Biological cells exhibit nearly transparent characteristics with weak absorption properties in the visible light spectrum, resulting in extremely low optical contrast between cells and the surrounding medium under traditional bright-field microscopy. To enhance imaging contrast, conventional methods rely on chemical staining or fluorescent labeling, introducing exogenous absorption/fluorescence probes to visualize cellular structures. However, these approaches suffer from drawbacks such as phototoxicity, photobleaching, and poor biocompatibility, severely limiting long-term dynamic observation of living cells. Quantitative phase imaging (QPI) utilizes the inherent physical property of cellular phase (thickness) as an endogenous “probe”, resolving cellular thickness, refractive index, and 3D topography with nanoscale accuracy. It provides a new avenue for dynamic observation of living cells and nanoscale biological studies.
As a significant branch of QPI technology, differential phase contrast (DPC) has attracted considerable attention due to its advantages of being non-interferometric and low-cost. However, its theoretical framework relies on the “weak object approximation”, linking intensity images to sample phase through a linear model. This simplified model introduces two fundamental limitations. First, the phase reconstruction result is highly dependent on the precise modeling of the phase transfer function (PTF) under an ideal pupil. In practical optical systems, however, wavefront aberrations couple with the sample phase, leading to significant reconstruction errors. Second, the conventional half-circle illumination suffers from the problem of PTF response cancellation, resulting in the loss of low-frequency phase information and making it difficult to accurately reconstruct the fine structure of weak phase objects. These limitations significantly compromise the robustness of DPC in non-ideal optical environments and restrict its practical applicability in frontier biological research, such as cellular morphology characterization and tracking of subcellular dynamic processes.
