Optical Measurement of Cell Membrane Tension

Abstract

Using a novel noncontact technique based on optical interferometry, we quantify the nanoscale thermal fluctuations of red blood cells (RBCs) and giant unilamellar vesicles (GUVs). The measurements reveal a nonvanishing tension coefficient for RBCs, which increases as cells transition from a discocytic shape to a spherical shape. The tension coefficient measured for GUVs is, however, a factor of 4-24 smaller. By contrast, the bending moduli for cells and vesicles have similar values. This is consistent with the cytoskeleton confinement model, in which the cytoskeleton inhibits membrane fluctuations [Gov et al., Phys. Rev. Lett. 90, 228101, (2003) The red blood cell (RBC) has a composite membrane consisting of a lipid bilayer coupled to a two-dimensional spectrin network, which grants the cell its characteristic properties of both softness and strong shear elasticity [1,2]. Because mature RBCs lack nuclei, they represent a convenient model for studying cell membranes, which have broad applications in both science and technology [3,4]. The lipid bilayer is 4 -5 nm thick, and exhibits fluidlike behavior, characterized by a finite bending modulus and a vanishing shear modulus, 0. The resistance to shear, crucial for RBC function, is provided by the spectrin network, which has a mesh size of 80 nm.Spontaneous membrane fluctuations, or ''flickering,'' have been modeled theoretically under both static and dynamic conditions [5][6][7][8][9]. These thermally induced membrane motions exhibit 100 nm scale amplitudes at frequencies of tens of Hz. In past studies, measurements of the membrane mean squared displacement versus spatial wave vector, u 2 q, revealed a q ÿ4 dependence predicted by the equipartition theorem, which is indicative of fluidlike behavior [6,10 -13]. These results conflict with the static deformation measurements provided by micropipette aspiration [14,15], high-frequency electric fields [16,17], and, more recently, optical tweezers [18], which indicate an average value for the shear elasticity of the order of 10 ÿ6 J=m 2 . Gov et al. predicted that the cytoskeleton pinning of the membrane has an overall effect of confining the fluctuations and, thus, gives rise to superficial tension much larger than in the case of free bilayers [5]. This confinement model may offer new insight into the cytoskeleton-bilayer interaction that determines the morphology and physiology of the cell [19].RBCs can be assumed optically homogeneous. Therefore, measurement of the cell optical path length via interferometric techniques can provide information about the physical topography of the membrane with subwavelength accuracy and without contact. However, existing optical methods, including phase contrast microscopy (PCM) [6], reflection interference contrast microscopy (RICM) [10], and fluorescence interference contrast (FLIC) [20], are limited in their ability to measure cell membrane displacements. PCM provides phase shifts quantitatively only for samples that are optically much thinner than the wavelength of light, which ...

Measurement of red blood cell mechanics during morphological changes

“…Previous theoretical work has postulated that the presence of the spectrin network leads to additional terms in the deformation energy of the bilayer. Specifically, the ω ∼ q dependence is explained by an effective surface tension generated by the attachment of the membrane to the underlying spectrin network, while the ω ∼ q −1 dependence arises from the uniform confining potential due to this same network (12,17). The validity of these assumptions and the relation between these additional parameters and the microscale structure of the bilayer/spectrin complex remain somewhat elusive.…”

“…RBC thermal fluctuations ("flickering") have been studied for more than a century (7) to better understand the interaction between the lipid bilayer and the cytoskeleton (1, 8, 9). Nevertheless, quantifying these motions is experimentally challenging; reliable spatial and temporal data are desirable (1,8,(10)(11)(12)(13).We use diffraction phase microscopy (DPM) (14, 15) to study the undulatory dynamics of RBC membranes over the commonly occurring DC-EC-SC shape transition. DPM is a highly sensitive imaging technique that provides quantitative, high-stability maps of the optical paths across living cells (Figs.…”

Model for Probing Membrane-Cortex Adhesion by Micropipette Aspiration and Fluctuation Spectroscopy

“…Here, the confinement contribution explicitly arises from the attachment kinetics of the linkers via the adiabatic approximation. This fact allows us to experimentally determine the density of bound linkers, r b,eq , from measurements of the static structure factor of the cell membrane (42). Specifically, the long-wavelength limit q / 0 needs to be measured in fluctuation microscopy experiments in order to determine r b,eq from Eq.…”

Metabolic remodeling of the human red blood cell membrane

“…The RBC membrane is remarkably soft and elastic, and thus exhibits fluctuations with amplitudes of the order of tens of nanometers. The dynamics of the RBC membrane is strongly related to the membrane structure and mechanical properties and has been explored extensively (2)(3)(4)(5)(6). However, experimental results available to date on RBC membrane fluctuations have provided only limited information on select regions of the cell membrane with limited spatial and/or temporal resolution (7)(8)(9).…”