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Typical (hyper)spectral imaging approaches. (A) Point scan. (B) Line scan (i.e. "pushbroom"). (C) Wavelength scan. (D) Snapshot.
Source publication
In recent decades, various classes of nanoparticles have been developed for optical imaging of cancers. Many of these nanoparticles are designed to specifically target tumor sites, and specific cancer biomarkers, to facilitate the visualization of tumors. However, one challenge for accurate detection of tumors is that the molecular profiles of most...
Contexts in source publication
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... two major spectral imaging categories are scanning-based imaging and wide-field imaging ( Figure 3). The former is usually implemented by scanning a collimated or focused laser beam (in the form of a spot or a line) across the specimen ( Figure 4A and 4B). For example, raster scanning may be achieved by steering the laser beam with a galvanometric scanning mirror (high speed) or translating the specimen with a mechanical stage (low speed) [86]. ...
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... we define "wide-field imaging" as a method that illuminates a region and detects the emitted light from an array of points (pixels) within the area using a camera (a 2D detector array). Compared with scanning-based imaging methods, wide-field imaging eliminates the need for mechanical scanning and can image a large area with one acquisition ( Figure 4C and 4D), which is typically simpler and faster. Wide-field imaging methods are often utilized to achieve high spatial resolution, including super-resolution with structured illumination [93]. ...
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... and newly developed scientific CMOS (sCMOS) sensor arrays are the most widely used detector arrays for wide-field imaging, allowing for the acquisition of millions of spatial pixels with one exposure in time. Rapid hyperspectral wide-field imaging can be achieved by either acquiring two-dimensional (2D) images at multiple wavelength channels over time ( Figure 4C) [65] or taking a snapshot to acquire both spatial and spectral information (sometimes referred to "snapshot hyperspectral imaging") ( Figure 4D) [99][100][101]. The former approach can achieve a high spectral resolution (channel bandwidth) of 1-5 nm by using monochromators, filter wheels, or electronic tunable filters such as liquid crystal tunable filters (LCTFs) and acousto-optic tunable filters (AOTFs) [100]. ...
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... and newly developed scientific CMOS (sCMOS) sensor arrays are the most widely used detector arrays for wide-field imaging, allowing for the acquisition of millions of spatial pixels with one exposure in time. Rapid hyperspectral wide-field imaging can be achieved by either acquiring two-dimensional (2D) images at multiple wavelength channels over time ( Figure 4C) [65] or taking a snapshot to acquire both spatial and spectral information (sometimes referred to "snapshot hyperspectral imaging") ( Figure 4D) [99][100][101]. The former approach can achieve a high spectral resolution (channel bandwidth) of 1-5 nm by using monochromators, filter wheels, or electronic tunable filters such as liquid crystal tunable filters (LCTFs) and acousto-optic tunable filters (AOTFs) [100]. ...
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... main disadvantage of filter-based spectral imaging is low light throughput and prolonged imaging times due to the fact that only one filter channel is collected at a time and the rest of the photons are discarded. In comparison, the latter "snapshot" approach typically utilizes a combination of image-division elements and dispersive elements (typically a prism or diffractive optical element) to image a 2D field onto sub-regions of a detector array such that all spatial pixels and spectral channels are imaged simultaneously ( Figure 4D shows the image mapping spectrometry approach [104,105]; detailed discussion of all snapshot imaging approaches can be found in [101,106]). Snapshot hyperspectral imaging can enable enhanced light throughput and imaging speed, but may require a tradeoff between spatial and spectral resolution since a limited number of camera pixels must be used to collect signals from both different spectral channels as well as different spatial positions [100,107,108]. ...
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Citations
... 20 These examples highlight the use of SERS for the detection of clinically relevant targets because SERS provides a number of key advantages, including ultra-sensitivity, high multiplexing capability, quantification based on spectral intensities, high photostabil-ity, and the requirement for only a single laser source for excitation. [21][22][23][24] Therefore, one of the main strengths of SERS is its ability to detect several analytes in a single sample with complex mixture. Recently, multiplexed real-time PCR-based SERS detection was reported for the simultaneous detection of two biomarker genes from Staphylococcus aureus using a thermoplastic chip and labelled probes, 25 and compared with this strategy, our proposed PCR/SERS assay offers several advantages, including (i) using small molecules rather than fluorophores as Raman reporters for strong and well-defined Raman peaks, enabling improved multiplexing capabilities, (ii) a simpler scheme using a magnetic-bead/PCR products/SERS nanotags sandwich structure for the SERS signal readout, and (iii) a much shorter assay time (80 min). ...
