Time-lapse imaging using dual-color coded quantitative differential phase contrast microscopy.

Abstract

.Significance: Quantitative differential phase contrast (qDPC) microscopy enhances phase contrast by asymmetric illumination using partially coherent light and multiple intensity measurements. However, for live cell imaging, motion artifacts and image acquisition time are important issues. For live cell imaging, a large number of intensity measurements can limit the imaging quality and speed. The minimum number of intensity measurements in qDPC can greatly enhance performance for live imaging.Aim: To obtain high-contrast, isotropic qDPC images with two intensity measurements and perform time-lapse imaging of biological samples.Approach: Based on the color-coded design, a dual-color linear-gradient pupil is proposed to achieve isotropic phase contrast response with two intensity measurements. In our method, the purpose of designing a dual-color coded pupil is twofold: first, to obtain a linear amplitude gradient for asymmetric illumination, which is required to get a circular symmetry of transfer function, and second, to reduce the required number of frames for phase retrieval.Results: To demonstrate the imaging performance of our system, standard microlens arrays were used as samples. We performed time-lapse quantitative phase imaging of rat astrocytes under a low-oxygen environment. Detailed morphology and dynamic changes such as the apoptosis process and migration of cells were observed.Conclusions: It is shown that dual-color linear-gradient pupils in qDPC can outperform half-circle and vortex pupils, and isotropic phase transfer function can be achieved with only two-axis measurements. The reduced number of frames helps in achieving faster imaging speed as compared to the typical qDPC system. The imaging performance of our system is evaluated by time-lapse imaging of rat astrocytes. Different morphological changes in cells during their life cycle were observed in terms of quantitative phase change values.