Methods in Bioengineering: Nanoscale Bioengineering and Nanomedicine -- Contents -- Preface -- Chapter 1 Preparation and Characterization of Carbon Nanotube-Protein Conjugates -- 1.1 Introduction -- 1.2 Materials -- 1.3 Methods -- 1.3.1 Physical Adsorption of Proteins on Carbon Nanotubes -- 1.3.2 Protein Assisted Solubilization of Carbon Nanotubes -- 1.3.3 Covalent Attachment of Proteins onto Carbon Nanotubes -- 1.4 Data Acquisition, Anticipated Results, and Interpretation of Data -- 1.4.1 Characterization of Proteins Physically Adsorbed onto Carbon Nanotubes -- 1.4.2 Characterization of Protein-Solubilized Carbon Nanotubes -- 1.4.3 Characterization of Covalently Attached Carbon Nanotube-Protein Conjugates -- 1.5 Discussion and Commentary -- 1.6 Applications Notes -- 1.7 Summary Points -- Acknowledgments -- References -- Chapter 2 Peptide-Nanoparticle Assemblies -- 2.1 Introduction -- 2.2 Materials -- 2.3 Methods -- 2.3.1 Coil-Coil Peptide Mediated NP Assembly -- 2.3.2 Synthesis of Hybrid Structures Using Multifunctional Peptides -- 2.4 Assembly Mediated by Metal Ion-Peptide Recognition -- 2.5 Peptides as Antibody Epitopes for Nanoparticle Assembly -- 2.6 DATA Acquisition, Anticipated Results, and Interpretation -- 2.7 Discussion and Commentary -- 2.8 Application Notes -- 2.9 Summary Points -- Acknowledgments -- References -- Chapter 3 Nanoparticle-Enzyme Hybrids as Bioactive Materials -- 3.1 Introduction -- 3.2 Materials -- 3.3 Methods -- 3.3.1 Enzyme-Attached Polystyrene Nanoparticles -- 3.3.2 Polyacrylamide Hydrogel Nanoparticles for Entrapment of Enzymes -- 3.3.3 Magnetic Nanoparticles with Porous Silica Coating for Enzyme Attachment -- 3.3.4 Enzyme Loading and Activity Assay -- 3.4 Results -- 3.4.1 Polystyrene-Enzyme Hybrid Nanoparticles -- 3.4.2 Polyacrylamide Hydrogel Nanoparticles with Entrapped Enzymes.

3.4.3 Magnetic Nanoparticles for Enzyme Attachment -- 3.5 Discussion and Commentary -- 3.6 Troubleshooting -- 3.7 Application Notes -- 3.8 Summary Points -- Acknowledgments -- References -- Chapter 4 Self-Assembled QD-Protein Bioconjugates and Their Use in Fluorescence Resonance Energy Transfer -- 4.1 Introduction -- 4.2 Materials -- 4.2.1 Reagents -- 4.2.2 Equipment -- 4.3 Methods -- 4.3.1 Quantum Dot Synthesis -- 4.3.2 Surface Ligand Exchange -- 4.3.3 Biomolecule Conjugation -- 4.3.4 Fluorescence Measurements -- 4.4 Data Analysis and Interpretation -- 4.4.1 Calculating Donor-Acceptor Distances -- 4.4.2 Calculating Reaction Rates of Surface-Bound Substrates -- 4.5 Summary Points -- 4.6 Conclusions -- References -- Annotated References -- Chapter 5 Tracking Single Biomolecules in Live Cells Using Quantum Dot Nanoparticles -- 5.1 Introduction -- 5.2 Materials -- 5.2.1 Reagents -- 5.2.2 Imaging Equipment -- 5.3 Methods -- 5.3.1 Forming QD Bioconjugates -- 5.3.2 Treating Cells with QD Bioconjugates -- 5.4 Data Acquisition, Anticipated Results, and Interpretation -- 5.4.1 Imaging QD-Bound Complexes in Cells -- 5.4.2 Analysis of the Real-Time QD Dynamics -- 5.5 Discussion and Commentary -- References -- Chapter 6 Nanoparticles as Biodynamic Substrates for Engineering Cell Fates -- 6.1 Introduction -- 6.2 Experimental Design -- 6.3 Materials -- 6.3.1 Cell Culture, Fixing, Staining, and Analysis Reagents -- 6.3.2 Nanoparticle Fabrication and Functionalization -- 6.3.3 Microscale Plasma Initiated Patterning -- 6.4 Methods -- 6.4.1 Albumin Nanoparticle Fabrication -- 6.4.2 Albumin Nanoparticle Functionalization -- 6.4.3 Albumin Nanoparticle Pattern Creation-Microscale Plasma Initiated Patterning (uPIP) -- 6.4.4 Cell Culture -- 6.4.5 Keratinocyte Morphology and Migration -- 6.4.6 Fibroblast Extracellular Matrix Assembly -- 6.4.7 Cell Attachment Assay.

