Optical Diffraction Tomography (ODT)
Optical Diffraction Tomography (ODT)
Embedded Files
Our Vision:
We are trying to push the boundaries of Optical Tomography by developing a refractive index-sensitive tomographic imaging technique tailored for thick and optically scattering biological samples. The goal is to reconstruct three-dimensional refractive index distributions with minimal information loss, even in highly heterogeneous or diffusive media. This will be achieved primarily through the development of novel experimental schemes, innovations in inverse problem solving, multi-angle illumination strategies, and advanced computational reconstruction algorithms. Such a technique is expected to provide deep insight into the internal structure of tissues and complex cellular environments without the limitations imposed by staining, sectioning, or phototoxicity.
Lateral Shearing Optical Diffraction Tomography (LS-ODT)
Lateral Shearing Optical Diffraction Tomography (LS-ODT)
Optical Diffraction Tomography (ODT) is a powerful label-free technique for reconstructing the three-dimensional refractive index (RI) distribution of biological samples. While conventional ODT performs well for thin and weakly scattering specimens, its performance degrades significantly for heterogeneous and optically thick samples such as tissues and organoids due to multiple scattering and reduced phase stability.
In this work, we introduce Lateral Shearing Optical Diffraction Tomography (LS-ODT), a novel common-path interferometric ODT approach specifically designed for imaging thick and strongly scattering samples with high temporal stability. The system is based on a Sagnac-type common-path geometry using beam splitters and mirrors, enabling partial lateral shearing off-axis interferometry.
Inspired by the contrast mechanism of Differential Interference Contrast (DIC) microscopy, LS-ODT suppresses multiple-scattering background by ensuring that the interfering wavefronts experience similar scattering distortions. In addition, the use of dynamic speckle illumination significantly enhances spatial phase sensitivity and refractive-index accuracy compared to conventional laser-based ODT systems.
The method is experimentally validated on:
Cell phantoms,
Mouse kidney tissue sections,
Human iPSC-derived brain organoids (both thin and thick samples).
Correlative fluorescence and RI tomography further demonstrate LS-ODT’s ability to support histological and biomedical investigations. This work establishes LS-ODT as a robust and stable platform for quantitative, label-free 3D imaging of complex biological specimens.
Figure: Lateral-Shearing Optical Diffraction Tomography (LS-ODT) for imaging thick biological samples.
Schematic of the LS-ODT system and representative three-dimensional refractive index (RI) reconstructions of biological specimens. Panels (A–H) show cell phantom RI imaging comparing fully coherent and partially coherent illumination. RI maps with orthogonal views are presented for mouse kidney tissue (G) and thin brain organoids (E). Panels (I–J) compare RI tomography of a thick human brain organoid obtained using (a) a commercial ODT system (NanoLive) and (b) LS-ODT approach, demonstrating improved image quality in highly scattering samples.
References:
Azeem Ahmad, Balpreet Singh Ahluwalia “Quantitative differential interference contrast microscopy (QDIC): Common-path, high-speed, single shot and highly sensitive Optical Diffraction Tomography (ODT)” Patent Filed in 2025, Patent Application No. 2513325.7. Contribution: 50%
Paweł Gocłowski, Julianna Winnik, et al., Azeem Ahmad*. "Lateral shearing optical diffraction tomography of brain organoid with reduced spatial coherence." (Under Review, Research). arXiv Link
Gocłowski, Paweł, Julianna Winnik, Vishesh Dubey, Piotr Zdańkowski, Maciej Trusiak, Ujjwal Neogi, Mukesh Varshney, Balpreet S. Ahluwalia, and Azeem Ahmad*. "Correlative common-path refractive index tomography and fluorescence of organoid." In Digital Holography and Three-Dimensional Imaging, pp. DW3C-3. Optica Publishing Group, 2025. Link
Gradient Optical Diffraction Tomography (GODT)
Gradient Optical Diffraction Tomography (GODT)
High-quality Optical Diffraction Tomography requires stable interference, typically achieved using low-coherence illumination and common-path shearing interferometry. However, conventional shearing approaches are restricted to sparse samples because they rely on object-free regions for self-interference. Moreover, many gradient-based imaging techniques require phase integration and axial scanning, leading to quasi-3D reconstructions and increased sensitivity to noise.
In this work, we introduce Gradient Optical Diffraction Tomography (GODT), a rigorous tomographic framework that directly reconstructs the three-dimensional refractive index gradient from phase-gradient measurements, avoiding phase integration and z-scanning altogether.
GODT is built on a polarization diffraction grating based common-path interferometric principle, enabling high phase stability and compatibility with low-coherence illumination. Unlike traditional gradient methods, GODT provides a mathematically consistent inverse scattering formulation for reconstructing the RI derivative in three dimensions.
The method is validated through:
Numerical simulations,
Experiments on nano-printed cell phantoms,
Fixed neural cell samples.
Results show that GODT offers:
Enhanced contrast,
Increased sensitivity to subtle RI variations,
Improved visualization of fine cellular structures compared to standard ODT approaches.
GODT represents a new class of gradient-based tomographic imaging that combines the robustness of common-path interferometry with rigorous 3D reconstruction, opening new possibilities for differential RI imaging of complex biological specimens.