Bacterial infections are a leading cause of death globally. The detection of DNA sequences correlated to the causative pathogen has become a vital tool in medical diagnostics. In practice, PCR-based assays for the simultaneous detection of multiple pathogens currently rely on probe-based quantitative strategies that require expensive equipment but have limited sensitivity or multiplexing capabilities. Hence, novel approaches to address the limitations of the current gold standard methods are still in high demand. In this study, we propose a simple multiplex PCR/SERS assay for the simultaneous detection of four bacterial pathogens, namely P. aeruginosa, S. aureus, S. epidermidis, and M. smegmatis. Wherein, specific primers for amplifying each target gDNA were applied, followed by applying SERS nanotags functionalized with complementary DNA probes and Raman reporters for specific identification of the target bacterial pathogens. The PCR/SERS assay showed high specificity and sensitivity for genotyping bacterial pathogen gDNA, whereby as few as 100 copies of the target gDNA could be detected. With high sensitivity and the convenience of standard PCR amplification, the proposed assay shows great potential for the sensitive detection of multiple pathogen infections to aid clinical decision-making.
... Hyperspectral image acquisition devices are designed in four different imaging approaches, where images are acquired one point at a time (point-scan), one line at a time, comprising the entire spectral range (line-scan), one spatial image at a time at each wavelength (wavelength scan), and one spatial image at a time at each wavelength (snapshot) (Wang et al., 2017). ...
Traditional techniques for quality assessment of fresh agricultural products are onerous and costly. Imaging systems based on the visible wavelength range have been widely used for automated processing because they are relatively inexpensive and fast and designed to represent the human eye to detect external features such as size, shape, colour, or the presence of damages and diseases. However, traditional cameras cannot identify damage that is internal or non-visible to the human eye. Hyperspectral imaging combines conventional two-dimensional imaging systems with spectroscopy to simultaneously obtain spatial and spectral information from an object. This technology is being progressively applied for postharvest inspection of fruits and vegetables to develop predictive models for estimating internal quality or identifying key wavelengths related to specific problems. Nevertheless, to obtain accurate and trustable detection, the use of this technology requires adequate equipment, robust calibrations, and powerful statistical tools to create prediction or classification models that are consistent over time. This chapter reviews the application, advantages, and requirements of visible and near-infrared hyperspectral imaging for mechanical/bruise damage and quality assessment of fresh horticultural produce.
... Importantly, NIR MFLI Forster Resonance Energy Transfer (FRET) measurements can quantify the fraction of drug binding to the target in live and intact preclinical animal models [13][14][15]. Thus, optical imaging has emerged as an invaluable tool to quantify the binding of antibody drugs to their target in a noninvasive and longitudinal manner [16,17]. We have demonstrated the utility of NIR MFLI FRET to measure receptor-ligand binding via dark quencher (QC-1)-mediated FRET in the transferrin (Tf) -transferrin receptor system [18]. ...
Rationale
Trastuzumab (TZM) is a monoclonal antibody that targets the human epidermal growth factor receptor (HER2) and is clinically used for the treatment of HER2-positive breast tumors. However, the tumor microenvironment can limit the access of TZM to the HER2 targets across the whole tumor and thereby compromise TZM’s therapeutic efficacy. An imaging methodology that can non-invasively quantify the binding of TZM-HER2, which is required for therapeutic action, and distribution within tumors with varying tumor microenvironments is much needed.
Methods
We performed near-infrared (NIR) fluorescence lifetime (FLI) Forster Resonance Energy Transfer (FRET) to measure TZM-HER2 binding, using in vitro microscopy and in vivo widefield macroscopy, in HER2 overexpressing breast and ovarian cancer cells and tumor xenografts, respectively. Immunohistochemistry was used to validate in vivo imaging results.
Results
NIR FLI FRET in vitro microscopy data show variations in intracellular distribution of bound TZM in HER2-positive breast AU565 and AU565 tumor-passaged XTM cell lines in comparison to SKOV-3 ovarian cancer cells. Macroscopy FLI (MFLI) FRET in vivo imaging data show that SKOV-3 tumors display reduced TZM binding compared to AU565 and XTM tumors, as validated by ex vivo immunohistochemistry. Moreover, AU565/XTM and SKOV-3 tumor xenografts display different amounts and distributions of TME components, such as collagen and vascularity. Therefore, these results suggest that SKOV-3 tumors are refractory to TZM delivery due to their disrupted vasculature and increased collagen content.
