Genetically Encodable Contrast Agents for Optical Coherence Tomography.

Optical coherence tomography (OCT) has gained wide adoption in biological research and medical imaging due to its exceptional tissue penetration, 3D imaging speed, and rich contrast. However, OCT plays a relatively small role in molecular and cellular imaging due to the lack of suitable biomolecular contrast agents. In particular, while the green fluorescent protein has provided revolutionary capabilities to fluorescence microscopy by connecting it to cellular functions such as gene expression, no equivalent reporter gene is currently available for OCT. Here, we introduce gas vesicles, a class of naturally evolved gas-filled protein nanostructures, as genetically encodable OCT contrast agents. The differential refractive index of their gas compartments relative to surrounding aqueous tissue and their nanoscale motion enables gas vesicles to be detected by static and dynamic OCT. Furthermore, the OCT contrast of gas vesicles can be selectively erased in situ with ultrasound, allowing unambiguous assignment of their location. In addition, gas vesicle clustering modulates their temporal signal, enabling the design of dynamic biosensors. We demonstrate the use of gas vesicles as reporter genes in bacterial colonies and as purified contrast agents in vivo in the mouse retina. Our results expand the utility of OCT to image a wider variety of cellular and molecular processes.

Automatic proximal airway volume segmentation using optical coherence tomography for assessment of inhalation injury.

BACKGROUND: Acute respiratory distress syndrome (ARDS) is a severe form of acute lung injury with a mortality rate of up to 40%. Early management of ARDS has been difficult due to the lack of sensitive imaging tools and robust analysis software. We previously designed an optical coherence tomography (OCT) system to evaluate mucosa thickness (MT) after smoke inhalation, but the analysis relied on manual segmentation. The aim of this study is to assess in vivo proximal airway volume (PAV) after inhalation injury using automated OCT segmentation and correlate the PAV to lung function for rapid indication of ARDS. METHODS: Anesthetized female Yorkshire pigs (n = 14) received smoke inhalation injury (SII) and 40% total body surface area thermal burns. Measurements of PaO2-to-FiO2 ratio (PFR), peak inspiratory pressure (PIP), dynamic compliance, airway resistance, and OCT bronchoscopy were performed at baseline, postinjury, 24 hours, 48 hours, 72 hours after injury. A tissue segmentation algorithm based on graph theory was used to reconstruct a three-dimensional (3D) model of lower respiratory tract and estimate PAV. Proximal airway volume was correlated with PFR, PIP, compliance, resistance, and MT measurement using a linear regression model. RESULTS: Proximal airway volume decreased after the SII: the group mean of proximal airway volume at baseline, postinjury, 24 hours, 48 hours, 72 hours were 20.86 cm (±1.39 cm), 17.61 cm (±0.99 cm), 14.83 cm (±1.20 cm), 14.88 cm (±1.21 cm), and 13.11 cm (±1.59 cm), respectively. The decrease in the PAV was more prominent in the animals that developed ARDS after 24 hours after the injury. PAV was significantly correlated with PIP (r = 0.48, p < 0.001), compliance (r = 0.55, p < 0.001), resistance (r = 0.35, p < 0.01), MT (r = 0.60, p < 0.001), and PFR (r = 0.34, p < 0.01). CONCLUSION: Optical coherence tomography is a useful tool to quantify changes in MT and PAV after SII and burns, which can be used as predictors of developing ARDS at an early stage. LEVEL OF EVIDENCE: Prognostic, level III.

Point-of-care endoscopic optical coherence tomography detects changes in mucosal thickness in ARDS due to smoke inhalation and burns.

BACKGROUND: The prevalence of acute respiratory distress syndrome (ARDS) in mechanically ventilated burn patients is 33%, with mortality varying from 11-46% depending on ARDS severity. Despite the new Berlin definition for ARDS, prompt bedside diagnosis is lacking. We developed and tested a bedside technique of fiberoptic-bronchoscopy-based optical coherence tomography (OCT) measurement of airway mucosal thickness (MT) for diagnosis of ARDS following smoke inhalation injury (SII) and burns. METHODS: 16 female Yorkshire pigs received SII and 40% thermal burns. OCT MT and PaO2-to-FiO2 ratio (PFR) measurements were taken at baseline, after injury, and at 24, 48, and 72h after injury. RESULTS: Injury led to thickening of MT which was sustained in animals that developed ARDS. Significant correlations were found between MT, PFR, peak inspiratory pressure (PIP), and total infused fluid volume. CONCLUSIONS: OCT is a useful tool to quantify MT changes in the airway following SII and burns. OCT may be effective as a diagnostic tool in the early stages of SII-induced ARDS and should be tested in humans.

In vivo cross-sectional imaging of the phonating larynx using long-range Doppler optical coherence tomography.

Diagnosis and treatment of vocal fold lesions has been a long-evolving science for the otolaryngologist. Contemporary practice requires biopsy of a glottal lesion in the operating room under general anesthesia for diagnosis. Current in-office technology is limited to visualizing the surface of the vocal folds with fiber-optic or rigid endoscopy and using stroboscopic or high-speed video to infer information about submucosal processes. Previous efforts using optical coherence tomography (OCT) have been limited by small working distances and imaging ranges. Here we report the first full field, high-speed, and long-range OCT images of awake patients' vocal folds as well as cross-sectional video and Doppler analysis of their vocal fold motions during phonation. These vertical-cavity surface-emitting laser source (VCSEL) OCT images offer depth resolved, high-resolution, high-speed, and panoramic images of both the true and false vocal folds. This technology has the potential to revolutionize in-office imaging of the larynx.

