■无标记定量相位成像可以以最小的扰动潜在地测量细胞动力学,激励努力开发更快、更灵敏的仪器。我们描述快速,单次定量相位梯度显微镜(ss-QPGM),可同时获取重建相位图像所需的多个偏振分量。我们集成了一个计算高效的最小二乘算法来提供实时,视频速率成像(高达75帧/秒)。开发的仪器用于观察细胞形态的变化,并将其与通常通过染色获得的分子测量值相关联。
■我们旨在表征ss-QPGM的快速方法,并记录单细胞相位图像中的形态变化。我们还使用同时获得的荧光图像将这些与指示细胞死亡的生化变化相关联。
■这里,我们研究了两种不同的乳腺细胞系中营养剥夺和抗癌药物诱导的细胞死亡,viz.,M2和MCF7。我们的方法涉及对ss-QPGM进行在线测量,并对生化标记的细胞进行荧光成像。
■我们使用USAF1951模式相位目标验证了相位测量的准确性。ss-QPGM系统的分辨率为912.3lp/mm,并且我们的分析方案准确地检索到具有高相关系数(~0.99)的相位,通过校准样品厚度测量。分析阶段的对比度,我们估计该显微镜可实现的空间分辨率为0.55μm。ss-QPGM延时活细胞成像揭示了生物化学诱导的细胞死亡过程中的多种细胞内和形态学变化。来自定量相位和荧光的共配准图像的推断表明坏死的可能性,这与之前的发现一致。
■演示了具有高时间分辨率和高空间保真度的无标签ss-QPGM。它在监测活细胞动态变化方面的应用前景广阔。
UNASSIGNED: 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.
UNASSIGNED: 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.
UNASSIGNED: 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.
UNASSIGNED: 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.
UNASSIGNED: 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.