| 00:00:00 | Welcome |
| 00:00:11 | Ragy Ragheb |
| 00:00:44 | Characterizing Liposome Formation, Structure, and Stability with Complementary Techniques |
| 00:00:58 | What are liposomes ? |
| 00:01:49 | Liposomes |
| 00:03:03 | Liposome preparation |
| 00:03:37 | Liposome extrusion |
| 00:04:45 | Malvern Panalytical’s tools for this study |
| 00:05:13 | Liposome characterization solutionsoffered by Malvern Panalytical |
| 00:06:33 | Zetasizer: Dynamic and Electrophoretic Light Scattering |
| 00:06:52 | Dynamic Light Scattering and Brownian Motion |
| 00:07:15 | Dynamic Light Scattering (DLS) |
| 00:07:44 | Intensity Fluctuations, Correlation, and Size Distributions |
| 00:08:21 | Zeta Potential |
| 00:09:08 | Electrophoretic Light Scattering |
| 00:09:46 | Laser Doppler Electrophoresis |
| 00:10:07 | NanoSight – Nanoparticle Tracking Analysis |
| 00:10:18 | Seeing is Believing! |
| 00:10:18 | Patented Optical Arrangement |
| 00:11:14 | Concentration: Quantify particles in known volume |
| 00:11:14 | Parameters measured by NTA |
| 00:12:09 | Zetasizer and NanoSight results |
| 00:12:20 | DOPC, 2mg/mL, checking size by extrusion pass # with various pore-sizes |
| 00:12:56 | Effect of varying lipid concentration on extrusion sizing by pass # through 100nm pore size |
| 00:13:18 | Effect of Freeze / Thaw cycles (5x) on particle size and concentration as extruded through 100 nm pores |
| 00:13:43 | Effect of Freeze / Thaw cycles (5x) on particle size and concentration as extruded through 100 nm pores |
| 00:14:24 | Effect of both Freeze / Thaw cycles (5x) and Step-Down extrusion on particle size and concentration (11x passes/pore size) |
| 00:15:34 | Effect of both Freeze / Thaw cycles (5x) and Step-Down extrusion on particle size and concentration (11x passes/pore size) |
| 00:16:06 | Effect of % Cholesterol on size, concentration, and zeta potential (1mM NaCl + 0.1mM MOPS) with diffusion barrier method |
| 00:17:40 | Effect of % Cholesterol on size, concentration, and zeta potential (1mM NaCl + 0.1mM MOPS) with diffusion barrier method |
| 00:18:04 | 2 mg/mL DOPC with varying mol% Rh-DPPE Fluorescence-mode concentration efficiency of Rhodamine labeled liposomes as a function of increasing dye loadings. |
| 00:19:32 | SAXS / WAXS on liposomes |
| 00:20:52 | What is SAXS ? |
| 00:22:04 | Schematics of a SAXS experiment |
| 00:22:09 | Schematics of a SAXS / WAXS experiment |
| 00:23:24 | EmpyreanVersatile platform for X-ray scattering & diffraction experiments |
| 00:24:13 | Experimental setup for SAXS / WAXS |
| 00:25:14 | SAXS and WAXS measurements |
| 00:25:41 | Temperature-controlled capillary holder |
| 00:25:46 | Investigated samples |
| 00:27:20 | Untitled |
| 00:27:28 | DPPC @ 20°CBefore extrusion - multilamellar vesicles |
| 00:27:33 | DPPC at different temperaturesBefore extrusion |
| 00:28:31 | DPPC at differenttemperatures Before extrusion |
| 00:30:58 | DPPC at different temperaturesBefore extrusion |
| 00:32:41 | DPPC vs. DOPC @ T = 20°CBefore extrusion |
| 00:33:41 | Untitled |
| 00:33:56 | DPPC @ T = 20°Cbefore and after extrusion |
| 00:34:50 | DPPC @ T = 20°Cafter extrusion |
| 00:35:42 | Electron density profile across a lipid bilayer |
| 00:37:12 | Model calculationUni-lamellar vesicle |
| 00:38:42 | Model calculationVariation of lamellarity N |
| 00:38:47 | DPPC @ T = 20°Cafter extrusion |
| 00:39:29 | Summary and conclusions |
| 00:41:31 | Thank you for attention |
Liposomes have been used in drug discovery and drug delivery for some time, and the biophysical characterization of these systems and their payloads is critical to understanding and optimizing their fabrication and function. This study looks at optimal conditions for extruding liposomes as well as their stability under different conditions.
Our aim is to further educate the public about the intricacies of liposome formation and characterization as measured by Nanoparticle Tracking Analysis (NTA) from the NanoSight product range, Dynamic and Electrophoretic Light Scattering (DLS/ELS) from the Zetasizer product range, and Small-angle and Wide-angle X-ray scattering (SAXS/WAXS) from the X-ray analytical product range within Malvern Panalytical.
A broad range of characterization information and combination of both NanoSight and Zetasizer systems helped further optimize fabrication and understand the function of liposomes as well as labeling efficiency of fluorescent lipids. NTA through NanoSight provided number-based high resolution sizing, accurate distribution profiles, concentration (particles/mL), and fluorescence measurements. DLS provided excellent reproducibility, mean size and PDI measurements over a broad range and non-invasive trend analysis. ELS provided zeta potential as a functionality and stability metric of particles. SAXS allowed to investigate the bilayer stacking in MLVs and from WAXS data the order of the alkyl chains (gel phase vs. liquid phase) could be studied as a function of sample temperature.
Speakers
Ragy Ragheb is a Technical Specialist at Malvern Panalytical specializing in the NanoSight and Zetasizer product ranges. Ragy received his Ph.D. in Macromolecular Science and Engineering from Virginia Polytechnic Institute and State University and completed his post-doctoral work in Biomedical Engineering at Yale University. His research focused on the fabrication and characterization of several synthetic and metal-based nanoparticle systems for theranostic and drug delivery applications.
Joerg Bolze studied Chemistry at the University of Karlsruhe in Germany. In 1997 he received a doctorate for his thesis about the structural characterization of colloidal systems with small-angle X-ray scattering. He continued his research as a postdoctor at synchrotron radiation facilities in Japan and Korea. In 2006 Joerg joined PANalytical as an application specialist for small-angle X-ray scattering. In 2012 he was appointed as Product Marketing manager Nanomaterials. Over the years he contributed significantly to the development of Malvern PANalytical's small- and wide angle X-ray scattering solutions.
More information
Who should attend?
Anyone interested in characterizing both biological and synthetic nanoparticles.
What will you learn?
The benefits and complementary nature of both NanoSight (Nanoparticle Tracking Analysis) and Zetasizer (Dynamic and Electrophoretic Light Scattering) in characterizing liposomes.