Sciences and Technology, Massachusetts Institute of Technology; Center for Biomedical Engineering, Department of Medicine, Brigham and Women's Hospital
Other articles by Utkan Demirci on PubMed
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Capacitive Micromachined Ultrasonic Transducers: Next-generation Arrays for Acoustic Imaging?
IEEE Transactions on Ultrasonics, Ferroelectrics, and Frequency Control.
Nov, 2002 |
Pubmed ID: 12484483 Piezoelectric materials have dominated the ultrasonic transducer technology. Recently, capacitive micromachined ultrasonic transducers (CMUTs) have emerged as an alternative technology offering advantages such as wide bandwidth, ease of fabricating large arrays, and potential for integration with electronics. The aim of this paper is to demonstrate the viability of CMUTs for ultrasound imaging. We present the first pulse-echo phased array B-scan sector images using a 128-element, one-dimensional (1-D) linear CMUT array. We fabricated 64- and 128-element 1-D CMUT arrays with 100% yield and uniform element response across the arrays. These arrays have been operated in immersion with no failure or degradation in performance over the time. For imaging experiments, we built a resolution test phantom roughly mimicking the attenuation properties of soft tissue. We used a PC-based experimental system, including custom-designed electronic circuits to acquire the complete set of 128 x 128 RF A-scans from all transmit-receive element combinations. We obtained the pulse-echo frequency response by analyzing the echo signals from wire targets. These echo signals presented an 80% fractional bandwidth around 3 MHz, including the effect of attenuation in the propagating medium. We reconstructed the B-scan images with a sector angle of 90 degrees and an image depth of 210 mm through offline processing by using RF beamforming and synthetic phased array approaches. The measured 6-dB lateral and axial resolutions at 135 mm depth were 0.0144 radians and 0.3 mm, respectively. The electronic noise floor of the image was more than 50 dB below the maximum mainlobe magnitude. We also performed preliminary investigations on the effects of crosstalk among array elements on the image quality. In the near field, some artifacts were observable extending out from the array to a depth of 2 cm. A tail also was observed in the point spread function (PSF) in the axial direction, indicating the existence of crosstalk. The relative amplitude of this tail with respect to the mainlobe was less than -20 dB.
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Forward-viewing CMUT Arrays for Medical Imaging
IEEE Transactions on Ultrasonics, Ferroelectrics, and Frequency Control.
Jul, 2004 |
Pubmed ID: 15301009 This paper reports the design and testing of forward-viewing annular arrays fabricated using capacitive micromachined ultrasonic transducer (CMUT) technology. Recent research studies have shown that CMUTs have broad frequency bandwidth and high-transduction efficiency. One- and two-dimensional CMUT arrays of various sizes already have been fabricated, and their viability for medical imaging applications has been demonstrated. We fabricated 64-element, forward-viewing annular arrays using the standard CMUT fabrication process and carried out experiments to measure the operating frequency, bandwidth, and transmit/receive efficiency of the array elements. The annular array elements, designed for imaging applications in the 20 MHz range, had a resonance frequency of 13.5 MHz in air. The immersion pulse-echo data collected from a plane reflector showed that the devices operate in the 5-26 MHz range with a fractional bandwidth of 135%. The output pressure at the surface of the transducer was measured to be 24 kPa/V. These values translate into a dynamic range of 131.5 dB for 1-V excitation in 1-Hz bandwidth with a commercial low noise receiving circuitry. The designed, forward-viewing annular CMUT array is suitable for mounting on the front surface of a cylindrical catheter probe and can provide Doppler information for measurement of blood flow and guiding information for navigation through blood vessels in intravascular ultrasound imaging.
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Coherent Array Imaging Using Phased Subarrays. Part II: Simulations and Experimental Results
IEEE Transactions on Ultrasonics, Ferroelectrics, and Frequency Control.
