Yuye Ling

Giga-pixel fluorescent imaging over an ultra-large field-of-view using a flatbed scanner

We demonstrate a new fluorescent imaging technique that can screen for fluorescent micro-objects over an ultra-wide field-of-view (FOV) of ~532 cm(2), i.e., 19 cm × 28 cm, reaching a space-bandwidth product of more than 2 billion. For achieving such a large FOV, we modified the hardware and software of a commercially available flatbed scanner, and added a custom-designed absorbing fluorescent filter, a two-dimensional array of external light sources for computer-controlled and high-angle fluorescent excitation. We also re-programmed the driver of the scanner to take full control of the scanner hardware and achieve the highest possible exposure time, gain and sensitivity for detection of fluorescent micro-objects through the gradient index self-focusing lens array that is positioned in front of the scanner sensor chip. For example, this large FOV of our imaging platform allows us to screen more than 2.2 mL of undiluted whole blood for detection of fluorescent micro-objects within <5 minutes. This high-throughput fluorescent imaging platform could be useful for rare cell research and cytometry applications by enabling rapid screening of large volumes of optically dense media. Our results constitute the first time that a flatbed scanner has been converted to a fluorescent imaging system, achieving a record large FOV.

Phase-noise analysis of swept-source optical coherence tomography systems

We propose a new model to characterize the phase noise in swept-source optical coherence tomography (SS-OCT). The new model explicitly incorporates scanning variability, timing jitter, and sample location in addition to intensity noise (shot noise). The model was analyzed and validated by using both Monte Carlo methods and experiments. We suggest that the proposed model can be used as a guideline for future SS-OCT experimental designs.

Ex vivo visualization of human ciliated epithelium and quantitative analysis of induced flow dynamics by using optical coherence tomography: EX VIVO VISUALIZATION OF HUMAN CILIATED EPITHELIUM

Cilia‐driven mucociliary clearance is an important self‐defense mechanism of great clinical importance in pulmonary research. Conventional light microscopy possesses the capability to visualize individual cilia and its beating pattern but lacks the throughput to assess the global ciliary activities and flow dynamics. Optical coherence tomography (OCT), which provides depth‐resolved cross‐sectional images, was recently introduced to this area.

Highly phase-stable 200 kHz swept-source optical coherence tomography based on KTN electro-optic deflector

The rapid advance in swept-source optical coherence tomography (SS-OCT) technology has enabled exciting new applications in elastography, angiography, and vibrometry, where both high temporal resolution and phase stability are highly sought-after. In this paper, we present a 200 kHz SS-OCT system centered at 1321 nm by using an electro-optically tuned swept source. The proposed systems performance was fully characterized, and it possesses superior phase stability (0.0012% scanning variability and <1 ns timing jitter) that is promising for many phase-sensitive imaging applications. Biological experiments were demonstrated within ex vivo human tracheobronchial ciliated epithelium where both the ciliary motion and ciliary beat frequency were successfully extracted.

Multicontrast endomyocardial imaging by single-channel high-resolution cross-polarization optical coherence tomography

A single-channel high-resolution cross-polarization (CP) optical coherence tomography (OCT) system is presented for multicontrast imaging of human myocardium in one-shot measurement. The intensity and functional contrasts, including the ratio between the cross- and co-polarization channels as well as the cumulative retardation, are reconstructed from the CP-OCT readout. By comparing the CP-OCT results with histological analysis, it is shown that the system can successfully delineate microstructures in the myocardium and differentiate the fibrotic myocardium from normal or ablated myocardium based on the functional contrasts provided by the CP-OCT system. The feasibility of using A-line profiles from the 2 orthogonal polarization channels to identify fibrotic myocardium, normal myocardium and ablated lesion is also discussed.

High-speed phase-stable swept source optical coherence tomography: functional imaging and biomedical applications

High-speed phase-stable swept source optical coherence tomography: functional imaging and biomedical applications

A temporal-frequency variant on robust-principle component analysis for segmentation of motile cilia in optical coherence tomography images (Conference Presentation)

Optical Coherence Tomography (OCT) has established itself as an important tool for studying the role of cilia in Mucociliary Clearance (MCC) due to its ability to observe the cilia’s temporal characteristics over a large field of view. To obtain useful, quantitative measures of this dynamic morphology, the ciliated layer of tissue needs to be segmented from other static components. This is currently accomplished using Speckle Variance processing, a technique whose success relies on subjective thresholding and lacks sensitivity to other sources of speckle noise. We present a modified, frequency constrained, version of Robust Principle Component Analysis (RPCA) which we call Frequency Constrained RPCA (FC-RPCA) as an alternative method for dynamic segmentation of cilia from time-varying OCT B-scans. Based in Sparse Representation theory, FC-RPCA decomposes stacks of images in time into low-rank (static) and a sparse (dynamic) matrices. The sparse matrix represents the segmented cilia layer because of the sparse frequency spectrum exhibited by their characteristic beating pattern. This novel algorithm introduces an additional feature, a user defined frequency constraint on the sparse component, which prevents other sources of speckle noise, like slow moving mucus clouds at the tissues surface, from being segmented with the cilia. The algorithm was used to segment motile cilia in 17 datasets of ex-vivo human ciliated epithelium with high accuracy. Furthermore, FC-RPCA requires no parameter tuning across datasets, demonstrating its capability as a robust tool for processing large volumes of data. When compared with the standard Speckle Variance method, FC-RPCA performed with improved accuracy and selectivity.

