Detachment of agglutinin-bonded red blood cells. III. Mechanical analysis for large contact areas.

An experimental method and analysis are introduced which provide direct quantitation of the strength of adhesive contact for large agglutinin-bonded regions between macroscopically smooth membrane capsules (e.g., red blood cells). The approach yields intrinsic properties for separation of adherent regions independent of mechanical deformation of the membrane capsules during detachment. Conceptually, the micromechanical method involves one rigid test-capsule surface (in the form of a perfect sphere) held fixed by a micropipette and a second deformable capsule maneuvered with another micropipette to force contact with the test capsule. Only the test capsule is bound with agglutinin so that the maximum number of cross-bridges can be formed without steric interference. Following formation of a large adhesion region by mechanical impingement, the deformable capsule is detached from the rigid capsule surface by progressive aspiration into the micropipette. For the particular case modeled here, the deformable capsule is assumed to be a red blood cell which is preswollen by slight osmotic hydration before the test. The caliber of the detachment pipette is chosen so that the capsule will form a smooth cylindrical "piston" inside the pipette as it is aspirated. Because of the high flexibility of the membrane, the capsule naturally seals against the tube wall by pressurization even though it does not adhere to the glass. This arrangement maintains perfect axial symmetry and prevents the membrane from folding or buckling. Hence, it is possible to rigorously analyze the mechanics of deformation of the cell body to obtain the crucial "transducer" relation between pipette suction force and the membrane tension applied directly at the perimeter of the adhesive contact. Further, the geometry of the cell throughout the detachment process is predicted which provides accurate specification of the contact angle theta c between surfaces at the perimeter of the contact. A full analysis of red cell capsules during detachment has been carried out; however, it is shown that the shear rigidity of the red cell membrane can often be neglected so that the red cell can be treated as if it were an under filled lipid bilayer vesicle. From the analysis, the mechanical leverage factor (1-cos theta c) and the membrane tension at the contact perimeter are determined to provide a complete description of the local mechanics of membrane separation as functions of large-scale experimental variables (e.g., suction force, contact diameter, overall cell length). In a companion paper (Evans, E., D. Berk, A. Leung, and N. Mohandas. 1990. Biophys. J. 59:849-860), this approach was applied to the study of separation of large regions of adhesive contact formed between red blood cells by monoclonal antibodies and lectins.