Jan, 2005 |
Pubmed ID: 15742562 The basic principles and theory of phased subarray (PSA) imaging imaging provides the flexibility of reducing the number of front-end hardware channels between that of classical synthetic aperture (CSA) imaging--which uses only one element per firing event--and full-phased array (FPA) imaging-which uses all elements for each firing. The performance of PSA generally ranges between that obtained by CSA and FPA using the same array, and depends on the amount of hardware complexity reduction. For the work described in this paper, we performed FPA, CSA, and PSA imaging of a resolution phantom using both simulated and experimental data from a 3-MHz, 3.2-cm, 128-element capacitive micromachined ultrasound transducer (CMUT) array. The simulated system point responses in the spatial and frequency domains are presented as a means of studying the effects of signal bandwidth, reconstruction filter size, and subsampling rate on the PSA system performance. The PSA and FPA sector-scanned images were reconstructed using the wideband experimental data with 80% fractional bandwidth, with seven 32-element subarrays used for PSA imaging. The measurements on the experimental sector images indicate that, at the transmit focal zone, the PSA method provides a 10% improvement in the 6-dB lateral resolution, and the axial point resolution of PSA imaging is identical to that of FPA imaging. The signal-to-noise ratio (SNR) of PSA image was 58.3 dB, 4.9 dB below that of the FPA image, and the contrast-to-noise ratio (CNR) is reduced by 10%. The simulated and experimental test results presented in this paper validate theoretical expectations and illustrate the flexibility of PSA imaging as a way to exchange SNR and frame rate for simplified front-end hardware.
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A Microfluidic Device for Practical Label-free CD4(+) T Cell Counting of HIV-infected Subjects
Lab on a Chip.
Feb, 2007 |
Pubmed ID: 17268618 Practical HIV diagnostics are urgently needed in resource-limited settings. While HIV infection can be diagnosed using simple, rapid, lateral flow immunoassays, HIV disease staging and treatment monitoring require accurate counting of a particular white blood cell subset, the CD4(+) T lymphocyte. To address the limitations of current expensive, technically demanding and/or time-consuming approaches, we have developed a simple CD4 counting microfluidic device. This device uses cell affinity chromatography operated under differential shear flow to specifically isolate CD4(+) T lymphocytes with high efficiency directly from 10 microliters of unprocessed, unlabeled whole blood. CD4 counts are obtained under an optical microscope in a rapid, simple and label-free fashion. CD4 counts determined in our device matched measurements by conventional flow cytometry among HIV-positive subjects over a wide range of absolute CD4 counts (R(2) = 0.93). This CD4 counting microdevice can be used for simple, rapid and affordable CD4 counting in point-of-care and resource-limited settings.
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A Microchip Approach for Practical Label-free CD4+ T-cell Counting of HIV-infected Subjects in Resource-poor Settings
Journal of Acquired Immune Deficiency Syndromes (1999).
Jul, 2007 |
Pubmed ID: 17414933 Simple affordable CD4 cell counting is urgently needed to stage and monitor HIV-infected patients in resource-limited settings. To address the limitations of current approaches, we designed a simple, label-free, and cost-effective CD4 cell counting device using microfluidic technology. We previously described the fabrication of a microfluidic system for high-efficiency isolation of pure populations of CD4+ T cells based on cell affinity chromatography operated under controlled flow. Here, we compare the performance of a microfluidic CD4 cell counting device against standard flow cytometry in 49 HIV-positive subjects over a wide range of absolute CD4 cell counts. We observed a close correlation between CD4 cell counts from the microchip device and measurements by flow cytometry, using unprocessed whole blood from HIV-positive adult subjects. Sensitivities for distinguishing clinically relevant thresholds of 200, 350, and 500 cells/microL are 0.86, 0.90, and 0.97, respectively. Specificity is 0.94 or higher at all thresholds. This device can serve as a functional cartridge for fast, accurate, affordable, and simple CD4 cell counting in resource-limited settings.
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A Cell-laden Microfluidic Hydrogel
Lab on a Chip.
