Birefringence of a normal human red blood cell and related optomechanics in an optical trap

Abstract

A normal human red blood cell (RBC) when trapped with a linearly polarized laser, reorients about the electric polarization direction and then remains rotationally bound to this direction. This behavior is expected for a birefringent object. We have measured the birefringence of distortion-free RBCs in an isotonic medium using a polarizing microscope. The birefringence is confined to the cell's dimple region and the slow axis is along a diameter. We report an average retardation of 3.5 ± 1.5 nm for linearly polarized green light (λ = 546 nm). We also estimate a retardation of 1.87 ± 0.09 nm from the optomechanical response of the RBC in an optical trap. We reason that the birefringence is a property of the cell membrane and propose a simple model attributing the origin of birefringence to the phospholipid molecules in the lipid bilayer and the variation to the membrane curvature. We observe that RBCs reconstituted in shape subsequent to crenation show diminished birefringence along with a sluggish optomechanical response in a trap. As the arrangement of phospholipid molecules in the cell membrane is disrupted on crenation, this lends credence to our conjecture on the origin of birefringence. Dependence of the birefringence on membrane contours is further illustrated through studies on chicken RBCs.

Effect of Bovine Serum Albumin on Red Blood Cell Optical Anisotropy Probed Through the Optomechanical Response in an Optical Trap

“…The reorientation dynamics of trapped objects become more interesting when trapped entities possess optical anisotropy along with anisotropy of the shape. In a previous work, we showed that the optomechanical response of a trapped RBC in a linearly polarized OT can be used to obtain estimates of its birefringence and that such estimates of RBC birefringence corroborate with direct measurements carried out using a polarizing microscope. Furthermore, in Ref.…”

“…Furthermore, in Ref. , we also show by means of a geometric model, that the observed variation of birefringence along the RBC diameter may be attributed to the orientation of phospholipid molecules in the lipid bilayer and that any change in the membrane curvature results in an altered value of birefringence. It is thus plausible that pathological conditions and diseases that alter the shape of the RBC will reflect in a change in RBC birefringence.…”

“…This enhancement in birefringence may be thought of as due to a slight flattening of the central crater or the dimple region of the RBC which according to the model proposed by us in Ref. brings more phospolipid molecules into parallel alignment with each other. Thus, it may be seen that treatment with BSA induces changes in cell shape, leading to changes in the orientation of phospholipid molecules.…”

“…We show that changes in optical anisotropy of RBCs brought about by a treatment with BSA can be estimated by studying their optomechanical response in an OT. Since our earlier studies with normal RBCs as well as those RBCs that are subjected to hyperosmotic shock have proved that the observed birefringence is strongly linked to the ordering of the phospholipids in the RBC membrane, enhancement of birefringence with BSA treatment leads us to believe that the lipid bilayer in the RBC membrane must be largely undisrupted by BSA. This work can be taken forward to study the variation of birefringence with BSA concentration and the theoretical model proposed in Ref.…”

“…After trapping an RBC, we rotate the plane of polarization of the trapping laser beam by introducing a half‐wave plate in the optical path. The trapped RBC now rotates about the laser propagation direction till it aligns its slow axis which we identified to be along a diameter to the electric field direction. We show this process in Figure .…”