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Label-Free Biochemical Recognition Using MEMS Resonators for Microarray Technology

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Label-Free Biochemical Recognition Using MEMS Resonators for Microarray Technology University of Denver Digital Commons @ DU Electronic Theses and Dissertations Graduate Studies 1-1-2011 Label-Free Biochemical Recognition Using MEMS Resonators for Microarray Technology Babak Tousifar University of Denver Follow this and additional works at: https://digitalcommons.du.edu/etd Part of the Biomechanical Engineering Commons Recommended Citation Tousifar, Babak, "Label-Free Biochemical Recognition Using MEMS Resonators for Microarray Technology" (2011). Electronic Theses and Dissertations. 655. https://digitalcommons.du.edu/etd/655 This Thesis is brought to you for free and open access by the Graduate Studies at Digital Commons @ DU. It has been accepted for inclusion in Electronic Theses and Dissertations by an authorized administrator of Digital Commons @ DU. For more information, please contact [email protected],[email protected]. LABEL-FREE BIOCHEMICAL RECOGNITION USING MEMS RESONATORS FOR MICROARRAY TECHNOLOGY __________ A Thesis Presented to the Faculty of Engineering and Computer Science University of Denver __________ In Partial Fulfillment of the Requirements for the Degree Master of Science __________ by Babak Tousifar November 2011 Advisor: Dr. Siavash Pourkamali ©Copyright by Babak Tousifar 2011 All Rights Reserved Author: Babak Tousifar Title: LABEL-FREE BIOCHEMICAL RECOGNITION USING MEMS RESONATORS FOR MICROARRAY TECHNOLOGY Advisor: Dr. Siavash Pourkamali Degree Date: November 2011 Abstract Highly sensitive biosensors capable of detecting and characterizing smallest quantities of cellular and molecular targets are needed in pharmaceutical and medical diagnostics industries. In this work, the importance of biological target recognition specifically proteins through microarray technologies has been discussed and the most successful tools and techniques have been studied. Moreover, a thermally actuated Micro Electro-Mechanical Systems (MEMS) resonator has been demonstrated and fabricated in this work as an accurate, reliable and low cost biotechnology tool. For the proof of concept, amine coupling of octadecylamine chain to the epoxide reactive groups of the functionalized silicon dioxide surfaces has been shown through resonator frequency monitoring. The frequency deviation of the sensors implied a meaningful surface coverage after target detection. Furthermore, X-ray Photoelectron Spectroscopy (XPS) analysis of the devices at different stages of the surface modification supported that the frequency deviations are due to receptor and target attachments. In addition, two surface passivation techniques against non-specific adsorption of proteins have been investigated on silicon surfaces and fluorescent microscopy was performed for quality control and validation of experiments. ii Acknowledgements Words fall short to convey my gratitude to those who helped and supported me during my Master studies. I would like to take this opportunity to express my individualized thanks to all of those, even though the list is long and the space allocation for this purpose is short. My special thanks and heartfelt gratitude go to my dear academic advisor Dr. Siavash Pourkamali and research co-adviser Dr. Byron W. Purse for their invaluable advice, patience, guidance, and consistent support throughout my studies. The accomplishment of this work would have been impossible without their help and support. I would like to extend my sincere appreciation to my dear professor Dr. Corinne Lengsfeld for her kind supports. I am also grateful of Professor Daniel L. Armentrout for his thoughtful advice in a section of this thesis. I would like to thank the Department of Mechanical and Material Engineering of University of Denver for its financial support and providing a friendly and progressive work atmosphere. I also appreciate the help and cooperation of Department of Chemistry at Colorado State University for providing the XPS facility. Furthermore, I would like to thank my colleagues and friends and at University of Denver Specifically Dr. Mirek Kvanisca and Jennifer Chapin for their valuable dedications. Finally, the deepest gratitude goes to my ever companion Saba Bakhshi, my dear family and friends for their eternal love and support. iii Table of Contents List of Figures .................................................................................................................... vi List of Tables ...................................................................................................................... x Chapter One: Introduction .................................................................................................. 1 1.1 Proteomics and Microarrays ............................................................................. 1 1.2 Gene Expression Review .................................................................................. 2 1.2.1 Transcription ...................................................................................... 3 1.2.2 Translation ......................................................................................... 3 1.3 Protein Microarray Applications ....................................................................... 7 1.3.1 Protein Therapeutics .......................................................................... 7 1.3.2 Medical Diagnostics........................................................................... 8 1.3.3 National Safety ................................................................................... 9 1.4 Microarray Sensing Mechanisms ...................................................................... 9 1.4.1 Sensor Interface and Substrates ....................................................... 10 1.4.2 Molecular Recognition Elements ..................................................... 11 1.4.3 Transduction Mechanisms ............................................................... 16 1.5 Microarray Challenges and its Prospective ..................................................... 24 Chapter Two: MEMS Biosensing Method and Mechanism ............................................. 27 2.1 Thermal-Piezoresistive Resonators ................................................................. 27 2.1.1 Thermal Actuation ........................................................................... 27 2.1.2 Device Concept and Operation Principles ....................................... 28 2.1.3 Fabrication Process .......................................................................... 30 2.1.5 Detection Sensitivity Limit .............................................................. 33 4.4 Resonator Static Temperature and its Effect on Biomolecules ...................... 35 2.1.7 Biochemical Sensing Mechanism .................................................... 37 Chapter Three: Biochemical Sensing Results and Discussion ......................................... 39 3.1 Octadecylamine Sensing ................................................................................. 39 3.1.1 Surface Functionalization ................................................................ 39 3.1.2 Octadecylamine Couplings .............................................................. 41 3.1.3 Frequency Measurements ................................................................ 41 3.1.4 Surface Coverage Analysis .............................................................. 44 3.1.5 XPS Surface Analysis ...................................................................... 45 3.2 Biotin-Avidin Conjugation Sensing ................................................................ 49 3.2.1 Surface Biotinylation ....................................................................... 50 iv 3.2.2 Avidin Conjugation .......................................................................... 53 3.3 Surface Passivation Methods and Characterization ........................................ 57 3.3.1 Polymer Chemistry .......................................................................... 57 3.3.2 Surface Chemistry ............................................................................ 60 Chapter Four: Conclusion, Discussion and Future Work ................................................. 64 4.1 Real Time Biochemical Sensing ..................................................................... 64 4.2 Array Implementation ..................................................................................... 66 4.3 Encapsulation in Micro-Fluidic Channels ...................................................... 67 References ......................................................................................................................... 69 v List of Figures Figure 1.1 (a) Gene locations in Eukaryotic Cells in the form of DNA double helix strand, (b) DNA double strand macromolecule structure and its biochemical components. Figure 1.2 Transcription process in the cell nucleolus starts from attachment of an RNA Polymeras to the specific genes and encode a single strand molecule (RNA) from the template strands. Figure 1.3 Three-nucleotide based Codons for amino acids. UAA, UAG and UGA are stop codons which terminate the translation process. Figure 1.4 Gene expression process in eukaryotic cells and detailed description of its steps from DNA transcription to Protein (polypeptide) translation. Figure 1.5 Biosensor schematic and its components including recognition element, interface and transducer. Figure 1.6 Microarray substrate with the grown interfaces to immobilize the molecule of interest i.e. protein. Figure 1.7 (a) Double Strand DNA (dsDNA) denatures
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