Jun, 2007 |
Pubmed ID: 17538718 The encapsulation of mammalian cells within the bulk material of microfluidic channels may be beneficial for applications ranging from tissue engineering to cell-based diagnostic assays. In this work, we present a technique for fabricating microfluidic channels from cell-laden agarose hydrogels. Using standard soft lithographic techniques, molten agarose was molded against a SU-8 patterned silicon wafer. To generate sealed and water-tight microfluidic channels, the surface of the molded agarose was heated at 71 degrees C for 3 s and sealed to another surface-heated slab of agarose. Channels of different dimensions were generated and it was shown that agarose, though highly porous, is a suitable material for performing microfluidics. Cells embedded within the microfluidic molds were well distributed and media pumped through the channels allowed the exchange of nutrients and waste products. While most cells were found to be viable upon initial device fabrication, only those cells near the microfluidic channels remained viable after 3 days, demonstrating the importance of a perfused network of microchannels for delivering nutrients and oxygen to maintain cell viability in large hydrogels. Further development of this technique may lead to the generation of biomimetic synthetic vasculature for tissue engineering, diagnostics, and drug screening applications.
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Single Cell Epitaxy by Acoustic Picolitre Droplets
Lab on a Chip.
Sep, 2007 |
Pubmed ID: 17713612 The capability to encapsulate single to few cells with micrometre precision, high viability, and controlled directionality via a nozzleless ejection technology using a gentle acoustic field would have great impact on tissue engineering, high throughput screening, and clinical diagnostics. We demonstrate encapsulation of single cells (or a few cells) ejected from an open pool in acoustic picolitre droplets. We have developed this technology for the specific purpose of printing cells in various biological fluids, including PBS and agarose hydrogels used in tissue engineering. We ejected various cell types, including mouse embryonic stem cells, fibroblasts, AML-12 hepatocytes, human Raji cells, and HL-1 cardiomyocytes encapsulated in acoustic picolitre droplets of around 37 microm in diameter at rates varying from 1 to 10,000 droplets per second. At such high throughput levels, we demonstrated cell viabilities of over 89.8% across various cell types. Moreover, this ejection method is readily adaptable to other biological applications, such as extracting data from single cells and generating large cell populations from single cells. The technique described in the current study may also be applied to investigate stem cell differentiation at the single cell level, to direct tissue printing, and to isolating pure RNA or DNA from a single cell at the picolitre level. Overall, the techniques described have the potential for widespread impact on many high-throughput testing applications in the biological and health sciences.
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Microcirculation Within Grooved Substrates Regulates Cell Positioning and Cell Docking Inside Microfluidic Channels
Lab on a Chip.
May, 2008 |
Pubmed ID: 18432345 Immobilization of cells inside microfluidic devices is a promising approach for enabling studies related to drug screening and cell biology. Despite extensive studies in using grooved substrates for immobilizing cells inside channels, a systematic study of the effects of various parameters that influence cell docking and retention within grooved substrates has not been performed. We demonstrate using computational simulations that the fluid dynamic environment within microgrooves significantly varies with groove width, generating microcirculation areas in smaller microgrooves. Wall shear stress simulation predicted that shear stresses were in the opposite direction in smaller grooves (25 and 50 microm wide) in comparison to those in wider grooves (75 and 100 microm wide). To validate the simulations, cells were seeded within microfluidic devices, where microgrooves of different widths were aligned perpendicularly to the direction of the flow. Experimental results showed that, as predicted, the inversion of the local direction of shear stress within the smaller grooves resulted in alignment of cells on two opposite sides of the grooves under the same flow conditions. Also, the amplitude of shear stress within microgrooved channels significantly influenced cell retainment in the channels. Therefore, our studies suggest that microscale shear stresses greatly influence cellular docking, immobilization, and retention in fluidic systems and should be considered for the design of cell-based microdevices.
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Rapid Automated Cell Quantification on HIV Microfluidic Devices
Lab on a Chip.
