Automated Quantification and Statistical Characterization of 3D Morphological Parameters of Red Blood Cells and Blood Coagulation Structures Using Flow Cytometry with Digital Holographic Microscopy

Label-free, high-throughput, and 3D morphological analysis of blood cells remains a major challenge in biomedical optics. In this study, we investigate this issue using flow cytometry with digital holographic microscopy (DHM) to enable real-time, label-free imaging of red blood cells (RBCs) and blood coagulation structures (BCSs) without the need for staining or chemical pretreatment. We demonstrate an approach for the automated quantification and statistical characterization of these cells using quantitative phase information reconstructed from digital holograms. Although established image processing techniques such as phase unwrapping and segmentation are used, this study presents, to the best of our knowledge, the first statistical characterization of the 3D morphological features of BCSs. This is particularly useful in analyzing the heterogeneous and complex 3D structures of BCSs, which are difficult to assess using conventional microscopy. The results suggest that this DHM-based flow cytometry system provides a promising platform for non-invasive, real-time morphological evaluation of blood samples and has potential applications in hematological diagnostics and research related to blood coagulation.