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黃正鄴 (Cheng-Yeh Huang)
黃正鄴 (Cheng-Yeh Huang)
E-mail: chengyeh.huang [at] gmail.com
Website: https://chengyehhuang.wordpress.com/
Development of digital microfluidic immunochip for detecting low level of biomarkers
Development of digital microfluidic immunochip for detecting low level of biomarkers
可偵測低濃度生物標誌之數位微流體免疫檢測晶片
可偵測低濃度生物標誌之數位微流體免疫檢測晶片
This thesis describes the development of a digital microfluidic immunochip for detecting low level of biomarkers in an ultra-low volume sample. Two new techniques on a digital microfluidic (DMF) setup are developed for performing immunoassays. The first technique enhances the detection signal and sensitivity even at the post-assay stage without altering the original assay procedures chemically or biologically by employing magnetically triggered post-assay aggregation of beads in a DMF setup. The effect of bead aggregation in such an immunoassay system is systematically investigated. Bead aggregation is shown to stably enhance the fluorescence intensity in DMF device with a proper gap distance. In our soluble tumour necrosis factor receptor I (sTNF-RI) model immunoassay, limit of detection (LOD) 15 pg/mL and coefficient of variation (CV) 3% are achieved. The second technique enables the manipulation of low number of beads to provide stronger fluorescence signal per bead with high washing efficiency by utilizing a series of unequal electrodes. The result shows that only one wash step is required to achieve 98% washing rate without any bead loss at bead number less than 100 in a droplet. Decreasing the starting number of antibody coupled beads in immunoassay is shown to effectively elevate the responding signal, since less starting bead number would allow more antigens to bind to the surface of each bead. Therefore, by combining both techniques, our sTNF-RI model DMF immunoassay with LOD 3.14 pg/mL is achieved, and only 200 nL sample volume and less than 1 hour of analysis time are needed. This DMF immunoassay platform hence allows a small consumption of sample volume at a nanoliter level to achieve low limit of detection at the pg/mL level. Another aspect demonstrated is a decreased uncertainty. It may aid in some clinical diagnostics by providing modified assays with lower required sample and LODs to give insights into disease biology. Especially for in vitro fertilization (IVF), the sample quantity of embryo culture medium (5-10 μL) and concentration of biomarkers (below several tens of pg/mL) are all very low, which is hard to use conventional immunoassay to detect. The developed DMF immunoassay can act as a powerful embryo immuno-detection chip to detect and investigate different embryonic growth factors during embryo culture in IVF. It is hoped that the proposed embryo immuno-detection chip can provide quantitative evaluation of cultured embryos to alter the current morphology-based embryo selection, and reduce the implanted embryo number with good pregnancy rate and delivery rate.
This thesis describes the development of a digital microfluidic immunochip for detecting low level of biomarkers in an ultra-low volume sample. Two new techniques on a digital microfluidic (DMF) setup are developed for performing immunoassays. The first technique enhances the detection signal and sensitivity even at the post-assay stage without altering the original assay procedures chemically or biologically by employing magnetically triggered post-assay aggregation of beads in a DMF setup. The effect of bead aggregation in such an immunoassay system is systematically investigated. Bead aggregation is shown to stably enhance the fluorescence intensity in DMF device with a proper gap distance. In our soluble tumour necrosis factor receptor I (sTNF-RI) model immunoassay, limit of detection (LOD) 15 pg/mL and coefficient of variation (CV) 3% are achieved. The second technique enables the manipulation of low number of beads to provide stronger fluorescence signal per bead with high washing efficiency by utilizing a series of unequal electrodes. The result shows that only one wash step is required to achieve 98% washing rate without any bead loss at bead number less than 100 in a droplet. Decreasing the starting number of antibody coupled beads in immunoassay is shown to effectively elevate the responding signal, since less starting bead number would allow more antigens to bind to the surface of each bead. Therefore, by combining both techniques, our sTNF-RI model DMF immunoassay with LOD 3.14 pg/mL is achieved, and only 200 nL sample volume and less than 1 hour of analysis time are needed. This DMF immunoassay platform hence allows a small consumption of sample volume at a nanoliter level to achieve low limit of detection at the pg/mL level. Another aspect demonstrated is a decreased uncertainty. It may aid in some clinical diagnostics by providing modified assays with lower required sample and LODs to give insights into disease biology. Especially for in vitro fertilization (IVF), the sample quantity of embryo culture medium (5-10 μL) and concentration of biomarkers (below several tens of pg/mL) are all very low, which is hard to use conventional immunoassay to detect. The developed DMF immunoassay can act as a powerful embryo immuno-detection chip to detect and investigate different embryonic growth factors during embryo culture in IVF. It is hoped that the proposed embryo immuno-detection chip can provide quantitative evaluation of cultured embryos to alter the current morphology-based embryo selection, and reduce the implanted embryo number with good pregnancy rate and delivery rate.
