Chapter 13
Biodegradable, Targeted Polymeric Nanoparticle Drug Delivery Formulation for Cancer Therapy

13.1 Introduction 13.2 Materials 13.2.1 Polymer Synthesis of PLA-PEG and PLGA-PEG 13.2.1.1 Materials for Conjugation via Carbodiimide Chemistry 13.2.1.2 Materials for Conjugation via Ring Opening Polymerization 13.2.2 Nanoparticle Formation 13.2.2.1 Materials 13.2.2.2 Facilities/Equipment 13.2.3 Ligand Conjugation 13.2.3.1 Materials 13.2.4 Quantification of Drug Encapsulation 13.2.4.1 Materials 13.2.4.2 Facilities/Equipment 13.2.5 Release Experiments 13.2.5.1 Materials 13.2.5.2 Facilities/Equipment 13.2.6 Post-Formulation Treatment 13.2.6.1 Materials 13.2.7 Cell Binding and Uptake Experiments 13.2.7.1 Materials 13.2.7.2 Facilities/Equipment 13.2.8 Cytotoxicity Experiments 13.2.8.1 Materials 13.2.8.2 Facilities/Equipment 13.3 Methods 13.3.1 Polymer Synthesis of PLA-PEG and PLGA-PEG 13.3.1.1 Protocol for Conjugation via Carbodiimide Chemistry (if X = -CH3, -OH, or –MAL) 13.3.1.2 Protocol for Conjugation via Carbodiimide Chemistry (if X = -COOH) 13.3.1.3 Protocol for Conjugation via Ring Opening Polymerization 13.3.2 Nanoparticle Formation 13.3.2.1 Protocol for Nanoprecipitation Method 13.3.2.2 Protocol for Single Emulsion Method 13.3.2.3 Protocol for Double Emulsion Method 13.3.3 Conjugation of Targeting Ligand 13.3.3.1 Protocol for Ligand Conjugation via Carbodiimide Chemistry 13.3.3.2 Protocol for Ligand Conjugation via Maleimide-Thiol Chemistry 13.3.4 Quantification of Drug Encapsulation 13.3.4.1 Protocol for Quantification of Drug Encapsulation 13.3.5 Drug Release Studies 13.3.5.1 Protocol for Release Experiment with Low-Solubility Drug 13.3.5.2 Protocol for Release Experiment with High-Solubility Drug 13.3.6 Post-formulation Treatment 13.3.6.1 Method for Particle Storage with Sucrose Lyoprotection 13.3.7 In vitro Experiments: Cell Binding and Uptake Studies 13.3.7.1 Protocol for Fluorescence Microscopy Imaging 13.3.7.2 Protocol for Quantification of Internalization by Flow Cytometry 13.3.8 In vitro Experiments: Cytotoxicity Studies 13.3.8.1 Protocol for Cytotoxicity Study 13.4 Data Acquisition, Results and Interpretation 13.4.1 Polymer Characterization 13.4.2 Nanoparticle Characterization 13.4.3 In vitro Experiments 13.5 Discussion and Commentary 13.5.1 Particle Size 13.5.2 Particle Shape 13.5.3 Surface Chemistry 13.5.4 Drug Loading 13.5.5 Drug Release 13.5.6 Active Targeting and Ligand Conjugation 13.6 Troubleshooting Tips 13.7 Application Notes 13.8 Summary Points 13.9 Acknowledgement 13.10 References

Chapter 14
Porous Silicon Particles for Multi-Stage Delivery

14.1		Introduction
14.2		Fabrication PSPs
14.2.1		Materials
14.2.2		Methods
14.2.2.1		Thin Film Deposition
14.2.2.2		Photolitography
14.2.2.3		Dry Etch
14.2.2.4		Anodic Etch
14.2.2.5		Release of pSi elements to obtain PSPs of desired shape/size/pores
14.2.2.6		Scanning Electron Microscopy (SEM) characterization
14.2.2.7		Nitrogen absorption/desorption characterization
14.2.3		Characterization
14.3		Oxidation and Surface modification with APTES of PSPs
14.3.1		Reagents
14.3.2		Methods
14.3.2.1		Wet Oxidation of PSPs
14.3.2.2		Surface modification of PSPs with APTES
14.4		Fluorescent dye conjugation of PSPs
14.4.1		Reagents
14.4.2		Methodology
14.5		Zeta Potential measurement
14.5.1		Equipment
14.5.2		Reagents
14.5.3		Methodology
14.5.4		Results
14.6		Count and Size Analysis of PSPs
14.6.1		Materials
14.6.1.1		Reagents
14.6.1.2		Facilities/Equipment
14.6.2		Methods
14.6.3		Data Acquisition, Anticipated Results, and Interpretation
14.7		Using Inductively Coupled Plasma – Atomic Emission Spectroscopy (ICP-AES) to determine the amount of degraded silicon in solution
14.7.1		Materials
14.7.1.1		Reagents
14.7.1.2		Facilities/Equipment
14.7.2		Methods
14.7.3		Data Acquisition, Anticipated Results, and Interpretation
14.7.3.1		Data Acquisition
14.7.3.2		Analyzing Data
14.8		Flow Cytometry to characterize PSP shape, size and fluorescence intensity
14.8.1		Materials
14.8.1.1		Reagents
14.8.1.2		Facilities/Equipment
14.8.2		Methods
14.8.3		Data Acquisition, Anticipated Results, and Interpretation
14.9		Loading and Release of Second-stage NPs from PSPs
14.9.1		Loading of NP into PSPs
14.9.1.1		Materials
14.9.1.1.1		Reagents
14.9.1.1.2		Facilities/Equipment
14.9.1.2		Methods
14.9.2		Release of NPs from PSPs
14.9.2.1		Materials
14.9.2.1.1		Reagents
14.9.2.1.2		Facilities/Equipment
14.9.2.2		Methods
14.9.3		Data Acquisition, Anticipated Results, and Interpretation
14.10		Discussion and commentary
14.11		Troubleshooting Table
14.12		Acknowledgements
		References

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