國立虎尾科技大學 |

Wide-Field Mid-Infrared Photothermal Microscopy.

Record Type:	Language materials, manuscript : Monograph/item
Title/Author:	Wide-Field Mid-Infrared Photothermal Microscopy./
Author:	Zong, Haonan.
Description:	1 online resource (182 pages)
Notes:	Source: Dissertations Abstracts International, Volume: 85-07, Section: B.
Contained By:	Dissertations Abstracts International85-07B.
Subject:	Optics. -
Online resource:	click for full text (PQDT)
ISBN:	9798381407365

Wide-Field Mid-Infrared Photothermal Microscopy.
Zong, Haonan.

Wide-Field Mid-Infrared Photothermal Microscopy. - 1 online resource (182 pages)

Source: Dissertations Abstracts International, Volume: 85-07, Section: B.

Thesis (Ph.D.)--Boston University, 2024.

Includes bibliographical references

Infrared (IR) imaging has been a widely used method for chemical imaging of a variety of samples. Traditional infrared imaging method that is directly measuring IR absorption such as Fourier transform infrared (FTIR) microscopy has limited resolution due to the intrinsic long wavelength of the IR beam. Atomic force microscope-infrared (AFM-IR) spectroscopy was developed to break the IR diffraction-limited resolution of FTIR. However, it has limited speed and is only applicable for dry and flat samples. In recent years, mid-infrared photothermal (MIP) microscopy was invented to overcome the limitations of FTIR and AFM-IR. The first developed point-scanning-based MIP achieved sub-micron resolution and depth-resolved IR imaging of live biological samples. Later, wide-field-based MIP schemes were developed and dramatically improved the imaging speed of MIP. In Chapter One, this dissertation introduces the MIP development background and the MIP signal formation mechanism for general samples. In Chapters Two, Three, and Four, this dissertation further developed wide-field MIP and achieved optimal detection of three different specific types of samples by three different methodologies, which are described as follows. (1) Chapter Two, bond-selective interferometric scattering microscopy. (2) Chapter Three, back-ground suppressed high-throughput mid-infrared photothermal microscopy via pupil engineering. (3) Chapter Four, bond-selective full-field optical coherence tomography. In Chapter Five, this dissertation summarizes the work and provides an outlook of future work on video-rate bond-selective intensity diffraction tomography. In the first methodology, we describe a bond-selective interferometric scattering microscope where the mid-IR induced photothermal signal is detected by a visible beam in a wide-field common-path interferometry configuration. Single-particle interferometric reflectance imaging sensor (SP-IRIS) has been a very promising technology for highly sensitive label-free imaging of a broad spectrum of biological nanoparticles from proteins to viruses in a high-throughput manner. Although it can reveal the specimen's size and shape information, the chemical composition is inaccessible in interferometric measurements. IR spectroscopic imaging provides chemical specificity based on inherent chemical bond vibrations of specimens but lacks the ability to image and resolve individual nanoparticles due to long IR wavelengths. Here, a bond-selective interferometric scattering microscope is achieved by detecting the mid-IR-induced photothermal modulation of a visible beam in a wide-field common-path interferometry configuration. A thin film layered substrate is utilized to reduce the reflected