6.5 Results -- 6.5.1 Enhanced Cell Migration -- 6.5.2 Enhanced Extracellular Matrix Assembly -- 6.6 Discussion of Pitfalls -- 6.6.1 Spatial Guidance of Cell Attachment-Microscale Plasma Initiated Patterning -- 6.6.2 Three-Dimensional Presentation of Albumin Nanoparticles -- 6.7 Summary Points -- Acknowledgments -- References -- Chapter 7 Magnetic Cell Separation to Enrich for Rare Cells -- 7.1 Introduction -- 7.1.1 Principle -- 7.1.2 Examples of Cell Magnetic Separation Applications -- 7.2 Materials and Methods -- 7.2.1 Enrichment Process -- 7.2.2 Red Cell Lysis Step -- 7.2.3 Immunomagnetic Labeling -- 7.2.4 Magnetic Cell Separation Step -- 7.3 Data Acquisition, Results, and Interpretation -- 7.4 Discussion and Commentary -- 7.5 Summary Points to Obtain High-Performance, Magnetic Cell Separations -- Acknowledgments -- References -- Chapter 8 Magnetic Nanoparticles for Drug Delivery -- 8.1 Introduction -- 8.2 Experimental Design -- 8.3 Materials -- 8.3.1 Reagents -- 8.3.2 Facilities and Equipment -- 8.4 Methods -- 8.4.1 Synthesis of Magnetic Nanoparticles -- 8.4.2 Physical Characterization of Magnetic Nanoparticles -- 8.4.3 Conversion of DOX HCl -- 8.4.4 Drug Loading and Release Kinetics -- 8.4.5 Kinetics of DOX Release from Magnetic Nanoparticles -- 8.4.6 Antiproliferative Activity of Doxorubicin Loaded Magnetic Nanoparticles on MCF-7 Cells -- 8.4.7 Antiproliferative Activity of Doxorubicin Loaded MagneticNanoparticles on MCF-7 Cells in the Presence of a Magnetic Field -- 8.5 Data Acquisition, Anticipated Results, and Interpretation -- 8.6 Discussion and Commentary -- 8.7 Application Notes -- 8.8 Summary Points -- Acknowledgments -- References -- Chapter 9 Imaging and Therapy of Atherosclerotic Lesions with Theranostic Nanoparticles -- 9.1 Introduction -- 9.2 Experimental Design -- 9.3 Materials -- 9.3.1 Reagents -- 9.3.2 Facilities/Equipment.