Dec, 2009 |
Pubmed ID: 19904402 Lab-chip device analysis often requires high throughput quantification of fluorescent cell images, obtained under different conditions of fluorescent intensity, illumination, focal depth, and optical magnification. Many laboratories still use manual counting--a tedious, expensive process prone to inter-observer variability. The manual counting process can be automated for fast and precise data gathering and reduced manual bias. We present a method to segment and count cells in microfluidic chips that are labeled with a single stain, or multiple stains, using image analysis techniques in Matlab and discuss its advantages over manual counting. Microfluidic based cell capturing devices for HIV monitoring were used to validate our method. Captured CD4(+) CD3(+) T lymphocytes were stained with DAPI, AF488-anti CD4, and AF647-anti CD3 for cell identification. Altogether 4788 (76 x 3 x 21) gray color images were obtained from devices using discarded 10 HIV infected patient whole blood samples (21 devices). We observed that the automatic method performs similarly to manual counting for a small number of cells. However, automated counting is more accurate and more than 100 times faster than manual counting for multiple-color stained cells, especially when large numbers of cells need to be quantified (>500 cells). The algorithm is fully automatic for subsequent microscope images that cover the full device area. It accounts for problems that generally occur in fluorescent lab-chip cell images such as: uneven background, overlapping cell images and cell detection with multiple stains. This method can be used in laboratories to save time and effort, and to increase cell counting accuracy of lab-chip devices for various applications, such as circulating tumor cell detection, cell detection in biosensors, and HIV monitoring devices, i.e. CD4 counts.
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Microscale Electroporation: Challenges and Perspectives for Clinical Applications
Integrative Biology : Quantitative Biosciences from Nano to Macro.
Mar, 2009 |
Pubmed ID: 20023735 Microscale engineering plays a significant role in developing tools for biological applications by miniaturizing devices and providing controllable microenvironments for in vitro cell research. Miniaturized devices offer numerous benefits in comparison to their macroscale counterparts, such as lower use of expensive reagents, biomimetic environments, and the ability to manipulate single cells. Microscale electroporation is one of the main beneficiaries of microscale engineering as it provides spatial and temporal control of various electrical parameters. Microscale electroporation devices can be used to reduce limitations associated with the conventional electroporation approaches such as variations in the local pH, electric field distortion, sample contamination, and the difficulties in transfecting and maintaining the viability of desired cell types. Here, we present an overview of recent advances of the microscale electroporation methods and their applications in biology, as well as current challenges for its use for clinical applications. We categorize microscale electroporation into microchannel and microcapillary electroporation. Microchannel-based electroporation can be used for transfecting cells within microchannels under dynamic flow conditions in a controlled and high-throughput fashion. In contrast, microcapillary-based electroporation can be used for transfecting cells within controlled reaction chambers under static flow conditions. Using these categories we examine the use of microscale electroporation for clinical applications related to HIV-1, stem cells, cancer and other diseases and discuss the challenges in further advancing this technology for use in clinical medicine and biology.
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Layer by Layer Three-dimensional Tissue Epitaxy by Cell-laden Hydrogel Droplets
Tissue Engineering. Part C, Methods.
Feb, 2010 |
Pubmed ID: 19586367 The ability to bioengineer three-dimensional (3D) tissues is a potentially powerful approach to treat diverse diseases such as cancer, loss of tissue function, or organ failure. Traditional tissue engineering methods, however, face challenges in fabricating 3D tissue constructs that resemble the native tissue microvasculature and microarchitectures. We have developed a bioprinter that can be used to print 3D patches of smooth muscle cells (5 mm x 5 mm x 81 microm) encapsulated within collagen. Current inkjet printing systems suffer from loss of cell viability and clogging. To overcome these limitations, we developed a system that uses mechanical valves to print high viscosity hydrogel precursors containing cells. The bioprinting platform that we developed enables (i) printing of multilayered 3D cell-laden hydrogel structures (16.2 microm thick per layer) with controlled spatial resolution (proximal axis: 18.0 +/- 7.0 microm and distal axis: 0.5 +/- 4.9 microm), (ii) high-throughput droplet generation (1 s per layer, 160 droplets/s), (iii) cell seeding uniformity (26 +/- 2 cells/mm(2) at 1 million cells/mL, 122 +/- 20 cells/mm(2) at 5 million cells/mL, and 216 +/- 38 cells/mm(2) at 10 million cells/mL), and (iv) long-term viability in culture (>90%, 14 days). This platform to print 3D tissue constructs may be beneficial for regenerative medicine applications by enabling the fabrication of printed replacement tissues.