Journal papers
Journal papers
1. C.-Y. Huang, P.-Y. Tsai, I-C. Lee, H.-Y. Hsu, H.-Y. Huang, S.-K. Fan, D.-J. Yao, C.-H. Liu, W. Hsu, "A highly efficient bead extraction technique with low bead number for digital microfluidic immunoassay," Biomicrofluidics 2016, 10, 011901. (IF=3.357, N/M=3/31)
1. C.-Y. Huang, P.-Y. Tsai, I-C. Lee, H.-Y. Hsu, H.-Y. Huang, S.-K. Fan, D.-J. Yao, C.-H. Liu, W. Hsu, "A highly efficient bead extraction technique with low bead number for digital microfluidic immunoassay," Biomicrofluidics 2016, 10, 011901. (IF=3.357, N/M=3/31)
2. C.-Y. Huang, P. Sun, M.-S. Lee, S.-Y. Wu, Y.-C. Shieh, W. Hsu, "A resettable, wireless and passive fall-down recorder using a magnetic droplet with an LC circuit," IEEE Sensors Journal 2016, 16, 654-661. (IF=1.762, N/M=77/249)
2. C.-Y. Huang, P. Sun, M.-S. Lee, S.-Y. Wu, Y.-C. Shieh, W. Hsu, "A resettable, wireless and passive fall-down recorder using a magnetic droplet with an LC circuit," IEEE Sensors Journal 2016, 16, 654-661. (IF=1.762, N/M=77/249)
3. C.-Y. Huang, P.-H. Shih, P.-Y. Tsai, I-C. Lee, H.-Y. Hsu, H.-Y. Huang, S.-K. Fan, and W. Hsu, "AMPFLUID: Aggregation magnified post-assay fluorescence for ultrasensitive immunodetection on digital microfluidics." Proceedings of the IEEE 2015, 103, 225-235. (IF=5.466, N/M=6/248)
3. C.-Y. Huang, P.-H. Shih, P.-Y. Tsai, I-C. Lee, H.-Y. Hsu, H.-Y. Huang, S.-K. Fan, and W. Hsu, "AMPFLUID: Aggregation magnified post-assay fluorescence for ultrasensitive immunodetection on digital microfluidics." Proceedings of the IEEE 2015, 103, 225-235. (IF=5.466, N/M=6/248)
International conference papers
1. Y.-B. Wang, M.-S. Lee, J.-H. Huang, C.-Y. Huang, W. Hsu, and Y.-Y. Tu, "An EWOD mixer with high flexibility on dilution ratio," in The 41st International Conference on Micro and Nano Engineering (MNE 2015), The Hague, The Netherlands, September 2015.
1. Y.-B. Wang, M.-S. Lee, J.-H. Huang, C.-Y. Huang, W. Hsu, and Y.-Y. Tu, "An EWOD mixer with high flexibility on dilution ratio," in The 41st International Conference on Micro and Nano Engineering (MNE 2015), The Hague, The Netherlands, September 2015.
2. Y.-H. Chung, C.-Y. Huang, P.-Y. Tsai, H.-Y. Hsu, and W. Hsu, "Investigation of three-dimension bead aggregation effect for optical sensing in digital microfluidics platform,” in The 5th International Conference on Optofluidics (Optofluidics 2015), Taipei, Taiwan, July 2015. (Oral)
2. Y.-H. Chung, C.-Y. Huang, P.-Y. Tsai, H.-Y. Hsu, and W. Hsu, "Investigation of three-dimension bead aggregation effect for optical sensing in digital microfluidics platform,” in The 5th International Conference on Optofluidics (Optofluidics 2015), Taipei, Taiwan, July 2015. (Oral)
3. H.-W. Tsai, C.-Y. Huang, C.-H. Chen, and W. Hsu, "Durability investigation of hydrophobic films for digital microfluidics," in The 5th International Conference on Optofluidics (Optofluidics 2015), Taipei, Taiwan, July 2015.
3. H.-W. Tsai, C.-Y. Huang, C.-H. Chen, and W. Hsu, "Durability investigation of hydrophobic films for digital microfluidics," in The 5th International Conference on Optofluidics (Optofluidics 2015), Taipei, Taiwan, July 2015.