light and provide a reference field for the interferometric detection of the weakly scattered field. A pulsed mid-IR laser is employed to modulate the interferometric signal. Subsequent demodulation via a virtual lock-in camera offers simultaneous chemical information about tens of micro- or nano-particles. The chemical contrast arises from a minute change in the particle's scattered field as a consequence of the vibrational absorption at the target molecule. We characterize the system with sub-wavelength polymer beads and highlight biological applications by chemically imaging several microorganisms including Staphylococcus aureus, Escherichia coli, and Candida albicans. A theoretical framework is established to extend bond-selective interferometric scattering microscopy to a broad range of biological micro- and nano-particles.In the second methodology, we demonstrate a back-ground suppressed high-throughput mid-infrared photothermal microscopy via pupil engineering. MIP microscopy has been a promising label-free chemical imaging technique for functional characterization of specimens owing to its enhanced spatial resolution and high specificity. Recently developed wide-field MIP imaging modalities have drastically improved speed and enabled high-throughput imaging of micron-scale subjects. However, the weakly scattered signal from subwavelength particles becomes indistinguishable from the shot-noise as a consequence of the strong background light, leading to limited sensitivity. Here, we demonstrate background-suppressed chemical fingerprinting at a single nanoparticle level by selectively attenuating the reflected light through pupil engineering in the collection path. Our technique provides over 3 orders of magnitude background suppression by quasi-darkfield illumination in the epi-configuration without sacrificing lateral resolution. We demonstrate 6-fold signal-to-background noise ratio improvement, allowing for simultaneous detection and discrimination of hundreds of nanoparticles across a field of view of 70 μm x 70 μm. A comprehensive theoretical framework for photothermal image formation is provided and experimentally validated with 300 and 500 nm PMMA beads. The versatility and utility of our technique are demonstrated via hyperspectral dark-field MIP imaging of S. aureus and E. coli bacteria and MIP imaging of subcellular lipid droplets inside C. albicans and cancer cells.In the third methodology, we present a bond-selective full-field optical coherence tomography. Optical coherence tomography (OCT) is a label-free, non-invasive 3D imaging tool widely used in both biological research and clinical diagnosis. Conventional OCT modalities can only visualize specimen tomography without chemical information. Here, we report a bond-selective full-field OCT (BS-FF-OCT), in which a pulsed mid-infrared laser is used to modulate the OCT signal through the photothermal effect, achieving label-free bond-selective 3D sectioned imaging of highly scattering samples. We first demonstrate BS-FF-OCT imaging of 1 µm PMMA beads embedded in agarose gel. Next, we show 3D hyperspectral imaging of up to 75 µm of polypropylene fiber mattress from a standard surgical mask. We then demonstrate BS-FF-OCT imaging on biological samples, including cancer cell spheroids and C. elegans. Using an alternative pulse timing configuration, we finally demonstrate the capability of BS-FF-OCT on imaging a highly scattering myelinated axons region in a mouse brain tissue slice.

