Browsing by Author "Huang, Hong-Yuan"

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A highly efficient bead extraction technique with low bead number for digital microfluidic immunoassay
(2016-01)
Huang, Cheng-Yeh
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Tsai, Po-Yen
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Lee, I-Chin
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Hsu, Hsin-Yun
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Huang, Hong-Yuan
Here, we describe a technique to manipulate a low number of beads to achieve high washing efficiency with zero bead loss in the washing process of a digital microfluidic (DMF) immunoassay. Previously, two magnetic bead extraction methods were reported in the DMF platform: (1) single-side electrowetting method and (2) double-side electrowetting method. The first approach could provide high washing efficiency, but it required a large number of beads. The second approach could reduce the required number of beads, but it was inefficient where multiple washes were required. More importantly, bead loss during the washing process was unavoidable in both methods. Here, an improved double-side electrowetting method is proposed for bead extraction by utilizing a series of unequal electrodes. It is shown that, with proper electrode size ratio, only one wash step is required to achieve 98% washing rate without any bead loss at bead number less than 100 in a droplet. It allows using only about 25 magnetic beads in DMF immunoassay to increase the number of captured analytes on each bead effectively. In our human soluble tumor necrosis factor receptor I (sTNF-RI) model immunoassay, the experimental results show that, comparing to our previous results without using the proposed bead extraction technique, the immunoassay with low bead number significantly enhances the fluorescence signal to provide a better limit of detection (3.14 pg/ml) with smaller reagent volumes (200 nl) and shorter analysis time (<1 h). This improved bead extraction technique not only can be used in the DMF immunoassay but also has great potential to be used in any other bead-based DMF systems for different applications. (C) 2016 AIP Publishing LLC.
AMPFLUID: Aggregation Magnified Post-Assay Fluorescence for Ultrasensitive Immunodetection on Digital Microfluidics
(2015-02-01)
Huang, Cheng-Yeh
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Shih, Po-Huai
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Tsai, Po-Yen
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Lee, I-Chin
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Hsu, Hsin-Yun
Several strategies are currently employed to enhance the detection limit of bead-based assays, but all these approaches improve the sensitivity by varying the assay procedures chemically or biologically. In previous digital microfluidic setups for bead-based immunoassay, the magnetic beads were suspended for detection. We investigated the effect of bead aggregation in such an immunoassay system and propose the aggregation magnified post-assay fluorescence for ultrasensitive immunodetection on digital microfluidics (AMPFLUID). The detection signal and sensitivity are further enhanced even at the post-assay stage without altering the original assay protocol on employing magnetically triggered post-assay aggregation of beads in a digital microfluidic setup followed by processing of the fluorescent signal. This method is shown to enhance the fluorescent signal with increased consistency and sensitivity after appropriate charge-coupled device (CCD) calibration. This method of on-chip detection allows the fulfilment of consumption of a volume at the nanoliter level and a limit of detection in the range picogram/mL. In our sTNF-RI model immunoassay, only 2.5 nL of sample is required; a detection limit 15 pg/mL is achieved. The decreased uncertainty of the measure is indicated by the error bars and coefficient of variation.
Digital Microfluidic Dynamic Culture of Mammalian Embryos on an Electrowetting on Dielectric (EWOD) Chip
(2015-05-01)
Huang, Hong-Yuan
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Shen, Hsien-Hua
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Tien, Chang-Hung
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Li, Chin-Jung
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Fan, Shih-Kang
Current human fertilization in vitro (IVF) bypasses the female oviduct and manually inseminates, fertilizes and cultivates embryos in a static microdrop containing appropriate chemical compounds. A microfluidic microchannel system for IVF is considered to provide an improved in-vivo-mimicking environment to enhance the development in a culture system for an embryo before implantation. We demonstrate a novel digitalized microfluidic device powered with electrowetting on a dielectric (EWOD) to culture an embryo in vitro in a single droplet in a microfluidic environment to mimic the environment in vivo for development of the embryo and to culture the embryos with good development and live births. Our results show that the dynamic culture powered with EWOD can manipulate a single droplet containing one mouse embryo and culture to the blastocyst stage. The rate of embryo cleavage to a hatching blastocyst with a dynamic culture is significantly greater than that with a traditional static culture (p<0.05). The EWOD chip enhances the culture of mouse embryos in a dynamic environment. To test the reproductive outcome of the embryos collected from an EWOD chip as a culture system, we transferred embryos to pseudo-pregnant female mice and produced live births. These results demonstrate that an EWOD-based microfluidic device is capable of culturing mammalian embryos in a microfluidic biological manner, presaging future clinical application.
