SEM imaging can be used to quantify RBC shape. RBCs are sampled from fresh blood samples or from packed RBC units during storage. RBCs are resuspended into PBS to make up 500 μL of solution at 5% haematocrit. RBCs are then fixed by progressively adding a concentrated 2% glutaraldehyde solution in the cell suspension. The cells are incubated for 30 min at RT and in the dark, before being centrifuged and washed in PBS. This fixation protocol was established in order to limit RBC shape changes due to the presence of glutaraldehyde.After fixation, a RBC suspension at 2.5% haematocrit is adhered to coverslips coated with poly-D-lysine. A second fixation step is realised on the adhered RBCs by incubating the coverslips in osmium tetroxide in cacodylate buffer (1%) for one hour. The coverslips are then dried by incubating them in an ascending series of ethanol, then by incubating them with hexamethyldisilizane (HMDS) for 30 min. The HMDS incubation step is repeated twice before the coverslips are dried in open air. Finally, the coverslips are gold coated and imaged with a Zeiss Sigma FESEM.