We develop new computational and adaptive optics approaches to push the speed and imaging depth limits of optical microscopy, and OCT-based methods for 4D imaging of biomechanical cell-matrix interactions that are of great interest in the burgeoning field of Cell Mechanics and Mechanobiology. A brief description and highlights from the main areas of our research can be found below.

Computational image formation and adaptive optics for faster and deeper volumetric microscopy

The spatiotemporal coverage of an imaging technology plays a key role in the information that can be acquired about the dynamic interactions within 3D biological systems or tissues. High-throughput cellular-resolution imaging with large volumetric coverage is desirable for many biological studies. This can be beneficial for studying collective (emergent) behavior of cell populations in both engineered biological systems and in vivo models of diseases. In many biological tissues, imaging depth is limited by optical scattering to the superficial layers that are within 1-2 mm of the tissue surface. Extending imaging depth into scattering biological samples and tissues can be considered one of the ‘grand challenges’ of optical microscopy.

From an imaging science perspective, we seek out new ways to integrate computational image formation and hardware adaptive optics (HAO), in order to split the ‘work’ of image formation between computation and optical hardware and do better than each method alone. We developed hybrid adaptive optics (hyAO) to enhance volumetric throughput of optical coherence microscopy (OCM) by addressing both the resolution and signal-collection penalties suffered at away-from-focus depths. We have also utilized hyAO to suppress the effects of multiple scattering and speckle via aberration-diverse OCT, which acquires multiple measurements of the sample with different known aberration states (applied with hardware AO); after computational adaptive optics (CAO) correction of the known aberrations, ballistic scattered signals at each voxel in the image coherently add up in phase, whereas the multiply-scattered signals get randomized by the different interactions that each aberrated illumination beam has with the scattering medium.

High-throughput volumetric OCM of NIH-3T3 fibroblast cell population in Matrigel. Computed OCM enables volumetric microscopy at a 2-μm isotropic resolution over a 1-mm³ volumetric field-of-view, allowing minute-scale cellular dynamics to be captured at 3-minute temporal sampling.

For further information see our Editor’s Pick paper Liu et al., Biomed. Opt. Express, 2018 on high-throughput volumetric OCM, our aberration diverse OCT paper Liu et al., Biomed. Opt. Express, 2018,  and Wu et al., J. Biomed. Opt., 2019 on computed OCM in mouse brain.


Hybrid adaptive optics for multimodal microscopy

Hybrid adaptive optics (hyAO), which harnesses the benefits of both hardware and computational adaptive optics (CAO), can be applied to multimodal imaging by integrating OCT with other imaging modalities. In particular, we seek to leverage the ultra-deep aberration sensing capabilities of long-wavelength optical coherence microscopy (OCM) to rapidly sense wavefront aberrations deep in the mouse brain, and then to leverage the close connection between hardware and computational adaptive optics to enable adaptive optics three-photon microscopy (AO-3PM) image faster and deeper in the mouse and adult zebrafish brains. This research area is a collaboration with the Xu Research Group.

For further information see Liu et al., Proc. SPIE 11630, 2021 for a presentation on wavefront sensing and correction in mouse brain by CAO-OCM, and Yang et al., Proc. SPIE 1164817, 2021 for recent results from our multimodal microscopy system that combines long-wavelength OCM, 3PM and third-harmonic generation (THG) microscopy.


Traction force optical coherence microscopy for volumetric time-lapse imaging of cell forces

Cellular traction forces (CTFs) play a role in many physiological and disease processes, including cell migration, cancer metastasis, stem cell differentiation, wound healing, and synapse formation. Traction force microscopy (TFM) is a family of optical techniques used to quantify CTF. Typical methods for 3D TFM are based on confocal fluorescence microscopy, and can suffer from limited penetration depth, slow acquisition speeds, and photobleaching/phototoxicity concerns. Our lab developed traction force optical coherence microscopy (TF-OCM) to overcome these challenges and enable 4D TFM for the study of both single cells and multi-cellular constructs in scattering media. We collaborate with the Fischbach Lab to apply our new imaging capabilities to study biomechanical cell-matrix interactions in physiologically-relevant tumor spheroid systems.

Computational 4D-OCM of collective cell invasion into surrounding collagen matrix by adipose stromal cell spheroid. Top row: depth projection of traditional OCM image (left), temporal speckle contrast delineating green ‘cell’ and gray-scale ‘collagen’ channels (middle), and depth-to-color projection of cell channel. Bottom row: 3D rendering and 3D rendering with cutaway of cell channel.

For further information see Mulligan et al., Sci. Rep., 2019 on quantitative 3D TF-OCM and Mulligan et al., Sci. Rep., 2021 on 4D imaging of collective cell invasion and force-mediated matrix deformation.


Optical coherence elastography techniques for high-resolution mechanical microscopy of 3D engineered cell cultures and biological tissues

Biophysical interactions between cell and extracellular matrix (ECM) play an important role in biological processes, including initiation and progression of cancer, stem cell differentiation, morphogenesis, and wound healing. Importantly, cell-ECM interactions have been shown to differ in 2D versus 3D environment, driving the pursuit of cellular-scale studies in the more physiologically relevant 3D environment. However, existing approaches for characterizing ECM mechanical properties are typically limited to bulk mechanical testing or atomic force microscopy (which only probes the 2D surface of the sample). Our lab developed optical coherence elastography (OCE) techniques based on highly localized mechanical excitation provided by photonic and acoustic forces, and ultra-precise displacement detection by phase-sensitive OCT.

3D mechanical microscopy of side-by-side agarose hydrogel with photonic force (PF)-OCE. PF-OCE utilizes photonic force from a weakly-focused laser beam to locally “push” on individual micro-beads embedded in the 3D viscoelastic medium. The force-mediated bead “mechanical response” (i.e., bead motion) is interferometrically detected by phase-sensitive OCT, after compensating for the confounding absorption-mediated “photothermal response” of the medium. A larger bead mechanical response implies a more compliant “microenvironment” around that particular bead.

For further information see Leartprapun et al., Nat. Commun., 2018 on 3D mechanical microscopy with PF-OCE and Leartprapun et al., Biomed. Opt. Express, 2019 on spatial resolution and localized excitation in acoustic radiation force (ARF)-OCE.