## 3 Results and Discussion  
### Evolution of RBC Morphology Characteristics during In-Vitro Storage  
The functionality of a RBC is linked with its cell geometry. The evolution of cell dimensions (i.e., cell length, cell diameter and cell thickness) with respect to in-vitro storage effects can be described through the cell shape factor (_SF_) [76]. The cell is assumed to be a rigid body having uniform density, and the centre of mass of the whole cell and its three-principal axes of inertia are determined for each numerically predicted cell shape at \(t\) (Days) = 0, 7, 14, 21, 23, 25, 28, 35 and 42. \(H_{1}\), \(H_{2}\) and \(H_{3}\) are defined as the distance between the furthest vertex points on the RBC membrane surface along the three-principal axes of inertia of the cell such that \(\text{H}_{1}\ \leq\ \text{H}_{2}\ \leq\ \text{H}_{3}\). _SF_ is given by \(SF=H_{1}\ /\ \sqrt{H_{2}\times H_{3}}\)[76; 80], and indicates the sphericity of the cell. The cell becomes more and more spherical as _SF_ reaches the value 1, and becomes a more and more flattened disc as _SF_ reaches 0. _SF_ at \(t\) (Days) = 0, 7, 14, 21, 23, 25, 28, 35 and 42 was estimated and is presented in Figure 5. The _SF_ of stored RBCs gradually increases at \(0\leq\ t<20\) and at \(26\leq\ t\leq 42\), whereas a sudden increment of _SF_ was observed at \(20\leq\ t<26\). The evolution of _SF_ is comparable to the evolution of cell morphology, \(\mu(t)\) and \(\kappa(t)\) during storage (refer to Figures 1 and 2), and can be attributed primarily to the evolution of cell morphology as \(\mu(t)\) and \(\kappa(t)\) are influenced by the cell morphology as well [8; 47]. It is important to distinguish the morphology characteristics of stored RBCs against other pathophysiological conditions where similar morphologies are observed, for better diagnostic purposes. Each shape-transformation scenario might affect the cell details differently. Therefore, Figure 5 also compares the morphology characteristics of storage lesion-produced discocyte, echinocyte I, echinocyte II, echinocyte III and sphero-echinocyte RBC morphologies against that of echinocytogenic shape-transforming conditions (e.g., amphipaths, extracellular ionic strength and pH) produced RBC morphologies. The CG-RBC membrane model [76] is capable of predicting RBC morphologies under varying shape-transformation scenarios, and [PERSON] et al. [76] discussed the morphology characteristics of discocyte, echinocyte I, echinocyte II, echinocyte III and sphero-echinocyte morphologies of a healthy RBC at \(t\) = 0 (Days), which were obtained via echinocytogenic shape-transforming conditions. The echinocytogenic shape-transforming conditions transform the biconcave discocyte shape into echinocyte forms at constant cell surface area and volume [48; 49; 50; 51; 52; 53; 54]. _SF_ values of discocyte, echinocyte I, echinocyte II, echinocyte III and sphero-echinocyte RBC morphologies obtained via the echinocyte shape-transforming condition (from [PERSON] et al. [76]) are presented in Figure 5 at \(t\) where analogous morphology was observed during storage. Figure 5: Shape factors (_SF_) of CG-RBC membrane model-predicted RBC morphologies during in-vitro storage (discocyte at \(0\leq\ t<20\); echinocyte I at \(20\leq\ t<22\); echinocyte II at \(22\leq\ t<24\); echinocyte III at \(24\leq\ t<26\); and sphero-echinocyte at \(26\leq\ t\leq 42\)), and at echinocytic shape-transforming conditions. The dependency of cell \(SF\) on its morphology was confirmed by the similar behaviour of \(SF\) values for the two considered shape-transforming scenarios; in-vitro storage and echinocytogenic condition. Overall, \(SF\) of stored RBCs is higher than that of echinocytogenic conditions-produced RBCs, which shows the influence of the loss of membrane surface area and cell volume during storage. The cell surface area and volume of the stored RBC gradually decreases during storage, whereas the morphological changes in the presence of echinocytogenic conditions take place at constant cell surface area and volume.