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Microporous Cell-laden Hydrogels for Engineered Tissue Constructs
Biotechnology and Bioengineering.
May, 2010 |
Pubmed ID: 20091766 In this article, we describe an approach to generate microporous cell-laden hydrogels for fabricating biomimetic tissue engineered constructs. Micropores at different length scales were fabricated in cell-laden hydrogels by micromolding fluidic channels and leaching sucrose crystals. Microengineered channels were created within cell-laden hydrogel precursors containing agarose solution mixed with sucrose crystals. The rapid cooling of the agarose solution was used to gel the solution and form micropores in place of the sucrose crystals. The sucrose leaching process generated homogeneously distributed micropores within the gels, while enabling the direct immobilization of cells within the gels. We also characterized the physical, mechanical, and biological properties (i.e., microporosity, diffusivity, and cell viability) of cell-laden agarose gels as a function of engineered porosity. The microporosity was controlled from 0% to 40% and the diffusivity of molecules in the porous agarose gels increased as compared to controls. Furthermore, the viability of human hepatic carcinoma cells that were cultured in microporous agarose gels corresponded to the diffusion profile generated away from the microchannels. Based on their enhanced diffusive properties, microporous cell-laden hydrogels containing a microengineered fluidic channel can be a useful tool for generating tissue structures for regenerative medicine and drug discovery applications.
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Vitrification and Levitation of a Liquid Droplet on Liquid Nitrogen
Proceedings of the National Academy of Sciences of the United States of America.
Mar, 2010 |
Pubmed ID: 20176969 The vitrification of a liquid occurs when ice crystal formation is prevented in the cryogenic environment through ultrarapid cooling. In general, vitrification entails a large temperature difference between the liquid and its surrounding medium. In our droplet vitrification experiments, we observed that such vitrification events are accompanied by a Leidenfrost phenomenon, which impedes the heat transfer to cool the liquid, when the liquid droplet comes into direct contact with liquid nitrogen. This is distinct from the more generally observed Leidenfrost phenomenon that occurs when a liquid droplet is self-vaporized on a hot plate. In the case of rapid cooling, the phase transition from liquid to vitrified solid (i.e., vitrification) and the levitation of droplets on liquid nitrogen (i.e., Leidenfrost phenomenon) take place simultaneously. Here, we investigate these two simultaneous physical events by using a theoretical model containing three dimensionless parameters (i.e., Stefan, Biot, and Fourier numbers). We explain theoretically and observe experimentally a threshold droplet radius during the vitrification of a cryoprotectant droplet in the presence of the Leidenfrost effect.
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Advances in Developing HIV-1 Viral Load Assays for Resource-limited Settings
Biotechnology Advances.
Nov-Dec, 2010 |
Pubmed ID: 20600784 Commercial HIV-1 RNA viral load assays have been routinely used in developed countries to monitor antiretroviral treatment (ART). However, these assays require expensive equipment and reagents, well-trained operators, and established laboratory infrastructure. These requirements restrict their use in resource-limited settings where people are most afflicted with the HIV-1 epidemic. Inexpensive alternatives such as the Ultrasensitive p24 assay, the reverse transcriptase (RT) assay and in-house reverse transcription quantitative polymerase chain reaction (RT-qPCR) have been developed. However, they are still time-consuming, technologically complex and inappropriate for decentralized laboratories as point-of-care (POC) tests. Recent advances in microfluidics and nanotechnology offer new strategies to develop low-cost, rapid, robust and simple HIV-1 viral load monitoring systems. We review state-of-the-art technologies used for HIV-1 viral load monitoring in both developed and developing settings. Emerging approaches based on microfluidics and nanotechnology, which have potential to be integrated into POC HIV-1 viral load assays, are also discussed.
