PATTERNED NANOARRAY SERS SUBSTRATES FOR PATHOGEN DETECTION A Thesis Presented to The Academic Faculty by Nicole Ella Marotta In Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the School of Chemistry and Biochemistry Georgia Institute of Technology August 2010 COPYRIGHT 2010 BY NICOLE ELLA MAROTTA PATTERNED NANOARRAY SERS SUBSTRATES FOR PATHOGEN DETECTION Approved by: Dr. Lawrence Bottomley, Advisor Dr. Mostafa El-Sayed School of Chemistry and Biochemistry School of Chemistry and Biochemistry Georgia Institute of Technology Georgia Institute of Technology Dr. L. Andrew Lyon Dr. Julia Kubanek School of Chemistry and Biochemistry School of Biology Georgia Institute of Technology Georgia Institute of Technology Dr. Loren Williams School of Chemistry and Biochemistry Georgia Institute of Technology Date Approved: August 12, 2010 To Mom and Dad. I love you. ACKNOWLEDGMENTS I want to first and foremost thank my parents who have always encouraged me to do my best. Whether it was playing a sport, performing in a musical or doing well in school you were always there to cheer me on. You sat on the sidelines when I was playing field hockey, sat in the front row when I was singing, and traveled down the east coast for my defense. I am so happy to know you will always be there, even when I am in a different state. Without your constant love and support, I would not be where I am today, and I honestly cannot thank you enough. I want to thank my advisor, Prof. Larry Bottomley, or Dr. B as I call him, for helping me become a better scientist. I remember when I was struggling with the this project and debating whether or not to complete the PhD. program. I had a meeting with you and you told me to trust you, it was going to work out. I did and I could not be happier. Thank you for believing that I could do it and helping me see that. I would also like to thank the rest of my thesis committee, Professors Andrew Lyon, Julia Kubanek, Nick Hud, and Mostafa El-Sayed for their words of wisdom and encouragement over the past few years. In addition to my family, advisor, and committee, I want to thank the friends I have made here in Atlanta and at Tech. I love all of you but a few deserve acknowledgment. Laura, thanks for being such an awesome friend. You are always there to talk if I need to, or just sit by the pool and hang out. Susan, thanks for being a great carpool partner and an even better date on Thursday nights. Meg, you lived with me for three of the most stressful years of my life and have become one of my closest friends. Thanks for helping me realize I didn't have to look up what a plasmon was again before my OP. I would also like to thank all of the individuals that have helped me with my research. Thank you to current group members Megan Damm, Kelsey Beavers and David Futur and to past group members Jabulani Barber and Kane Barker for their thought provoking discussions and suggestions. I would like to thank past undergraduate researchers Katherine Siemens, Morgan Nunn, and Kit Ledford for helping me acquire and analyze data, Prof. Mohan Srinivasarao for allowing extended use of his Raman spectrometer and Dr. Matija Crne and Minsang Park for their help in keeping the spectrometer in working order. I also want to thank Charlie Suh, Mikkel Thomas, Janet Cobb-Sulivan and Gary Spinner of the Georgia Tech MiRC Cleanroom for all of their suggestions and guidance concerning clearnroom tools and processes. I want to acknowledge Profs. R. A. Dluhy, R. A. Tripp, and Y. Zhao and Drs. Jeremy Driskell and Vivien Chu from University of Georgia for their fruitful discussions regarding the fabrication and characterization of substrates. Lastly, I graciously acknowledge Georgia Tech/UGA Biomedical Research program, Georgia Research Alliance VentureLab, Argent Diagnostics and SensorTech Inc. for providing the necessary funding. v TABLE OF CONTENTS Page ACKNOWLEDGMENTS iv LIST OF TABLES xiv LIST OF FIGURES xv LIST OF SYMBOLS AND ABBREVIATIONS xxv SUMMARY xxxii CHAPTER 1 Introduction to Surface Enhanced Raman Scattering 1 1.1 History of Raman 1 1.1.1. Raman Scattering 1 1.1.2. Polarizability 5 1.2. Instrumentation 5 1.2.1. Laser Source 5 1.2.2. Sample Cells 6 1.2.3. Wavelength Selector 6 1.2.4. Transducer and Readout System 8 1.3. Early Raman Studies 