Binding kinetics of biomolecule interaction at ultralow concentrations based on gold nanoparticle enhancement.

Measuring the kinetic constants of protein-protein interactions at ultralow concentrations becomes critical in characterizing biospecific affinity, and exploring the feasibility of clinical diagnosis with respect to detection sensitivity, efficiency and accuracy. In this study, we propose a method that can calculate the binding constants of protein-protein interactions in sandwich assays at ultralow concentrations at the pg/mL level, using a localized surface plasmon coupled fluorescence fiber-optic biosensor (LSPCF-FOB). We discuss a two-compartment model to achieve reaction-limited kinetics under the stagnant conditions of the reaction chamber. The association rate constant, dissociation rate constant, and the equilibrium dissociation constant, that is, k(a), k(d), K(D), respectively, of the kinetics of binding between total prostate-specific antigen (t-PSA) and anti-t-PSA at concentrations from 0.1 pg/mL to 1 ng/mL, were measured either in PBS or in human serum. This is the first time that k(a), k(d), and K(D) have been measured at such a low concentration range in a complex sample such as human serum.

Zeeman laser scanning confocal microscope and its ability on reduction of specimen-induced spherical aberration.

The spherical aberration induced by refractive-index mismatch results in the degradation on the quality of sectioning images in conventional confocal laser scanning microscope (CLSM). In this research, we have derived the theory of image formation in a Zeeman laser scanning confocal microscope (ZLSCM) and conducted experiments in order to verify the ability of reducing spherical aberration in ZLSCM. A Zeeman laser is used as the light source and produces the linearly polarized photon-pairs (LPPP) laser beam. With the features of common-path propagation of LPPP and optical heterodyne detection, ZLSCM shows the ability of reducing the specimen-induced spherical aberration and improving the axial resolution simultaneously.

Kinetics of glucose mutarotation assessed by an equal-amplitude paired polarized heterodyne polarimeter.

A dual-frequency equal-amplitude paired polarization heterodyne polarimeter (DEPHP) was set up in order to precisely measure the mutarotation rate constants of D-glucose in tridistilled water. The DEPHP is based on a balanced detector detection scheme for measurement of the optical rotation angle of D-glucose/water solution during the conversion process between alpha-D-glucose and beta-D-glucose while in a nonequilibrium state. The DEPHP can perform shot-noise-limited detection, so that the total optical rotation angle together with the mutarotation rate constants of alpha-D-glucose and beta-D-glucose conversion can be measured with high sensitivity. In this experiment, the sensitivity of the optical rotation angle measurement was 8.3 x 10(-5) deg/cm, while the total (k), forward (k(1)) and reverse (k(2)) mutarotation rate constants of D-glucose were found to be k = 7.67 x 10(-5) s(-1), k(1) = 2.76 x 10(-5) s(-1), and k(2) = 4.91 x 10(-5) s(-1), respectively, in tridistilled water. Moreover, using the DEPHP, we can measure the specific rotation angles of alpha-D-glucose and beta-D-glucose in water at 632.8 nm. They were 105 degrees and 12 degrees, respectively. Finally, the detection sensitivity of the DEPHP system is also discussed.

Dual-frequency heterodyne ellipsometer for characterizing generalized elliptically birefringent media.

This research proposed a dual-frequency heterodyne ellipsometer (DHE) in which a dual-frequency collinearly polarized laser beam with equal amplitude and zero phase difference between p- and s-polarizations is setup. It is based on the polarizer-sample-analyzer, PSA configuration of the conventional ellipsometer. DHE enables to characterize a generalized elliptical phase retarder by treating it as the combination of a linear phase retarder and a polarization rotator. The method for measuring elliptical birefringence of an elliptical phase retarder based on the equivalence theorem of an unitary optical system was derived and the experimental verification by use of DHE was demonstrated too. The experimental results show the capability of DHE on characterization of a generalized phase retardation plate accurately.

Analog differential-phase detection in optical coherence reflectometer.

A novel differential-phase optical coherence reflectometer (DP-OCR) was proposed using a low-coherence source, integrating it with an analog differential-phase decoding method. In the experiment, the DP-OCR performed a localized surface profile measurement of an optical grating (1200 lp/mm) and demonstrated its ability to measure the translation speed of a tilted mirror. Experimentally, the resolution of the axial displacement of proposed DP-OCR at 185 pm was demonstrated.

Optical activity measurement by use of a balanced detector optical heterodyne interferometer.

What we believe to be a novel amplitude sensitive optical heterodyne polarimeter in which a Zeeman laser is associated with balanced detector detection was set up. The aim was to measure the optical activity of a quartz crystal with a Cornu depolarizer at high accuracy. The features of this novel polarimeter, which include the use of a two-frequency laser that ensures the accuracy of the measurement, are discussed. Furthermore, the detection sensitivity of the optical activity of a quartz crystal was measured as 8.5x10(-10). To our knowledge, this is the highest sensitivity obtained for optical activity measurement of a quartz crystal when the error of the measurement is also analyzed.