References:
Winnik, Julianna, Piotr Zdańkowski, Marzena Stefaniuk, Azeem Ahmad, Chao Zuo, Balpreet S. Ahluwalia, and Maciej Trusiak. "Gradient optical diffraction tomography." Communications Physics 8, 510 (2025) Link
Current Projects
Single-Objective Reflection-Mode Optical Diffraction Tomography (SO-rODT)
Single-Objective Reflection-Mode Optical Diffraction Tomography (SO-rODT)
SO-rODT is a next-generation three-dimensional refractive-index (RI) imaging platform developed to overcome key limitations of conventional transmission-based optical diffraction tomography (ODT). Unlike existing systems that rely on light passing through transparent samples, SO-rODT operates in reflection mode using a single microscope objective. This design enables volumetric imaging of both transparent and opaque specimens with improved flexibility and accessibility.
The reflection-mode architecture enhances axial resolution and phase sensitivity while maintaining easy sample handling and straightforward system alignment. By eliminating the need for a dual-objective configuration typical of transmission ODT, SO-rODT reduces experimental complexity and expands compatibility with a broader range of samples.
A major strength of SO-rODT lies in its ability to capture both forward- and backward-scattered light. This expanded collection of scattering information provides broader spatial-frequency coverage and significantly reduces elongation artifacts that often limit depth resolution in conventional systems. As a result, SO-rODT is particularly well suited for opaque specimens, for which transmission-based techniques are unable to provide RI tomography.
The platform is designed for wide applicability across multiple disciplines. In the life sciences, SO-rODT enables label-free three-dimensional RI imaging of complex biological samples with high phase sensitivity, supporting investigations of cellular structure, tissue organization, and dynamic biological processes without the need for fluorescent markers. At the same time, its reflection-mode operation makes it naturally compatible with opaque materials, opening new opportunities for imaging semiconductor wafers, photonic components, and silicon chips—application areas that are beyond the reach of current transmission-based ODT systems.
By combining single-objective operation, reflection-mode imaging, high sensitivity, and broad sample compatibility, SO-rODT establishes a new class of quantitative three-dimensional optical tomography. This unique combination positions the technology as a versatile tool for both advanced biomedical research and industrial inspection, extending quantitative phase imaging into domains not accessible with existing ODT approaches.
References:
Azeem Ahmad, Joaquín Romero Hernández, "SO-rODT: Single-Objective Reflection-Mode Optical Diffraction Tomography" Disclosure of Invention (DOFI) submitted in 2025. Contribution: 80%
Sample z-scanning partial spatial coherence common-path ODT
Sample z-scanning partial spatial coherence common-path ODT
Sample z-scanning partial spatial coherence common-path Optical Diffraction Tomography (ODT) refers to a specialized implementation of ODT designed for high-resolution, label-free 3D imaging of transparent or semi-transparent samples. In this method, the sample is axially scanned along the z-axis to acquire diffraction patterns at different depths, rather than changing the illumination angle as in conventional angular scanning ODT. This approach simplifies the optical setup while still enabling volumetric reconstruction of the sample’s refractive index distribution.
The system operates using a common-path interferometric configuration, where both the reference and object beams travel along nearly identical optical paths. This design significantly enhances phase stability by reducing sensitivity to mechanical vibrations and environmental fluctuations, making it highly suitable for quantitative phase imaging.
To further improve image quality, the system employs partially spatially coherent illumination—typically from a low-coherence light source such as an LED or a filtered laser. Partial spatial coherence helps suppress speckle noise and minimizes multiple scattering effects, resulting in cleaner phase reconstructions, especially in optically thick or inhomogeneous samples.
Together, the integration of z-scanning, partial spatial coherence, and common-path interferometry forms a robust and stable ODT system capable of producing high-fidelity 3D refractive index maps. This approach is particularly valuable for imaging biological specimens, such as single cells and thick and thin tissue sections, with minimal information loss and without the need for labeling or staining.
Angular illumination scanning partial spatial coherence common-path ODT
Angular illumination scanning partial spatial coherence common-path ODT
This is a high-resolution, label-free imaging technique designed to reconstruct the three-dimensional (3D) refractive index distribution of transparent or weakly scattering specimens, such as biological cells and tissues. This specific implementation of ODT combines several key features—angular scanning, partial spatial coherence, and a common-path interferometric setup—each contributing to the robustness, sensitivity, and resolution of the system.
In this approach, angular illumination scanning is used to probe the sample from multiple directions. The illumination angle is varied either mechanically or digitally (e.g., galvo mirrors, DMDs, MEMS or spatial light modulator), while the sample remains stationary. Each illumination angle yields a different 2D projection of the optical field transmitted through or scattered by the sample. These angular projections are then computationally combined using tomographic reconstruction algorithms—typically based on the first Born or Rytov approximations—to recover the full 3D refractive index distribution. Angular scanning is particularly effective for capturing fine structural features and anisotropies in the sample.
The use of partially spatially coherent illumination (as opposed to fully coherent laser light) serves to suppress coherent artifacts such as speckle and unwanted interference fringes, which are common in biological samples with complex microstructures. Sources like filtered lasers or LED arrays with limited spatial coherence help balance the trade-off between coherence (needed for interferometric detection) and image quality, leading to clearer, more artifact-free reconstructions.
The system also employs a common-path interferometric configuration, in which the reference and object beams share nearly identical optical paths. This design offers superior phase stability and robustness to external disturbances such as vibration and air currents. It also simplifies the optical alignment and reduces system complexity, making it well-suited for long-term or high-throughput imaging applications.
Applications
In progress ...
Page updated
Google Sites
Report abuse