Refined contour analysis of giant unilamellar vesicles

Abstract

The fluctuation spectrum of giant unilamellar vesicles is measured using a high-resolution contour detection technique. An analysis at higher q vectors than previously achievable is now possible due to technical improvements of the experimental setup and of the detection algorithm. The global fluctuation spectrum is directly fitted to deduce the membrane tension and the bending modulus of lipid membranes. Moreover, we show that the planar analysis of fluctuations is valid for spherical objects, even at low wave vectors. Corrections due to the integration time of the video camera and to the section of a 3D object by the observation plane are introduced. A precise calculation of the error bars has been done in order to provide reliable error estimate. Eventually, using this technique, we have measured bending moduli for EPC, SOPC and SOPC: CHOL membranes confirming previously published values. An interesting application of this technique can be the measurement of the fluctuation spectra for non-equilibrium membranes, such as "active membranes".

Direct Cytoskeleton Forces Cause Membrane Softening in Red Blood Cells

“…In real space, the RBC-equatorial fluctuations, namely h(x,t) ¼ R(x,t) À R 0 , are measured as local deviations of the equatorial radius, R(x,t), from an average value, R 0 , calculated with respect to the center of mass of the discoidal profile (29,30). Fig.…”

“…However, in the flickering experiment, we can only obtain information from the focal plane, which is adjusted at the equatorial site where the sharpest image can be focused. Several authors have clarified how the MS analysis can be adapted to the case where the equatorial contour of an axisymmetric membrane is tracked by time-resolved microscopy (29,30). In this case, the equatorial undulations correspond to the discrete sequence of circular eigenmodes with the wavevector determined by the azimuthal number, q ¼ m/R (m ¼ 2,3,4.N and R 0 is the spherical radius).…”

“…Experimental approaches to the flickering phenomenon suffer from restricted time resolution and the limited spatial discrimination of membrane motions in the submicron scale. The use of ultrafast optical microscopy (28) renders the classical flickering method (29,30) capable of tracking cell contour fluctuations at very high sampling rates with subpixel resolution, thus allowing noninvasive detection of signal correlations over broad timescales. Normal RBCs have a characteristic biconcave or discocyte shape, a symmetry property that facilitates the detection, in Fourier space, of the active contributions to the normal modes of membrane motion at the equatorial rim.…”

Red blood cell thermal fluctuations: comparison between experiment and molecular dynamics simulations

“…20 A series of contours (between 1500 and 2500, each consisting of more than 600 points) is then analysed to quantify the membrane fluctuations. Following a standard method 24,25 , each contour is represented by a Fourier series, Hydrogen peroxide is known to induce the formation of haemoglobin-spectrin complexes 27,28 resulting, amongst other 40 changes, in decreased cell deformability and therefore we used it here as a convenient agent to stiffen the membrane.…”

“…Normal physiological function of the red blood cell (RBC) depends, amongst other factors, on finely tuned elastic properties of its plasma membrane, allowing the cell to maintain its overall shape during blood flow, yet to be flexible 25 enough to pass through fine vessels of comparable or smaller size. Numerous experimental and theoretical studies have contributed to our understanding of the molecular and supramolecular origins of RBC elasticity.…”