8 1.4. Discovery of Surface Enhanced Raman Scattering 10 1.5. Surface Enhancement Mechanisms 14 1.5.1. Electromagnetic Enhancement 14 1.5.2. Chemical Enhancement 17 1.5.3. Distance Dependence 19 1.6. SERS Substrates 21 vi 1.6.1. Nanoparticles 22 1.6.1.1. Nanoparticle Aggregation 22 1.6.2. Nanoshells 23 1.6.3. Film Over Nanosphere 23 1.6.4. Nanorods 24 1.7. Applications of SERS 26 1.7.1. Single Molecule Surface Enhanced Raman Scattering 26 1.7.2. Biomolecular Sensing 27 1.7.2.1. Pathogen Detection 27 1.7.2.1.1. Viruses 27 1.7.2.1.2. Bacteria Cells 28 1.7.2.1.3. Nucleotides 28 2 Introduction to Glancing Angle Deposition 32 2.1. Physical Vapor Deposition 32 2.1.1. Sputterer Deposition 34 2.1.2. Thermal Evaporation 36 2.1.2.1. Resistive Evaporation 36 2.1.2.2. Electron Beam Evaporation 38 2.1.3. Plume Characteristics 40 2.2. Glancing Angle Deposition (GLAD) 43 2.2.1. Columnar Structure Formation 45 2.2.1.1. Nucleation 45 2.2.1.2. Ballistic Shadowing 47 2.2.1.3. Surface Diffusion and Temperature 48 2.2.2. Column Development 50 vii 2.2.2.1. Structure Control - Seeding 51 2.2.2.2. Structure Control - Angles and Rotation 53 2.3. Applications of Structures Fabricated Using GLAD 58 2.3.1. Optical Sensors 59 2.3.2. Increased Surface Area 60 2.3.2.1. Electrode Systems 60 2.3.2.2. Sensor Systems 60 2.3.2.3. Chemical Processes 61 2.3.3. Multifunctional Patchy Particles 61 2.3.4. Silver Nanorod Arrays 62 3 Patterned Silver Nanorod Array Substrates for Surface Enhanced Raman Spectroscopy 65 3.1. Introduction 65 3.2. Methods and Materials 66 3.2.1. Substrate Fabrication 66 3.2.2. Substrate Patterning 67 3.2.3. Substrate Characterization 68 3.3. Results and Discussion 69 3.4. Conclusion 86 4 Thermal Stability of Silver Nanorod Arrays 90 4.1. Introduction 90 4.2. Methods and Materials 91 4.2.1. Substrate Fabrication 91 4.2.1.1. Method One 91 4.2.1.2. Method Two 91 4.2.2. Substrate Characterization 91 viii 4.3. Results 92 4.4. Discussion 99 5 Polystyrene Beads as Probes of the SERS Response Characteristics of Silver Nanorod Arrays 102 5.1. Introduction 102 5.2. Methods and Materials 103 5.2.1. SERS Substrates 103 5.2.2. Analytes 103 5.2.3. Substrate Characterization 103 5.3. Results and Discussion 104 6 Surface Enhanced Raman Scattering of Bacterial Growth Culture Media 127 6.1. Introduction 127 6.2. Methods and Materials 128 6.2.1. Substrate Fabrication 128 6.2.2. Cell Culture Growth Media 128 6.2.3. Bacteria and Cell Culture Preparation 128 6.2.4. Substrate Characterization 129 6.3. Results 129 6.4. Discussion 140 7 Limitations of a SERS-Based Assay of DNA Hybridization 151 7.1. Introduction 151 7.2. Materials 153 7.3. Methods 153 7.3.1. SERS Substrates 153 7.3.2. SERS Characterization 154 7.3.3. Scanning Electron Microscopy (SEM) Characterization 154 ix 7.3.4. Transmission Electron Microscopy (TEM) Characterization 156 7.3.5. Dynamic Light Scattering (DLS) Measurements 156 7.3.6. DNA Melting Curves 156 7.3.7. AFM Imaging 157 7.4. Results and Discussion 158 7.5. Conclusion 215 8 Conclusions and Future Work 218 APPENDIX A: Synthesis of Unique DNA Structures for AFM Pulling Studies 222 A.1. Introduction 222 A.2. DNA Loop Structure 228 A.2.1. Materials 228 A.2.2. Methods 228 A.2.2.1. Digestion of Plasmids 228 A.2.2.2. Ligation of Plasmid Fragments and Insert 229 A.2.2.3. DNA Gel Electrophoresis 231 A.2.2.4. Preparation of AFM Cantilevers 231 A.2.3. Results 233 A.2.3.1. Plasmid pBR322 Digestion 233 A.2.3.2. Plasmid pBR322 Ligation 233 A.2.3.3. Plasmid pUC19 Digestion 234 A.2.3.4. Plasmid pUC19 Ligation 234 A.2.3.5. Plasmid pTWIN1 Digestion 241 A.2.3.6. Plasmid pTWIN1 Ligation 241 A.2.4. Work Left to Be Done 242 A.3. DNA Block Copolymer Structure 246 x A.3.1.Materials 246 A.3.2. Methods 246 A.3.2.1. Synthesis Reaction 246 A.3.2.2. HPLC Separation 247 A.3.2.3. AFM Studies 247 A.3.3. Results 247 A.3.4. Work Left to Be Done 249 APPENDIX B: Fabrication of a micro-DSC Heater Device 254 B.1. Introduction 254 B.1.1. Thermal Modeling 254 B.2.2. Process and Mask Design 255 B.2. Prototype #1 Initial Process Flow 256 B.2.1. Deposition of Silicon Nitride Mesas for the micro-Heater 256 B.2.2. Creation of Wiring for the micro-Heater 258 B.2.3. Deposition of the Nickel Serpentine for the micro-Heater 258 B.2.4. Deposition of Silicon Nitride Barrier Layer 261 B.2.5.