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A Three-dimensional in Vitro Ovarian Cancer Coculture Model Using a High-throughput Cell Patterning Platform
Biotechnology Journal.
Feb, 2011 |
Pubmed ID: 21298805 In vitro 3D cancer models that provide a more accurate representation of disease in vivo are urgently needed to improve our understanding of cancer pathology and to develop better cancer therapies. However, development of 3D models that are based on manual ejection of cells from micropipettes suffer from inherent limitations such as poor control over cell density, limited repeatability, low throughput, and, in the case of coculture models, lack of reproducible control over spatial distance between cell types (e.g., cancer and stromal cells). In this study, we build on a recently introduced 3D model in which human ovarian cancer (OVCAR-5) cells overlaid on Matrigelâ„¢ spontaneously form multicellular acini. We introduce a high-throughput automated cell printing system to bioprint a 3D coculture model using cancer cells and normal fi broblasts micropatterned on Matrigelâ„¢ . Two cell types were patterned within a spatially controlled microenvironment (e.g., cell density, cell-cell distance) in a high-throughput and reproducible manner; both cell types remained viable during printing and continued to proliferate following patterning. This approach enables the miniaturization of an established macro-scale 3D culture model and would allow systematic investigation into the multiple unknown regulatory feedback mechanisms between tumor and stromal cells and provide a tool for high-throughput drug screening.
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Automated and Adaptable Quantification of Cellular Alignment from Microscopic Images for Tissue Engineering Applications
Tissue Engineering. Part C, Methods.
Jun, 2011 |
Pubmed ID: 21370940 Cellular alignment plays a critical role in functional, physical, and biological characteristics of many tissue types, such as muscle, tendon, nerve, and cornea. Current efforts toward regeneration of these tissues include replicating the cellular microenvironment by developing biomaterials that facilitate cellular alignment. To assess the functional effectiveness of the engineered microenvironments, one essential criterion is quantification of cellular alignment. Therefore, there is a need for rapid, accurate, and adaptable methodologies to quantify cellular alignment for tissue engineering applications. To address this need, we developed an automated method, binarization-based extraction of alignment score (BEAS), to determine cell orientation distribution in a wide variety of microscopic images. This method combines a sequenced application of median and band-pass filters, locally adaptive thresholding approaches and image processing techniques. Cellular alignment score is obtained by applying a robust scoring algorithm to the orientation distribution. We validated the BEAS method by comparing the results with the existing approaches reported in literature (i.e., manual, radial fast Fourier transform-radial sum, and gradient based approaches). Validation results indicated that the BEAS method resulted in statistically comparable alignment scores with the manual method (coefficient of determination R(2)=0.92). Therefore, the BEAS method introduced in this study could enable accurate, convenient, and adaptable evaluation of engineered tissue constructs and biomaterials in terms of cellular alignment and organization.
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Lensless Imaging for Simultaneous Microfluidic Sperm Monitoring and Sorting
Lab on a Chip.
Aug, 2011 |
Pubmed ID: 21677993 5.3 million American couples of reproductive age (9%) are affected by infertility, among which male factors account for up to 50% of cases, which necessitates the identification of parameters defining sperm quality, including sperm count and motility. In vitro fertilization (IVF) with or without intra cytoplasmic sperm injection (ICSI) has become the most widely used assisted reproductive technology (ART) in modern clinical practice to overcome male infertility challenges. One of the obstacles of IVF and ICSI lies in identifying and isolating the most motile and presumably healthiest sperm from semen samples that have low sperm counts (oligozoospermia) and/or low sperm motility (oligospermaesthenia). Microfluidic systems have shown potential to sort sperm with flow systems. However, the small field of view (FOV) of conventional microscopes commonly used to image sperm motion presents challenges in tracking a large number of sperm cells simultaneously. To address this challenge, we have integrated a lensless charge-coupled device (CCD) with a microfluidic chip to enable wide FOV and automatic recording as the sperm move inside a microfluidic channel. The integrated system enables the sorting and tracking of a population of sperm that have been placed in a microfluidic channel. This channel can be monitored in both horizontal and vertical configuration similar to a swim-up column method used clinically. Sperm motilities can be quantified by tracing the shadow paths for individual sperm. Moreover, as the sperm are sorted by swimming from the inlet towards the outlet of a microfluidic channel, motile sperm that reach the outlet can be extracted from the channel at the end of the process. This technology can lead to methods to evaluate each sperm individually in terms of motility response in a wide field of view, which could prove especially useful, when working with oligozoospermic or oligospermaesthenic samples, in which the most motile sperm need to be isolated from a pool of small number of sperm.
