Dynamic phase differences based on quantitative phase imaging for the objective evaluation of cell behavior

Abstract

Quantitative phase imaging (QPI) brought innovation to noninvasive observation of live cell dynamics seen as cell behavior. Unlike the Zernike phase contrast or differential interference contrast, QPI provides quantitative information about cell dry mass distribution. We used such data for objective evaluation of live cell behavioral dynamics by the advanced method of dynamic phase differences (DPDs). The DPDs method is considered a rational instrument offered by QPI. By subtracting the antecedent from the subsequent image in a time-lapse series, only the changes in mass distribution in the cell are detected. The result is either visualized as a two dimensional color-coded projection of these two states of the cell or as a time dependence of changes quantified in picograms. Then in a series of time-lapse recordings, the chain of cell mass distribution changes that would otherwise escape attention is revealed. Consequently, new salient features of live cell behavior should emerge. Construction of the DPDs method and results exhibiting the approach are presented. Advantage of the DPDs application is demonstrated on cells exposed to an osmotic challenge. For time-lapse acquisition of quantitative phase images, the recently developed coherence-controlled holographic microscope was employed.

Quantitative phase imaging unravels new insight into dynamics of mesenchymal and amoeboid cancer cell invasion

“…Moreover, postprocessing of images acquired by CCHM enables analysis of dynamic changes of cell mass distribution in migrating cells by calculating the dynamic phase differences (DPD) between consequent images (see Methods for more detail). The advantages of this method were demonstrated previously 33 . Analysis of cell mass differences during mesenchymal invasion highlighted the dynamics of membrane protrusions at the cell front and revealed a large influx of cell mass into the leading edge, clearly demonstrating the polarization of the cell, which corresponds to directionality of cell migration (Fig.…”

“…Nevertheless, QPI techniques have already yielded several findings of cellular behaviour 56 . For example, dynamic phase differences were employed for the visualization of changes in distribution of cell dry mass upon nutritional stress 57 , an approach, which was later elaborated as an evaluation method 33 . QPI also permits to differentiate between cell death caused by apoptosis and oncosis 58 or reveal necrosis 59 .…”

Automated classification of cell morphology by coherence-controlled holographic microscopy

“…In recent years, digital holographic microscopy (DHM) has proven to be a very versatile noninvasive tool for the observation of live cells, [14][15][16][17] while overcoming the limitations of the previously mentioned approaches. DHM provides quantitative phase images (QPIs) with high intrinsic contrast without labeling and since the images contain quantitative information about cell mass, it may potentially improve the performance of the classification.…”

Distinctive behaviour of live biopsy-derived carcinoma cells unveiled using coherence-controlled holographic microscopy

“…Our development of coherence-controlled holographic microscopy (CCHM) has offered us an opportunity to examine the reactions of live cells. CCHM quantitative phase imaging (QPI), which can be feasibly exploited even through turbid media [ 8 ] to measure tiny differences in cell motion [ 9 ], presents an innovative objective analysis of live cancer cells in vitro achieved by simultaneously imaging the cell shape and position and measuring changes in the cell mass, i.e., growth. Indeed, CCHM in a Q-PHASE microscope (Tescan Orsay Holding, a.s., Brno, Czech Republic) recently contributed to the differentiation of oncosis from apoptosis [ 10 ] and the recognition of entosis, which emerged as a particularly important type of ostensible cell death but is potentially required for tumour survival [ 11 ].Head and neck squamous cell carcinoma (HNSCC) is a pressing clinical problem.…”