Quantitative phase imaging verification in large field-of-view lensless holographic microscopy via two-photon 3D...

Large field-of-view (FOV) microscopic imaging (over 100 mm²) with high lateral resolution (1-2 μm) plays a pivotal role in biomedicine and biophotonics, especially within the label-free regime. Lensless digital holographic microscopy (LDHM) is promising in this context but ensuring accurate quantitative phase imaging (QPI) in large FOV LDHM is challenging. While phantoms, 3D printed by two-photon polymerization (TPP), have facilitated testing small FOV lens-based QPI systems, an equivalent evaluation for lensless techniques remains elusive, compounded by issues such as twin-image and beam distortions, particularly towards the detector's edges. Here, we propose an application of TPP over large area to examine phase consistency in LDHM. Our research involves fabricating widefield phase test targets with galvo and piezo scanning, scrutinizing them under single-shot twin-image corrupted conditions and multi-frame iterative twin-image minimization scenarios. By measuring the structures near the detector's edges, we verified LDHM phase imaging errors across the entire FOV, with less than 12% phase value difference between areas. Our findings indicate that TPP, followed by LDHM and Linnik interferometry cross-verification, requires new design considerations for precise large-area photonic manufacturing. This research paves the way for quantitative benchmarking of large FOV lensless phase imaging, enhancing understanding and further development of LDHM technique.