9.3.3 Animal Model -- 9.3.4 Alternate Reagents and Equipment -- 9.4 Methods -- 9.4.1 Synthesis of Theranostic Nanoparticles -- 9.4.2 Intravital Fluorescence Microscopy -- 9.4.3 Light-Based Therapy -- 9.5 Data Acquisition, Anticipated Results, and Interpretation -- 9.5.1 Characterization of Theranostic Nanoparticles -- 9.5.2 Animal Experimentation -- 9.5.3 Intravital Fluorescence Microscopy -- 9.5.4 Statistical Analyses -- 9.5.5 Anticipated Results -- 9.6 Discussion and Commentary -- 9.7 Summary Points -- Acknowledgments -- References -- Chapter 10 Biomedical Applications of Metal Nanoshells -- 10.1 Introduction -- 10.1.1 Biomedical Applications of Metal Nanoshells -- 10.1.2 Nanoshells for Combined Optical Contrast and Therapeutic Application -- 10.2 Experimental Design -- 10.3 Materials -- 10.3.1 Nanoparticle Production -- 10.3.2 Protein Conjugation to Nanoshells Surface -- 10.3.3 Cell Culture -- 10.3.4 In Vitro Assays -- 10.4 Methods -- 10.4.1 Fabrication of Gold/Silica Core Nanoshells -- 10.4.2 Nanoshells for Combined Imaging and Therapy In Vivo -- 10.4.3 Passivation of Nanoshells with PEG -- 10.4.4 Conjugation of Biomolecules to Nanoshells -- 10.4.5 Quantification of Antibodies on Nanoshells -- 10.5 Results -- 10.5.1 Gold/Silica Nanoshells Allow Both Imaging Contrast Increase and Therapeutic Benefit -- 10.5.2 Evaluation of Antibody Concentration per Nanoshell -- 10.6 Discussion of Pitfalls -- 10.7 Statistical Analysis -- Acknowledgments -- References -- Chapter 11 Environmentally Responsive Multifunctional Liposomes -- 11.1 Introduction -- 11.1.1 Cis-Aconityl Linkage -- 11.1.2 Trityl Linkage -- 11.1.3 Acetal Linkage -- 11.1.4 Polyketal Linkage -- 11.1.5 Vinyl Ether Linkage -- 11.1.6 Hydrazone Linkage -- 11.1.7 Poly(Ortho-Esters) -- 11.1.8 Thiopropionates -- 11.2 Materials -- 11.2.1 Chemicals -- 11.2.2 Syntheses.

11.2.3 Preparation of the TATp-Bearing, Rhodamine-Labeled Liposomal Formulations -- 11.2.4 Preparation of the TAtp-Bearing, Rhodamine Labeled, pGFP Complexed Liposomal Formulations -- 11.3 Methods -- 11.3.1 Synthesis of Hydrazone-Based mPEG-HZ-PE Conjugates [63, 66] -- 11.3.2 Synthesis of PE-PEG1000-TATp Conjugate [66] -- 11.3.3 In Vitro pH-Dependant Degradation of PEG-HZ-PE Conjugates -- 11.3.4 Avidin-Biotin Affinity Chromatography -- 11.3.5 In Vitro Cell-Culture Study -- 11.3.6 In Vivo Study -- 11.3.7 In Vivo Transfection with pGFP -- 11.4 Discussion and Commentary -- 11.4.1 Synthesis of Hydrazone-Based mPEG-HZ-PE Conjugates -- 11.4.2 Synthesis of PE-PEG1000-TATp Conjugate -- 11.4.3 In Vitro pH-Dependant Degradation of PEG-HZ-PE Conjugates -- 11.4.4 Avidin-Biotin Affinity Chromatography -- 11.4.5 In Vitro Cell Culture Study -- 11.4.6 In Vivo Study -- 11.4.7 In Vivo pGFP Transfection Experiment -- 11.5 Conclusion -- 11.6 Summary Points -- Acknowledgments -- References -- Chapter 12 Biodegradable, Targeted Polymeric Nanoparticle Drug Delivery Formulation for Cancer Therapy -- 12.1 Introduction -- 12.2 Materials -- 12.2.1 Polymer Synthesis of PLA-PEG and PLGA-PEG -- 12.2.2 Nanoparticle Formation -- 12.2.3 Ligand Conjugation -- 12.2.4 Quantification of Drug Encapsulation -- 12.2.5 Release Experiments -- 12.2.6 Postformulation Treatment -- 12.2.7 Cell Binding and Uptake Experiments -- 12.2.8 Cytotoxicity Experiments -- 12.3 Methods -- 12.3.1 Polymer Synthesis of PLA-PEG and PLGA-PEG -- 12.3.2 Nanoparticle Formation -- 12.3.3 Conjugation of Targeting Ligand -- 12.3.4 Quantification of Drug Encapsulation -- 12.3.5 Drug Release Studies -- 12.3.6 Postformulation Treatment -- 12.3.7 In Vitro Experiments: Cell Binding and Uptake Studies -- 12.3.8 In Vitro Experiments: Cytotoxicity Studies -- 12.4 Data Acquisition, Results, and Interpretation.

12.4.1 Polymer Characterization.