Investigation of the Endothelial Cell Glycocalyx using Electrophoresis

Abstract

The electrophoretic mobilities of bovine and porcine endothelial cells were measured by means of a light-scattering technique. Cells were harvested mechanically and by collagenase or trypsin digestion, and measurements were made on both fresh and chemically fixed cells under a range of solution pH, ionic strength, and composition. The effects of adding serum albumin to the buffer solution were also investigated. Electrophoretic mobilities were comparable to those reported for cultured cells, and for bovine cells at pH 7.4 they ranged from -0.69 ±. 01 (μm/s)/ (V/cm) in 0.15 M NaCl to -2.04 ±. 08 (μn/s)/(V/cm) in 0.015 M NaCl. Cells harvested mechanically had the highest mobility and those released by trypsin the lowest. Mobility became positive at pH 3, indicating that the glycocalyx contains more basic than sulphate groups. Calcium reduced the mobility at acid and neutral pH and caused charge reversal at high pH. In the presence of bovine serum albumin (1% w/v), mobility was reduced, demonstrating charge shielding by either changes in conformation of the glycocalyx or direct electrostatic interactions. Fixation with formaldehyde had a small, but significant effect on the electrophoretic mobility of the trypsin- and collagenase-harvested cells.