Label-free single-shot imaging with on-axis phase-shifting holographic reflectance quantitative phase microscopy

Quantitative phase microscopy (QPM) has been emerged as an indispensable diagnostic and characterization tool in biomedical imaging with its characteristic nature of label-free, noninvasive, and real time imaging modality. The integration of holography to the conventional microscopy opens new advancements in QPM featuring high-resolution and quantitative three-dimensional image reconstruction. However, the holography schemes suffer in space-bandwidth and time-bandwidth issues in the off-axis and phase-shifting configuration, respectively. Here, we introduce an on-axis phase-shifting holography based QPM system with single-shot imaging capability. The technique utilizes the Fizeau interferometry scheme in combination with polarization phase-shifting and space-division multiplexing to achieve the single-shot recording of the multiple phase-shifted holograms. Moreover, the high-speed imaging capability with instantaneous recording of spatially phase shifted holograms offers the flexible utilization of the approach in dynamic quantitative phase imaging with robust phase stability. We experimentally demonstrated the validity of the approach by quantitative phase imaging and depth-resolved imaging of paramecium cells. Furthermore, the technique is applied to the phase imaging and quantitative parameter estimation of red blood cells. This integration of a Fizeau-based phase-shifting scheme to the optical microscopy enables a simple and robust tool for the investigations of engineered and biological specimen with real-time quantitative analysis.

Automated summary

Quantitative phase microscopy (QPM) is an important tool in biomedical imaging, providing label-free, noninvasive, and real-time imaging. This study introduces an on-axis phase-shifting holography based QPM system with single-shot imaging capability, demonstrating its effectiveness through experimental validation.