Tonicity induced changes in volume and refractive index of suspended cells quantified with digital holographic microscopy

Kemper, Björn; Klokkers, Jens; Przibilla, Sabine; Vollmer, Angelika; Ketelhut, Steffi; Bally, Gert von; Pavenstädt, Hermann-Joseph; Schlatter, Eberhard; Edemir, Bayram

doi:10.4302/plp.2012.2.02

Cited by 4 publications

(5 citation statements)

References 23 publications

(35 reference statements)

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“…However, they still follow the volume dependent trend as can be seen by fitting the analyzed values leaving all three parameters as open fitting parameters ( n ∝ r −3.2 ). This is also in agreement with previous studies, where such an inverse linear relationship between RI and volume has been reported for entire cells . Taken together, this suggests that the nuclear RI for a given dry mass depends only on the volume it occupies and the consequent material density, and not on the specific conformation of the solid material in the nucleus.…”

Section: Resultssupporting

confidence: 93%

Cell nuclei have lower refractive index and mass density than cytoplasm

et al. 2016

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Common perception regards the nucleus as a densely packed object with higher refractive index (RI) and mass density than the surrounding cytoplasm. Here, the volume of isolated nuclei is systematically varied by electrostatic and osmotic conditions as well as drug treatments that modify chromatin conformation. The refractive index and dry mass of isolated nuclei is derived from quantitative phase measurements using digital holographic microscopy (DHM). Surprisingly, the cell nucleus is found to have a lower RI and mass density than the cytoplasm in four different cell lines and throughout the cell cycle. This result has important implications for conceptualizing light tissue interactions as well as biological processes in cells.

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Section: Resultssupporting

confidence: 93%

Cell nuclei have lower refractive index and mass density than cytoplasm

et al. 2016

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“…The comparison of the average values for refractive index, volume, and dry mass in Figure 4 demonstrates the requirement of repeated independent measurements for statistically reliable parameter acquisition. Moreover, the slightly different slopes may be explained by a different response of the different cell types to the osmotic stimulation which is in agreement with previously reported observations (45). This finding correlates with the relative low values of the corresponding standard errors compared with cell volume and dry mass and identifies the refractive index as the quantity of the accessed biophysical parameter set that is most sensitive to osmolality variations.…”

Section: Discussionsupporting

confidence: 92%

“…5a and 5b). Moreover, the slightly different slopes may be explained by a different response of the different cell types to the osmotic stimulation which is in agreement with previously reported observations (45). However, further studies are required to identify the underlying physiological processes.…”

Section: Discussionsupporting

confidence: 91%

“…In contrast, the calculated dry mass values that quantify the intracellular content show no statistically significant differences. This is in agreement with assumptions of the preservation of intracellular content (39,45) during osmotic stimulation of cells and is supported by the results of the cell swelling and shrinking experiments in Figure 5. The average dry mass was found independent of osmolality changes within the measurement uncertainty while cellular refractive index and volume significantly decreased or increased for cell culture media with decreased or increased osmolality.…”

Section: Discussionsupporting

confidence: 90%

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Quantitative phase imaging for cell culture quality control

Kastl¹,

Isbach²,

Dirksen

et al. 2017

Cytometry Pt A

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The potential of quantitative phase imaging (QPI) with digital holographic microscopy (DHM) for quantification of cell culture quality was explored. Label-free QPI of detached single cells in suspension was performed by Michelson interferometer-based self-interference DHM. Two pancreatic tumor cell lines were chosen as cellular model and analyzed for refractive index, volume, and dry mass under varying culture conditions. Firstly, adequate cell numbers for reliable statistics were identified. Then, to characterize the performance and reproducibility of the method, we compared results from independently repeated measurements and quantified the cellular response to osmolality changes of the cell culture medium. Finally, it was demonstrated that the evaluation of QPI images allows the extraction of absolute cell parameters which are related to cell layer confluence states. In summary, the results show that QPI enables label-free imaging cytometry, which provides novel complementary integral biophysical data sets for sophisticated quantification of cell culture quality with minimized sample preparation. © 2017 International Society for Advancement of Cytometry.

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“…The cell thickness increases nearly by a factor two after changing the osmolality from 320 mOsmol kg −1 to 160 mOsmol kg −1 . This correlates with data from comparative DHM investigations on adherent and suspended cells in which near doubling of the cell volume was observed 35, 36…”

Section: Resultssupporting

confidence: 85%

Three‐Dimensional Exploration and Mechano‐Biophysical Analysis of the Inner Structure of Living Cells

Barroso

Woerdemann

Vollmer

et al. 2012

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A novel mechanobiological method is presented to explore qualitatively and quantitatively the inside of living biological cells in three dimensions, paving the way to sense intracellular changes during dynamic cellular processes. For this purpose, holographic optical tweezers, which allow the versatile manipulation of nanoscopic and microscopic particles by means of tailored light fields, are combined with self-interference digital holographic microscopy. This biophotonic holographic workstation enables non-contact, minimally invasive, flexible, high-precision optical manipulation and accurate 3D tracking of probe particles that are incorporated by phagocytosis in cells, while simultaneously quantitatively phase imaging the cell morphology. In a first model experiment, internalized polystyrene microspheres with 1 μm diameter are three-dimensionally moved and tracked in order to quantify distances within the intracellular volume with submicrometer accuracy. Results from investigations on cell swelling provoked by osmotic stimulation demonstrate the homogeneous stretching of the cytoskeleton network, and thus that the proposed method provides a new way for the quantitative 3D analysis of the dynamic intracellular morphology.

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Tonicity induced changes in volume and refractive index of suspended cells quantified with digital holographic microscopy

Cited by 4 publications

References 23 publications

Cell nuclei have lower refractive index and mass density than cytoplasm

Cell nuclei have lower refractive index and mass density than cytoplasm

Quantitative phase imaging for cell culture quality control

Three‐Dimensional Exploration and Mechano‐Biophysical Analysis of the Inner Structure of Living Cells

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