Dynamic Embryo Culture on a Digital Microfluidic Chip
(2012)
Li, Chun-Chao
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Hsu, Yao-Wen
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Fan, Shih-Kang
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Huang, Hong-Yuan
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交大名義發表
Dynamic Embryo Culture on a Digital Microfluidic Chip
(2012-01-01)
Li, Chun-Chao
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Hsu, Yao-Wen
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Huang, Hong-Yuan
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Fan, Shih-Kang
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材料科學與工程學系
Embryo culture on a digital microfluidic Chip is presented. The digital microfluidic chip consists of two parallel glass plates with conductive, dielectric, and hydrophobic coatings. ICR mouse embryos are cultivated in the culture media (KSOM + AA) droplets placed in a petri dish for static studies or between the parallel glass plates for dynamic studies. Embryo culture is successful with the conductive ITO, dielectric SU-8 2002, hydrophobic Teflon layers, and surfactant Pluronic F127 used in the digital microfluidic devices. Moreover, the blastocyst rate of the embryos cultivated in encapsulated droplets containing culture media core droplets and appropriate culture oil shells is comparable to that of the embryos cultivated in traditional petri dishes. Electrowetting-on-dielectric (EWOD) effectively actuates the investigated encapsulated droplets that will be used for dynamic embryo culture.
EMBRYO LAB CHIP TAKING ADVANTAGE OF MICROFLUIDICS AND CELL CO-CULTURING
(2015)
Liu, Cheng-Hsien
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Chang, Kuo-Wei
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Chang, Pei-Yu
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Sung, Yi-Jung
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Huang, Hong-Yuan
This research serves as an in vitro platform for fertilization and embryonic development. Thin polydimethylsiloxane ( PDMS) membrane was etched to create an array of through-holes, and sandwiched between two PDMS channels to form an upper chamber and lower chamber. This device integrates the concentration gradient design and perfusion system for proposed embryo coculture with stromal cells.
Fabrication of 3D Microfluidic Chip integrating Concentration gradient design and Perfusion system for Embryo coculture with Stromal cells
(2012-01-01)
Chang, Pei-Yu
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Chang, Kuo-Wei
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Lin, Ciou-Wen
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Shih, Po-Chen
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Liu, Cheng-Hsien
Fabrication of 3D chip that enables cell culture under continuous flow perfusion and repeated in situ observation by light microscopy is described. Thin polydimethylsiloxane PDMS) membrane was etched to create an array of through-holes, and sandwiched between two PDMS channels to form an upper chamber and lower chamber. Thus we fabricated a 3D microfluidic chip which integrates the concentration gradient design and perfusion system for proposed embryo coculture with stromal cells.
Innovation of Digitalized Microfluidic System with Electrowetting-on-Dielectric (EWOD) Chip Used in In Vitro Embryo Culture System.
(2012-03-01)
Huang, Hong-Yuan
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Yao, Da-Jeng
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Huang, Hung-Ju
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Fan, Shih-Kang
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Li, Chin-Jung
Simultaneous detection of two growth factors from human single-embryo culture medium by a bead-based digital microfluidic chip
(2020-02-15)
Lee, Meng-Shiue
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Hsu, Wensyang
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Huang, Hong-Yuan
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Tseng, Hsueh-Yang
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Lee, Chia-Tung
The measurement of growth factors released in a culture medium is considered to be an attractive non-invasive approach, apart from the embryo morphology, to identify the condition of an embryo development after fertilization in vitro (IVF), but the available embryo culture medium in the current method is only a few microlitres. This small sample volume, also of small concentration, makes difficult the application of a conventional detection method, such as an enzyme-linked immunosorbent assay (ELISA). A reliable detection of the growth factor from each embryo culture medium of such a small concentration hence remains a challenge. Here for the first time we report the results of measurement of not just one, but two, growth factors, human IL-1 beta and human TNF-alpha, from an individual droplet of embryo culture medium with a bead-based digital microfluidic chip. The required sample volume for a single measurement is only 520 nL; the total duration of the on-chip process is less than 40 min. Using the culture media of human embryos with normal morphologic features, we found that the concentrations of TNF-alpha change little from day 3 to day 5-6, but the concentrations of IL-1 beta for some embryos might double from day 3 to day 5-6. For other embryos even with similar normal morphologic features, some growth factors, such as IL-1 beta, might exhibit different expressions during the culture period. Those growth factors could serve to distinguish the development conditions of each embryo, not merely from an observation of embryo morphology.
Womb-on-a-chip biomimetic system for improved embryo culture and development
(2016-04-01)
Chang, Kuo-Wei
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Chang, Pei-Yu
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Huang, Hong-Yuan
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Li, Chin-Jung
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Tien, Chang-Hung
The 3D biomimetic microsystem described in this research integrates various functions of the fallopian tube to serve as an in vitro platform for fertilization and embryonic development. This device mimics the in vivo embryo-endometrial stromal cell monolayer coculture to enhance the group culture of embryos and mechanically culture the embryos in a dynamic manner to enhance their development. The microdevice presented here is compared with static culture for embryonic development rate in the presence and absence of epithelial cells. The results suggest that the dynamic biomimetic microsystem enhances the rate of embryo development, providing a culture environment that closely mimics the in vivo microenvironment. This all-in-one biomimetic microsystem has the potential to replace the present embryo culture platforms used for assisting in vitro fertilization. (C) 2015 Elsevier B.V. All rights reserved.