Physics Chat
A Label Free Method to Measure Lipid Membrane Dynamics of Giant Unilamellar Vesicles
Speaker: Freya Turley (Prof Wolfgang Langbein's group)
Date: Thursday 23 March 2023
Time: 14:30
Venue: Queen's buildings North N3.23 with zoom live streaming
The interaction between proteins and lipid membranes is a fundamental process underpinning key functions in cell biology. Despite the importance of such system, many questions are still unanswered concerning the mechanism of protein organisation and function within the cell membrane, and how this is modulated by the lipid environment, due to the lack of suitable measurement techniques. We are developing a novel optical microscopy technique called Interferometric Gated Off-axis reflectometry (iGOR) which promises to be a step-change, enabling us to monitor the motion of single membrane proteins with high sensitivity (~10kDa estimated smallest size limit) rapidly (~1ms) in 3D label-free with ~10nm localisation precision, simultaneously with local membrane properties at the protein site, such as membrane curvature and thickness changes. As lipid membrane model systems, we are using giant unilamellar vesicles (GUVs) of about 30-50um diameter. This is a versatile system where we can control the lipid composition and the osmolarity of the internal and external solutions. We have optimised GUV stability using sucrose concentration differences, characterised via a quantitative differential interference contrast microscopy (qDIC) method developed in house. We found that using a concentration difference between the internal and external sucrose solution >0.2mM establishes an internal pressure too great for a unilamellar vesicle to withstand. We developed a numerical simulation model fitting the experimental qDIC data to accurately determine the lamellarity. An overview of these qDIC results and our progress on characterising lipid membrane dynamics and the insertion of pore forming proteins using iGOR will be presented.