Electronic reproduction.
Ann Arbor, Mich. :
ProQuest,
2024

Mode of access: World Wide Web

ISBN: 9798381407365Subjects--Topical Terms:

595336
Optics.
Subjects--Index Terms:

Infrared imagingIndex Terms--Genre/Form:

554714
Electronic books.

Wide-Field Mid-Infrared Photothermal Microscopy.
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520 $a Infrared (IR) imaging has been a widely used method for chemical imaging of a variety of samples. Traditional infrared imaging method that is directly measuring IR absorption such as Fourier transform infrared (FTIR) microscopy has limited resolution due to the intrinsic long wavelength of the IR beam. Atomic force microscope-infrared (AFM-IR) spectroscopy was developed to break the IR diffraction-limited resolution of FTIR. However, it has limited speed and is only applicable for dry and flat samples. In recent years, mid-infrared photothermal (MIP) microscopy was invented to overcome the limitations of FTIR and AFM-IR. The first developed point-scanning-based MIP achieved sub-micron resolution and depth-resolved IR imaging of live biological samples. Later, wide-field-based MIP schemes were developed and dramatically improved the imaging speed of MIP. In Chapter One, this dissertation introduces the MIP development background and the MIP signal formation mechanism for general samples. In Chapters Two, Three, and Four, this dissertation further developed wide-field MIP and achieved optimal detection of three different specific types of samples by three different methodologies, which are described as follows. (1) Chapter Two, bond-selective interferometric scattering microscopy. (2) Chapter Three, back-ground suppressed high-throughput mid-infrared photothermal microscopy via pupil engineering. (3) Chapter Four, bond-selective full-field optical coherence tomography. In Chapter Five, this dissertation summarizes the work and provides an outlook of future work on video-rate bond-selective intensity diffraction tomography. In the first methodology, we describe a bond-selective interferometric scattering microscope where the mid-IR induced photothermal signal is detected by a visible beam in a wide-field common-path interferometry configuration. Single-particle interferometric reflectance imaging sensor (SP-IRIS) has been a very promising technology for highly sensitive label-free imaging of a broad spectrum of biological nanoparticles from proteins to viruses in a high-throughput manner. Although it can reveal the specimen's size and shape information, the chemical composition is inaccessible in interferometric measurements. IR spectroscopic imaging provides chemical specificity based on inherent chemical bond vibrations of specimens but lacks the ability to image and resolve individual nanoparticles due to long IR wavelengths. Here, a bond-selective interferometric scattering microscope is achieved by detecting the mid-IR-induced photothermal modulation of a visible beam in a wide-field common-path interferometry configuration. A thin film layered substrate is utilized to reduce the reflected light and provide a reference field for the interferometric detection of the weakly scattered field. A pulsed mid-IR laser is employed to modulate the interferometric signal. Subsequent demodulation via a virtual lock-in camera offers simultaneous chemical information about tens of micro- or nano-particles. The chemical contrast arises from a minute change in the particle's scattered field as a consequence of the vibrational absorption at the target molecule. We characterize the system with sub-wavelength polymer beads and highlight biological applications by chemically imaging several microorganisms including Staphylococcus aureus, Escherichia coli, and Candida albicans. A theoretical framework is established to extend bond-selective interferometric scattering microscopy to a broad range of biological micro- and nano-particles.In the second methodology, we demonstrate a back-ground suppressed high-throughput mid-infrared photothermal microscopy via pupil engineering. MIP microscopy has been a promising label-free chemical imaging technique for functional characterization of specimens owing to its enhanced spatial resolution and high specificity. Recently developed wide-field MIP imaging modalities have drastically improved speed and enabled high-throughput imaging of micron-scale subjects. However, the weakly scattered signal from subwavelength particles becomes indistinguishable from the shot-noise as a consequence of the strong background light, leading to limited sensitivity. Here, we demonstrate background-suppressed chemical fingerprinting at a single nanoparticle level by selectively attenuating the reflected light through pupil engineering in the collection path. Our technique provides over 3 orders of magnitude background suppression by quasi-darkfield illumination in the epi-configuration without sacrificing lateral resolution. We demonstrate 6-fold signal-to-background noise ratio improvement, allowing for simultaneous detection and discrimination of hundreds of nanoparticles across a field of view of 70 μm x 70 μm. A comprehensive theoretical framework for photothermal image formation is provided and experimentally validated with 300 and 500 nm PMMA beads. The versatility and utility of our technique are demonstrated via hyperspectral dark-field MIP imaging of S. aureus and E. coli bacteria and MIP imaging of subcellular lipid droplets inside C. albicans and cancer cells.In the third methodology, we present a bond-selective full-field optical coherence tomography. Optical coherence tomography (OCT) is a label-free, non-invasive 3D imaging tool widely used in both biological research and clinical diagnosis. Conventional OCT modalities can only visualize specimen tomography without chemical information. Here, we report a bond-selective full-field OCT (BS-FF-OCT), in which a pulsed mid-infrared laser is used to modulate the OCT signal through the photothermal effect, achieving label-free bond-selective 3D sectioned imaging of highly scattering samples. We first demonstrate BS-FF-OCT imaging of 1 µm PMMA beads embedded in agarose gel. Next, we show 3D hyperspectral imaging of up to 75 µm of polypropylene fiber mattress from a standard surgical mask. We then demonstrate BS-FF-OCT imaging on biological samples, including cancer cell spheroids and C. elegans. Using an alternative pulse timing configuration, we finally demonstrate the capability of BS-FF-OCT on imaging a highly scattering myelinated axons region in a mouse brain tissue slice.
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