In recent years, the dynamic role of Lipid Droplets (LDs) in many cellular activities has been increasingly brought to light. In fact, it has been discovered that LDs are involved in many pathologies (e.g., diabetes, atherosclerosis, pathogen infections, neurodegenerative diseases and cancer). Moreover, it has been demonstrated that their number and size increase during an inflammation or infectious inside the immune cells, also with the COVID-19. Therefore, detecting LDs within single cells could aid the diagnosis of several pathologies. Currently, the gold-standard technique in this field is Fluorescence Imaging Flow Cytometry (FIFC), in which the single-cell analysis of fluorescence microscopy is implemented in high-throughput modality thanks to the flow-cytometry module. However, to overcome the drawbacks related to the fluorescence staining, Holographic Imaging Flow Cytometry (HIFC) has gaining momentum as label-free alternative to the FIFC tool. Thanks to the interferometric principles at the basis of digital holography, it has been already demonstrated that a suspended cell acts as a biological lens with specific focusing features. Here we show that the presence of intracellular LDs inside the cell is able to change its focalization features, measured through a HIFC system. Therefore, based on this property, we demonstrate that a detection of single cells containing intracellular LDs is possible by means of a direct analysis of the digital holograms recorded in flow cytometry modality. The attained results open the route to the development of a fast, non-destructive, and high-throughput tool for the diagnosis of LDs-related pathologies by exploiting the biolens’ signature in HIFC.
Detection and quantification of intracellular structures is fundamental in biomedical sciences. New emerging inspection tools based on holographic microscopy and quantitative phase imaging can give answers to such critical demands. Holographic tomography (HT) systems are the best candidates for this challenge. Recently, HT has been demonstrated working in flow-cytometry (FC) modality. Results show that the novel HTFC tool is capable to furnish 3D visualization and quantifications of the different intracellular particles. In particular, here we report that exogenous nanographene oxide particles as well as endogenous lipid droplets can be detected, measured, and visualized in each flowing cell by label-free HTFC. This method opens the way for accurate and high-throughput measurements at the 3D single-cell level for different applications such as diagnosis of diseases, development of drug delivery applications, and examination of cell functionalities. Experiments and processing methods will be described, and several examples will be discussed.
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