4. J.-H. Huang, H.-W. Tsai, Y.-B. Wang, C.-Y. Huang, C.-H. Chen, and W. Hsu, "Volume analysis of creating droplets on different hydrophobic materials in digital microfluidics platform," in The 5th International Conference on Optofluidics (Optofluidics 2015), Taipei, Taiwan, July 2015.
4. J.-H. Huang, H.-W. Tsai, Y.-B. Wang, C.-Y. Huang, C.-H. Chen, and W. Hsu, "Volume analysis of creating droplets on different hydrophobic materials in digital microfluidics platform," in The 5th International Conference on Optofluidics (Optofluidics 2015), Taipei, Taiwan, July 2015.
5. Y.-B. Wang, M.-H. Siao, C.-Y. Huang, and W. Hsu, "A novel inclined lipid bilayer formation based on DIB method," in The 5th International Conference on Optofluidics (Optofluidics 2015), Taipei, Taiwan, July 2015. (Best Poster Paper Finalist)
5. Y.-B. Wang, M.-H. Siao, C.-Y. Huang, and W. Hsu, "A novel inclined lipid bilayer formation based on DIB method," in The 5th International Conference on Optofluidics (Optofluidics 2015), Taipei, Taiwan, July 2015. (Best Poster Paper Finalist)
6. M.-H. Siao, H.-Y. Lin, C.-Y. Huang, W. Hsu, "Lipid bilayer formation by moving magnetic droplet with magnetic force," in The 8th IEEE International Conference on Nano/Molecular Medicine and Engineering (IEEE-NANOMED 2014), Kaohsiung, Taiwan, November 2014.
6. M.-H. Siao, H.-Y. Lin, C.-Y. Huang, W. Hsu, "Lipid bilayer formation by moving magnetic droplet with magnetic force," in The 8th IEEE International Conference on Nano/Molecular Medicine and Engineering (IEEE-NANOMED 2014), Kaohsiung, Taiwan, November 2014.
7. Y.-H. Chung, P.-Y. Tsai, C.-Y. Huang, H.-Y. Hsu, and W. Hsu, "Effects of gap size and bead aggregation on fluorescence detection intensity in digital microfluidic immunoassay," in The 40th International Conference on Micro and Nano Engineering (MNE 2014), Lausanne, Switzerland, September 2014.
7. Y.-H. Chung, P.-Y. Tsai, C.-Y. Huang, H.-Y. Hsu, and W. Hsu, "Effects of gap size and bead aggregation on fluorescence detection intensity in digital microfluidic immunoassay," in The 40th International Conference on Micro and Nano Engineering (MNE 2014), Lausanne, Switzerland, September 2014.
8. C.-Y. Huang, P.-H. Shih, P.-Y. Tsai, I-C. Lee, H.-Y. Hsu, S.-K. Fan, and W. Hsu, "Enhanced fluorescence detection through post-immunoassay beads aggregation on bead-based immunoassay using digital microfluidics," in The 9th Annual IEEE International Conference on Nano/Micro Engineered and Molecular Systems (IEEE-NEMS 2014), Hawaii, USA, April 2014. (Oral)
8. C.-Y. Huang, P.-H. Shih, P.-Y. Tsai, I-C. Lee, H.-Y. Hsu, S.-K. Fan, and W. Hsu, "Enhanced fluorescence detection through post-immunoassay beads aggregation on bead-based immunoassay using digital microfluidics," in The 9th Annual IEEE International Conference on Nano/Micro Engineered and Molecular Systems (IEEE-NEMS 2014), Hawaii, USA, April 2014. (Oral)
9. P.-Y. Tsai, C.-Y. Huang, H.-Y. Hsu, and W. Hsu, "Low bead number washing for bead-based immunoassay using double sided EWOD and magnetic force," in The 9th Annual IEEE International Conference on Nano/Micro Engineered and Molecular Systems (IEEE-NEMS 2014), Hawaii, USA, April 2014. (Oral)
9. P.-Y. Tsai, C.-Y. Huang, H.-Y. Hsu, and W. Hsu, "Low bead number washing for bead-based immunoassay using double sided EWOD and magnetic force," in The 9th Annual IEEE International Conference on Nano/Micro Engineered and Molecular Systems (IEEE-NEMS 2014), Hawaii, USA, April 2014. (Oral)
10. C.-Y. Huang, M.-Y. Chiang, S.-K. Fan, A. M. Ghaemmaghami, and W. Hsu, "Label-free separation and sorting of human monocytes and T-cells by electrowetting and dielectrophoresis," in The 8th Annual IEEE International Conference on Nano/Micro Engineered and Molecular Systems (IEEE-NEMS 2013), Suzhou, China, April 2013. (Oral)
10. C.-Y. Huang, M.-Y. Chiang, S.-K. Fan, A. M. Ghaemmaghami, and W. Hsu, "Label-free separation and sorting of human monocytes and T-cells by electrowetting and dielectrophoresis," in The 8th Annual IEEE International Conference on Nano/Micro Engineered and Molecular Systems (IEEE-NEMS 2013), Suzhou, China, April 2013. (Oral)
11. T.-M. Chuo, P.-H. Shih, W. Hsu, C.-Y. Huang, Y.-S. Huang, Y.-T. Chen, and S.-K. Fan, "Signal enhancement by particle concentration using dielectrophoresis in an electrowetting-driven droplet," in The 6th IEEE International Conference on Nano/Molecular Medicine and Engineering (IEEE-NANOMED 2012), Bangkok, Thailand, November 2012. (Oral, Best Student Paper Finalist)
12. C.-Y. Huang, S.-K. Fan, and W. Hsu, "Capillary electrophoresis in virtual microchannel based on dielectrophoresis," in The 15th International Conference on Miniaturized Systems for Chemistry and Life Sciences (MicroTAS 2011), Seattle, USA, September 2011.