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Microengineering Methods for Cell-based Microarrays and High-throughput Drug-screening Applications
Biofabrication.
Sep, 2011 |
Pubmed ID: 21725152 Screening for effective therapeutic agents from millions of drug candidates is costly, time consuming, and often faces concerns due to the extensive use of animals. To improve cost effectiveness, and to minimize animal testing in pharmaceutical research, in vitro monolayer cell microarrays with multiwell plate assays have been developed. Integration of cell microarrays with microfluidic systems has facilitated automated and controlled component loading, significantly reducing the consumption of the candidate compounds and the target cells. Even though these methods significantly increased the throughput compared to conventional in vitro testing systems and in vivo animal models, the cost associated with these platforms remains prohibitively high. Besides, there is a need for three-dimensional (3D) cell-based drug-screening models which can mimic the in vivo microenvironment and the functionality of the native tissues. Here, we present the state-of-the-art microengineering approaches that can be used to develop 3D cell-based drug-screening assays. We highlight the 3D in vitro cell culture systems with live cell-based arrays, microfluidic cell culture systems, and their application to high-throughput drug screening. We conclude that among the emerging microengineering approaches, bioprinting holds great potential to provide repeatable 3D cell-based constructs with high temporal, spatial control and versatility.
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The Assembly of Cell-encapsulating Microscale Hydrogels Using Acoustic Waves
Biomaterials.
Nov, 2011 |
Pubmed ID: 21820734 Microscale hydrogels find widespread applications in medicine and biology, e.g., as building blocks for tissue engineering and regenerative medicine. In these applications, these microgels are assembled to fabricate large complex 3D constructs. The success of this approach requires non-destructive and high throughput assembly of the microgels. Although various assembly methods have been developed based on modifying interfaces, and using microfluidics, so far, none of the available assembly technologies have shown the ability to assemble microgels using non-invasive fields rapidly within seconds in an efficient way. Acoustics has been widely used in biomedical arena to manipulate droplets, cells and biomolecules. In this study, we developed a simple, non-invasive acoustic assembler for cell-encapsulating microgels with maintained cell viability (>93%). We assessed the assembler for both microbeads (with diameter of 50 μm and 100 μm) and microgels of different sizes and shapes (e.g., cubes, lock-and-key shapes, tetris, saw) in microdroplets (with volume of 10 μL, 20 μL, 40 μL, 80 μL). The microgels were assembled in seconds in a non-invasive manner. These results indicate that the developed acoustic approach could become an enabling biotechnology tool for tissue engineering, regenerative medicine, pharmacology studies and high throughput screening applications.
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Integration of Cell Phone Imaging with Microchip ELISA to Detect Ovarian Cancer HE4 Biomarker in Urine at the Point-of-care
Lab on a Chip.