Functional endomyocardial imaging by single-channel high resolution cross-polarization OCT (Conference Presentation)

Subcellular resolution is required for OCT to portray the microstructural information of myocardium issue that is comparable to histology. Compare with its intrinsic intensity contrast, functional OCT system may provide contrast related to the tissue composition. We present a high-resolution (HR) cross-polarization OCT system that can provide functional contrast of human myocardium tissue in one-shot measurement. The system is implemented based on our previously reported high-resolution long imaging range OCT system with minimal modification. It features a broadband supercontinuum source, single-channel and one-shot detection, with moderate signal processing. The system has an axial resolution of 3.07 μm, and it is capable to produce accurate polarization information by calibrating the reconstruction performance with a quarter wave plate. The orthogonal polarization channels are multiplexed to fit within one imaging range. Following CP-OCT detection, the retardation can be reconstructed based on the complex signals, and the depolarization effect can be depicted by the channel intensity ratio. Tissue specimens from ten fresh human hearts are used to demonstrate the capability of CP-OCT contrasts. By analyzing the intrinsic and functional OCT contrasts of fresh human myocardium tissues against histology slides, we show that various tissue structures and tissue types of the myocardium, such as fibrosis and ablated lesions, can be better depicted by the function contrasts. We also suggest the possibility of using A-line features from the two orthogonal polarization channels to distinguish normal myocardium, fibrotic myocardium, and ablated lesions. This may serve as a rapid and cost-efficient solution for assessment of myocardium and further facilitate automatic tissue classification.

Visualization of ex vivo human ciliated epithelium and induced flow using optical coherence tomography (Conference Presentation)

The ciliated epithelium is important to the human respiratory system because it clears mucus that contains harmful microorganisms and particulate matter. We report the ex vivo visualization of human trachea/bronchi ciliated epithelium and induced flow characterized by using spectral-domain optical coherence tomography (SD-OCT). A total number of 17 samples from 7 patients were imaged. Samples were obtained from Columbia University Department of Anesthesiology’s tissue bank. After excision, the samples were placed in Gibco Medium 199 solution with oxygen at 4°C until imaging. The samples were maintained at 36.7°C throughout the experiment. The imaging protocol included obtaining 3D volumes and 200 consecutive B-scans parallel to the head-to-feet direction (superior-inferior axis) of the airway, using Thorlabs Telesto system at 1300 nm at 28 kHz A-line rate and a custom built high resolution SDOCT system at 800nm at 32 kHz A-line rate. After imaging, samples were processed with H and E histology. Speckle variance of the time resolved datasets demonstrate significant contrast at the ciliated epithelium sites. Flow images were also obtained after injecting 10μm polyester beads into the solution, which shows beads traveling trajectories near the ciliated epithelium areas. In contrary, flow images taken in the orthogonal plane show no beads traveling trajectories. This observation is in line with our expectation that cilia drive flow predominantly along the superior-inferior axis. We also observed the protective function of the mucus, shielding the epithelium from the invasion of foreign objects such as microspheres. Further studies will be focused on the cilia’s physiological response to environmental changes such as drug administration and physical injury.

Investigating mechanically induced phase response of the tissue by using high-speed phase-resolved optical coherence tomography (Conference Presentation)

Phase-resolved optical coherence tomography (OCT), a functional extension of OCT, provides depth-resolved phase information with extra contrast. In cardiology, changes in the mechanical properties have been associated with tissue remodeling and disease progression. Here we present the capability of profiling structural deformation of the sample in vivo by using a highly stable swept source OCT system The system, operating at 1300 nm, has an A-line acquisition rate of 200 kHz. We measured the phase noise floor to be 6.5 pm±3.2 pm by placing a cover slip in the sample arm, while blocking the reference arm. We then conducted a vibrational frequency test by measuring the phase response from a polymer membrane stimulated by a pure tone acoustic wave from 10 kHz to 80 kHz. The measured frequency response agreed with the known stimulation frequency with an error < 0.005%. We further measured the phase response of 7 fresh swine hearts obtained from Green Village Packing Company through a mechanical stretching test, within 24 hours of sacrifice. The heart tissue was cut into a 1 mm slices and fixed on two motorized stages. We acquired 100,000 consecutive M-scans, while the sample is stretched at a constant velocity of 10 um/s. The depth-resolved phase image presents linear phase response over time at each depth, but the slope varies among tissue types. Our future work includes refining our experiment protocol to quantitatively measured the elastic modulus of the tissue in vivo and building a tissue classifier based on depth-resolved phase information.