13. C.-Y. Huang, S.-K. Fan, and W. Hsu, "Electroosmotic flow control in a virtual microchannel based on liquid dielectrophoresis" in The 6th Annual IEEE International Conference on Nano/Micro Engineered and Molecular Systems (IEEE NEMS 2011), Kaohsiung, Taiwan, February 2011. (Oral)
14. H.-Y. Hu, C.-Y. Huang, S.-K. Fan, and W. Hsu, "Electroosmotic flow in dielectrophoresis-formed microchannel," in The 3rd IEEE International Conference on Nano/Molecular Medicine and Engineering (IEEE-NANOMED 2009), Tainan, Taiwan, October 2009. (Oral)
14. H.-Y. Hu, C.-Y. Huang, S.-K. Fan, and W. Hsu, "Electroosmotic flow in dielectrophoresis-formed microchannel," in The 3rd IEEE International Conference on Nano/Molecular Medicine and Engineering (IEEE-NANOMED 2009), Tainan, Taiwan, October 2009. (Oral)
Domestic conference paper
Domestic conference paper
1. C.-Y. Huang, H.-Y. Hu, S.-K. Fan, and W. Hsu, "General digital microfluidic platform: capillary electrophoresis-based separation of micro-particle in electric-field-controlled virtual microchannel," in Taiwan Scientific Symposium, Birmingham, UK. (Invited talk)
1. C.-Y. Huang, H.-Y. Hu, S.-K. Fan, and W. Hsu, "General digital microfluidic platform: capillary electrophoresis-based separation of micro-particle in electric-field-controlled virtual microchannel," in Taiwan Scientific Symposium, Birmingham, UK. (Invited talk)
Patents
Patents
1. 徐文祥,黄正鄴,蔡泊諺,施博懷,范士岡,饒達仁,劉承賢,黃泓淵. 磁珠式數位微流體免疫分析裝置與其方法, 中華民國專利證號: 發明 I542879, 民國105年7月21日.
1. 徐文祥,黄正鄴,蔡泊諺,施博懷,范士岡,饒達仁,劉承賢,黃泓淵. 磁珠式數位微流體免疫分析裝置與其方法, 中華民國專利證號: 發明 I542879, 民國105年7月21日.
2. W. Hsu, C.-Y. Huang, P.-Y. Tsai, P.-H. Shih, S.-K. Fan, D.-J. Yao, C.-H. Liu, and H.-Y. Huang. Magnetic bead–based digital microfluidic immunoanalysis device and method thereof. US Patent. (Pending)
2. W. Hsu, C.-Y. Huang, P.-Y. Tsai, P.-H. Shih, S.-K. Fan, D.-J. Yao, C.-H. Liu, and H.-Y. Huang. Magnetic bead–based digital microfluidic immunoanalysis device and method thereof. US Patent. (Pending)
Honours
Honours
1. 2016 中國機械工程學會 博士論文獎佳作
1. 2016 中國機械工程學會 博士論文獎佳作
2. 2016 國立交通大學 學生優良研究成果獎勵
2. 2016 國立交通大學 學生優良研究成果獎勵
3. 2015 國立交通大學 學生優良期刊論文獎勵
3. 2015 國立交通大學 學生優良期刊論文獎勵
4. 2015 Optofluidics Best Poster Paper Finalist
4. 2015 Optofluidics Best Poster Paper Finalist
5. 2012 IEEE-NANOMED Best Student Paper Finalist
5. 2012 IEEE-NANOMED Best Student Paper Finalist
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