The Pennsylvania State University The Graduate School Department of Chemistry LANGMUIR-BLODGETT FILMS AS MODELS FOR TOF- SIMS INVESTIGATION OF BIOLOGICAL SYSTEMS A Dissertation in Chemistry by Leiliang Zheng © 2008 Leiliang Zheng Submitted in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy August 2008 The dissertation of Leiliang Zheng was reviewed and approved* by the following: Nicholas Winograd Evan Pugh Professor of Chemistry Dissertation Advisor Chair of Committee Barbara Garrison Shapiro Professor of Chemistry Christine D. Keating Associate Professor of Chemistry Ahmed A. Heikal Associate Professor of Bioengineering Ayusman Sen Professor of Chemistry Head of the Department of Chemistry *Signatures are on file in the Graduate School iii ABSTRACT The work presented in this thesis is aimed at utilizing Langmuir-Blodgett (LB) films as model biological systems to improve fundamental understandings and expand applications of time-of-flight secondary ion mass spectrometry (TOF-SIMS) bioanalysis. Chemical imaging of ternary lipid LB monolayer systems shows that it is possible to study lipid-lipid interactions by TOF-SIMS. The incorporation of membrane proteins into the lipid LB monolayer leads to a more representative model and TOF-SIMS imaging is able to identify the membrane protein in the model membrane. Chemically alternating LB multilayer films are developed as a model for fundamental investigations of molecular depth profiling. Organic-organic interface widths are quantitatively studies as a function of temperature, surface topography, and primary ion energy and incident angle. Three-dimensional imaging of the alternating multilayer films visibly displays the “crater effect” and the effect of data acquisition range on interface width measurement is quantitatively studied. iv TABLE OF CONTENTS LIST OF FIGURES ..................................................................................................... vii LIST OF TABLES ....................................................................................................... xi ACKNOWLEDGEMENTS ......................................................................................... xii Chapter 1 Introduction: Langmuir-Blodgett Films and TOF-SIMS Analysis ............ 1 1.1 Langmuir-Blodgett Films – a Biological Model System ................................ 2 1.1.1 Monolayer at the air-water interface .................................................... 2 1.1.2 Supported LB films .............................................................................. 5 1.2 TOF-SIMS – Moving Towards Bioanalysis ................................................... 8 1.2.1 Chemical imaging ................................................................................. 10 1.2.2 Molecular depth profiling ..................................................................... 11 1.2.3 Instrumentations ................................................................................... 13 1.3 References ....................................................................................................... 13 Chapter 2 Investigating Lipid-lipid Interactions by TOF-SIMS Imaging .................. 16 2.1 Introduction ..................................................................................................... 16 2.2 Experimental Section ...................................................................................... 17 2.2.1 Materials ............................................................................................... 17 2.2.2 Substrate Preparation ............................................................................ 18 2.2.3 LB film preparation .............................................................................. 18 2.2.4 Instrumentation and data analysis ........................................................ 19 2.3 Results and Discussion ................................................................................... 21 2.3.1 Lipid domains observed by SIMS ........................................................ 21 2.3.2 Cholesterol interactions with phospholipids ........................................ 25 2.3.3 Quantification of lipid content .............................................................. 26 2.4 Conclusions ..................................................................................................... 30 2.5 Acknowledgement .......................................................................................... 31 2.6 References ....................................................................................................... 31 Chapter 3 Mass Spectrometric Imaging of Membrane Proteins in a Model Langmuir-Blodgett Membrane System ................................................................ 34 3.1 Introduction ..................................................................................................... 34 3.2 Experimental Section ...................................................................................... 37 3.2.1 Materials ............................................................................................... 37 3.2.2 LB film construction ............................................................................. 37 3.2.3 Sample preparation for matrix effect study .......................................... 39 3.2.4 TOF-SIMS analysis .............................................................................. 40 v 3.2.5 Data analysis ......................................................................................... 40 3.3 Results and Discussion ................................................................................... 41 3.3.1 Reference LB films of lipids ................................................................ 41 3.3.2 Evaluation of possible matrix effect ..................................................... 45 3.3.3 GpA reference LB film ......................................................................... 48 3.3.4 Outer membrane leaflet mimic LB film (DPPC/cholesterol/GpA) ...... 48 3.3.5 Inner membrane leaflet mimic LB film (DPPE/Cholesterol/GpA) ...... 50 3.3.6 Principal component analysis ............................................................... 52 3.4 Conclusions ..................................................................................................... 57 3.5 Acknowledgements ......................................................................................... 59 3.6 References ....................................................................................................... 59 Chapter 4 Chemically Alternating Langmuir-Blodgett Multilayer Films as a Model for Molecular Depth Profiling ................................................................... 62 4.1 Introduction ..................................................................................................... 62 4.2 Experimental Section ...................................................................................... 64 4.2.1 Materials ............................................................................................... 64 4.2.2 Substrate and LB film preparation ....................................................... 65 4.2.3 Instrumentation ..................................................................................... 66 4.2.4 Ellipsometry and AFM measurements ................................................. 67 4.3 Results and Discussions .................................................................................. 67 4.3.1 LB film characterisitics ........................................................................ 67 4.3.2 Characterization of LB films by SIMS ................................................. 71 4.3.3 Calculation of depth resolution ............................................................ 75 4.4 Conclustions ................................................................................................... 76 4.5 Acknowledgement .......................................................................................... 77 4.6 References ....................................................................................................... 77 Chapter 5 Molecular Depth Profiling of Multilayer Langmuir-Blodgett Films to Investigate Optimal Depth Resolution .................................................................. 80 5.1 Introduction ..................................................................................................... 80 5.2 Experimental Section ...................................................................................... 83 5.2.1 Materials ............................................................................................... 83 5.2.2 Substrates and film preparation ............................................................ 83 5.2.3 Instrumentation ..................................................................................... 84 5.2.4 Ellipsometry and AFM measurement ................................................... 85 5.3 Results and Discussions .................................................................................. 86 5.3.1 Single component LB films .................................................................. 86 5.3.2 Multilayer structures ............................................................................. 91 5.3.3 Surface Topography ............................................................................. 95 5.3.4 Primary ion energy and incident angle effects ....................................