AbstractAs a label-free, nondestructive method, phase contrast is by far the most popular microscopy technique for routine inspection of cell cultures. Yet, features of interest such as extensions near cell bodies are often obscured by a glow, which came to be known as the halo. Advances in modeling image formation have shown that this artifact is due to the limited spatial coherence of the illumination. Yet, the same incoherent illumination is responsible for superior sensitivity to fine details in the phase contrast geometry. Thus, there exists a trade-off between high-detail (incoherent) and low-detail (coherent) imaging systems. In this work, we propose a method to break this dichotomy, by carefully mixing corrected low-frequency and high-frequency data in a way that eliminates the edge effect. Specifically, our technique is able to remove halo artifacts at video rates, requiring no manual interaction ora prioripoint spread function measurements. To validate our approach, we imaged standard spherical beads, sperm cells, tissue slices, and red blood cells. We demonstrate the real-time operation with a time evolution study of adherent neuron cultures whose neurites are revealed by our halo correction. We show that with our novel technique, we can quantify cell growth in large populations, without the need for thresholds and calibration.
Real-time label-free microscopy with adjustable phase-contrast