Influence of Chemical and Genetic Manipulations on Cellular Organelles Quantified by Label-Free Optical Diffraction Tomography

Label-free optical diffraction tomography provides three-dimensionalimaging of cells and organelles, along with their refractive index(RI) and volume. These physical parameters are valuable for quantitativeand accurate analysis of the subcellular microenvironment and itsconnections to intracellular biological properties. In biologicaland biochemical cell analysis, various invasive cell manipulationsare used, such as temperature change, chemical fixation, live cellstaining with fluorescent dye, and gene overexpression of exogenousproteins. However, it is not fully understood how these various manipulationsaffect the physicochemical properties of different organelles. Inthis study, we investigated the impact of these manipulations on thecellular properties of single HeLa cells. We found that after cellfixation and an increase in temperature, the RI value of organelles,such as the nucleus and cytoplasm, significantly decreased overall.Interestingly, unlike the cell nuclei, cytoplasmic RI values werehardly detected after membrane permeation, indicating that only intracytoplasmiccomponents were largely lost. Additionally, our findings revealedthat the expression of GFP and GFP-tagged proteins significantly increasedthe RI values of organelles in living cells compared to the less effectiveRI changes observed with chemical fluorescence staining for cell organelles.The result demonstrates that distinct types of invasive manipulationscan alter the microenvironment of organelles in different ways. Ourstudy sheds new light on how chemical and genetic manipulations affectorganelles.

Automated summary

Label-free optical diffraction tomography allows three-dimensional imaging of cells and organelles, providing valuable physical parameters such as refractive index and volume for quantitative analysis. The impact of various invasive cell manipulations, such as temperature change and chemical fixation, on cellular properties was investigated in this study. It was found that these manipulations can significantly alter the refractive index values of organelles, and the expression of GFP-tagged proteins had a larger effect compared to chemical fluorescence staining.