Oct, 2011 |
Pubmed ID: 21881677 Ovarian cancer is asymptomatic in the early stages and most patients present with advanced levels of disease. The lack of cost-effective methods that can achieve frequent, simple and non-invasive testing hinders early detection and causes high mortality in ovarian cancer patients. Here, we report a simple and inexpensive microchip ELISA-based detection module that employs a portable detection system, i.e., a cell phone/charge-coupled device (CCD) to quantify an ovarian cancer biomarker, HE4, in urine. Integration of a mobile application with a cell phone enabled immediate processing of microchip ELISA results, which eliminated the need for a bulky, expensive spectrophotometer. The HE4 level detected by a cell phone or a lensless CCD system was significantly elevated in urine samples from cancer patients (n = 19) than healthy controls (n = 20) (p < 0.001). Receiver operating characteristic (ROC) analyses showed that the microchip ELISA coupled with a cell phone running an automated analysis mobile application had a sensitivity of 89.5% at a specificity of 90%. Under the same specificity, the microchip ELISA coupled with a CCD had a sensitivity of 84.2%. In conclusion, integration of microchip ELISA with cell phone/CCD-based colorimetric measurement technology can be used to detect HE4 biomarker at the point-of-care (POC), paving the way to create bedside technologies for diagnostics and treatment monitoring.
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Controlled Viable Release of Selectively Captured Label-free Cells in Microchannels
Lab on a Chip.
Dec, 2011 |
Pubmed ID: 22002065 Selective capture of cells from bodily fluids in microchannels has broadly transformed medicine enabling circulating tumor cell isolation, rapid CD4(+) cell counting for HIV monitoring, and diagnosis of infectious diseases. Although cell capture methods have been demonstrated in microfluidic systems, the release of captured cells remains a significant challenge. Viable retrieval of captured label-free cells in microchannels will enable a new era in biological sciences by allowing cultivation and post-processing. The significant challenge in release comes from the fact that the cells adhere strongly to the microchannel surface, especially when immuno-based immobilization methods are used. Even though fluid shear and enzymes have been used to detach captured cells in microchannels, these methods are known to harm cells and affect cellular characteristics. This paper describes a new technology to release the selectively captured label-free cells in microchannels without the use of fluid shear or enzymes. We have successfully released the captured CD4(+) cells (3.6% of the mononuclear blood cells) from blood in microfluidic channels with high specificity (89% ± 8%), viability (94% ± 4%), and release efficiency (59% ± 4%). We have further validated our system by specifically capturing and controllably releasing the CD34(+) stem cells from whole blood, which were quantified to be 19 cells per million blood cells in the blood samples used in this study. Our results also indicated that both CD4(+) and CD34(+) cells released from the microchannels were healthy and amenable for in vitro culture. Manual flow based microfluidic method utilizes inexpensive, easy to fabricate microchannels allowing selective label-free cell capture and release in less than 10 minutes, which can also be used at the point-of-care. The presented technology can be used to isolate and purify a broad spectrum of cells from mixed populations offering widespread applications in applied biological sciences, such as tissue engineering, regenerative medicine, rare cell and stem cell isolation, proteomic/genomic research, and clonal/population analyses.
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Nanoliter Droplet Vitrification for Oocyte Cryopreservation
Nanomedicine (London, England).
Dec, 2011 |
Pubmed ID: 22188180 Aim: Oocyte cryopreservation remains largely experimental, with live birth rates of only 2-4% per thawed oocyte. In this study, we present a nanoliter droplet technology for oocyte vitrification. Materials & methods: An ejector-based droplet vitrification system was designed to continuously cryopreserve oocytes in nanoliter droplets. Oocyte survival rates, morphologies and parthenogenetic development after each vitrification step were assessed in comparison with fresh oocytes. Results: Oocytes were retrieved after cryoprotectant agent loading/unloading, and nanoliter droplet encapsulation showed comparable survival rates to fresh oocytes after 24 h in culture. Also, oocytes recovered after vitrification/thawing showed similar morphologies to those of fresh oocytes. Additionally, the rate of oocyte parthenogenetic activation after nanoliter droplet encapsulation was comparable with that observed for fresh oocytes. This nanoliter droplet technology enables the vitrification of oocytes at higher cooling and warming rates using lower cryoprotectant agent levels (i.e., 1.4 M ethylene glycol, 1.1 M dimethyl sulfoxide and 1 M sucrose), thus making it a potential technology to improve oocyte cryopreservation outcomes.
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