Marker-free automatic quantification of red blood cell fluctuations with different storage periods by holographic imaging

This paper overviews the methods to quantitatively measure the cell membrane fluctuation (CMF) rate of red blood cells (RBCs) with different storage periods with millisecond temporal sensitivity at the single-cell level by using marker-free holographic imaging techniques. We quantitatively measured fluctuations in the discocyte shape of RBCs membrane, ring and dimple in the case of storage lesion with time-lapse phase images of RBCs. Our experimental results demonstrate that normal RBCs with a discocyte shape become stiffer with storage period and there is a significant negative correlation between CMFs and the sphericity coefficient, which describes the RBCs morphology. Correlation between CMF and projected surface area is also performed. © 2019 SPIE

Quantification of stored red blood cell fluctuations by time-lapse holographic cell imaging.

We propose methods to quantitatively calculate the fluctuation rate of red blood cells with nanometric axial and millisecond temporal sensitivity at the single-cell level by using time-lapse holographic cell imaging. For this quantitative analysis, cell membrane fluctuations (CMFs) were measured for RBCs stored at different storage times. Measurements were taken over the whole membrane for both the ring and dimple sections separately. The measurements show that healthy RBCs that maintain their discocyte shape become stiffer with storage time. The correlation analysis demonstrates a significant negative correlation between CMFs and the sphericity coefficient, which characterizes the morphological type of erythrocyte. In addition, we show the correlation results between CMFs and other morphological properties such as projected surface area, surface area, mean corpuscular volume, and mean corpuscular hemoglobin

Red blood cells ageing markers: a multi-parametric analysis : Red blood cells ageing markers: a multi-parametric analysis

Background. Red blood cells collected in citrate-phosphate-dextrose can be stored for up to 42 days at 4 degrees C in saline-adenine-glucose-mannitol additive solution. During this controlled, but nevertheless artificial, ex vivo ageing, red blood cells accumulate lesions that can be reversible or irreversible upon transfusion. The aim of the present study is to follow several parameters reflecting cell metabolism, antioxidant defences, morphology and membrane dynamics during storage. Materials and methods. Five erythrocyte concentrates were followed weekly during 71 days. Extracellular glucose and lactate concentrations, total antioxidant power, as well as reduced and oxidised intracellular glutathione levels were quantified. Microvesiculation, percentage of haemolysis and haematologic parameters were also evaluated. Finally, morphological changes and membrane fluctuations were recorded using label-free digital holographic microscopy. Results. The antioxidant power as well as the intracellular glutathione concentration first increased, reaching maximal values after one and two weeks, respectively. Irreversible morphological lesions appeared during week 5, where discocytes began to transform into transient echinocytes and finally spherocytes. At the same time, the microvesiculation and haemolysis started to rise exponentially. After six weeks (expiration date), intracellular glutathione was reduced by 25%, reflecting increasing oxidative stress. The membrane fluctuations showed decreased amplitudes during shape transition from discocytes to spherocytes. Discussion. Various types of lesions accumulated at different chemical and cellular levels during storage, which could impact their in vivo recovery after transfusion. A marked effect was observed after four weeks of storage, which corroborates recent clinical data. The prolonged follow-up period allowed the capture of deep storage lesions. Interestingly, and as previously described, the severity of the changes differed among donors