White light diffraction phase microscopy for imaging of red blood cells for different storage times

Abstract

In this study, the effects of different storage times on the surface morphology of red blood cells (RBCs) were investigated using white light diffraction phase microscopy (WDPM). Blood samples collected from 10 volunteer and stored for 56 days, were imaged on WDPM at every 7 days without any sample preparation. To obtain the phase profiles of RCBs, first the sample and then the reference interferograms were obtained from the experimental setup. Then, surface profiles were calculated from these interferogram images using Fourier transform (FT). With the experiment performed every 7 days, 10 RBC phase information were obtained from each sample and surface profiles were created. From these profiles, 7 parameters related with RBC morphology (average cell thickness – ACT; mean corpuscular volume – MCV; projected surface area – PSA; total surface area – SA; diameter – D; mean corpuscular haemoglobin – MCH; surface area to volume ratio - SAV) have been calculated. Therefore, changes in the morphology of RBCs during storage were evaluated quantitatively. Additionally, phase calibration target was used to confirm the accuracy of our experimental setup system. From the interferogram images, the depth of the phase target (GRP 9 and element 3) was obtained as 309 nm, in accordance with the produced depth. By this way, the reliability of the WDPM setup is demonstrated. This study suggests that the morphology of RBCs can be quantitatively obtained in a reliable manner at higher resolution with WDPM without sample preparation.