Capturing cell morphology dynamics with high temporal resolution using single-shot quantitative phase gradient imaging

• Published in: Journal of biomedical optics Vol. 29; no. S2; p. S22712
• Main Authors: Hur, Sun Woong, Kwon, Minsung, Manoharaan, Revathi, Mohammadi, Melika Haji, Samuel, Ashok Zachariah, Mulligan, Michael P., Hergenrother, Paul J., Bhargava, Rohit
• Format: Journal Article
• Language: English
• Published: Society of Photo-Optical Instrumentation Engineers 01.06.2024
• Subjects: Special Issue Honoring Gabriel Popescu, Pioneer in Biomedical Optics
• ISSN: 1083-3668; 1560-2281; 1560-2281
• DOI: 10.1117/1.JBO.29.S2.S22712

Label-free quantitative phase imaging can potentially measure cellular dynamics with minimal perturbation, motivating efforts to develop faster and more sensitive instrumentation. We characterize fast, single-shot quantitative phase gradient microscopy (ss-QPGM) that simultaneously acquires multiple polarization components required to reconstruct phase images. We integrate a computationally efficient least squares algorithm to provide real-time, video-rate imaging (up to 75   frames / s ). The developed instrument was used to observe changes in cellular morphology and correlate these to molecular measures commonly obtained by staining.SignificanceLabel-free quantitative phase imaging can potentially measure cellular dynamics with minimal perturbation, motivating efforts to develop faster and more sensitive instrumentation. We characterize fast, single-shot quantitative phase gradient microscopy (ss-QPGM) that simultaneously acquires multiple polarization components required to reconstruct phase images. We integrate a computationally efficient least squares algorithm to provide real-time, video-rate imaging (up to 75   frames / s ). The developed instrument was used to observe changes in cellular morphology and correlate these to molecular measures commonly obtained by staining.We aim to characterize a fast approach to ss-QPGM and record morphological changes in single-cell phase images. We also correlate these with biochemical changes indicating cell death using concurrently acquired fluorescence images.AimWe aim to characterize a fast approach to ss-QPGM and record morphological changes in single-cell phase images. We also correlate these with biochemical changes indicating cell death using concurrently acquired fluorescence images.Here, we examine nutrient deprivation and anticancer drug-induced cell death in two different breast cell lines, viz., M2 and MCF7. Our approach involves in-line measurements of ss-QPGM and fluorescence imaging of the cells biochemically labeled for viability.ApproachHere, we examine nutrient deprivation and anticancer drug-induced cell death in two different breast cell lines, viz., M2 and MCF7. Our approach involves in-line measurements of ss-QPGM and fluorescence imaging of the cells biochemically labeled for viability.We validate the accuracy of the phase measurement using a USAF1951 pattern phase target. The ss-QPGM system resolves 912.3    lp / mm , and our analysis scheme accurately retrieves the phase with a high correlation coefficient ( ∼ 0.99 ), as measured by calibrated sample thicknesses. Analyzing the contrast in phase, we estimate the spatial resolution achievable to be 0.55    μ m for this microscope. ss-QPGM time-lapse live-cell imaging reveals multiple intracellular and morphological changes during biochemically induced cell death. Inferences from co-registered images of quantitative phase and fluorescence suggest the possibility of necrosis, which agrees with previous findings.ResultsWe validate the accuracy of the phase measurement using a USAF1951 pattern phase target. The ss-QPGM system resolves 912.3    lp / mm , and our analysis scheme accurately retrieves the phase with a high correlation coefficient ( ∼ 0.99 ), as measured by calibrated sample thicknesses. Analyzing the contrast in phase, we estimate the spatial resolution achievable to be 0.55    μ m for this microscope. ss-QPGM time-lapse live-cell imaging reveals multiple intracellular and morphological changes during biochemically induced cell death. Inferences from co-registered images of quantitative phase and fluorescence suggest the possibility of necrosis, which agrees with previous findings.Label-free ss-QPGM with high-temporal resolution and high spatial fidelity is demonstrated. Its application for monitoring dynamic changes in live cells offers promising prospects.ConclusionsLabel-free ss-QPGM with high-temporal resolution and high spatial fidelity is demonstrated. Its application for monitoring dynamic changes in live cells offers promising prospects.