Conclusion
Our study demonstrates that FLI is a powerful analytical tool to monitor the delivery of antibody drug tumor both in cell cultures and in vivo live systems. Especially, MFLI FRET is a unique imaging modality that can directly quantify target engagement with potential to elucidate the role of the TME in drug delivery efficacy in intact live tumor xenografts.
... Several pathways exist to increase the speed of the acquisition of hyperspectral images [18,19]. A common approach is to directly image the sample through different (variable) spectral filters. ...
Optical micro-spectroscopy is an invaluable tool for studying and characterizing samples ranging from classical semiconductors to low-dimensional materials and heterostructures. To date, most implementations are based on point-scanning techniques, which are flexible and reliable, but slow. Here, we describe a setup for highly parallel acquisition of hyperspectral reflection and photoluminescence microscope images using a push-broom technique. Spatial as well as spectral distortions are characterized and their digital corrections are presented. We demonstrate close-to diffraction-limited spatial imaging performance and a spectral resolution limited by the spectrograph. The capabilities of the setup are demonstrated by recording a hyperspectral photoluminescence map of a CVD-grown MoSe 2 -WSe 2 lateral heterostructure, from which we extract the luminescence energies, intensities and peak widths across the interface.
... Fluorescence can be used to label ligands, such as peptides, and provide high contrast to distinguish regions of pathology from surrounding areas of normal tissues. The optical spectrum provides a wide range of wavelengths (500-900 nm) that can be separated into different channels used to perform multiplexed detection of unique molecular targets [10][11][12] . A number of fluorophores, such as fluorescein and indocyanine green (ICG), are FDA-approved for clinical administration, and can be used to label ligands that bind specifically to biomarkers of disease 13 . ...
A wide-field endoscope that is sensitive to fluorescence can be used as an adjunct to conventional white light endoscopy by detecting multiple molecular targets concurrently. We aim to demonstrate a flexible fiber-coupled accessory that can pass forward through the instrument channel of standard medical endoscopes for clinical use to collect fluorescence images. A miniature scan mirror with reflector dimensions of 1.30 × 0.45 mm² was designed, fabricated, and placed distal to collimated excitation beams at λex = 488, 660, and 785 nm. The mirror was driven at resonance for wide angular deflections in the X and Y-axes. A large image field-of-view (FOV) was generated in real time. The optomechanical components were packaged in a rigid distal tip with dimensions of 2.6 mm diameter and 12 mm length. The scan mirror was driven at 27.6 and 9.04 kHz in the fast (X) and slow (Y) axes, respectively, using a square wave with 50% duty cycle at 60 Vpp to collect fluorescence images at 10 frames per sec. Maximum total divergence angles of ± 27.4° and ± 22.8° were generated to achieve a FOV of 10.4 and 8.4 mm, respectively, at a working distance of 10 mm. Multiplexed fluorescence images were collected in vivo from the rectum of live mice using 3 fluorescently-labeled peptides that bind to unique cell surface targets. The fluorescence images collected were separated into 3 channels. Target-to-background ratios of 2.6, 3.1, and 3.9 were measured. This instrument demonstrates potential for broad clinical use to detect heterogeneous diseases in hollow organs.
... Different Types of Scanners[17]. ...
Recording spectral information together with spatial information is useful for many applications. Hyperspectral Imaging (HSI) is a recently developed imaging modality that is capable of recording both the spatial (x, y) and spectral (λ) information of a scene. Thereafter, several applications have been shown using HSI. In this review, we discussed some of the recent implementations of HSI that aim to address several scientific research problems. We also gave a brief overview of some of the existing types of HSI cameras. In some recent works, Deep Learning (DL) frameworks have been extensively used for several HSI-related problems. We have provided insights on some of the recent implementations of DL for HSI-based problems.
... This limitation is stipulated by the bandgap energy of approximately ≈ 1.1 eV (1100 nm) beyond which there will be no photogenerated carriers. Other materials such as mercury cadmium telluride (MCT) are sensitive to much lower energy in the SWIR band and beyond [25], making them suitable for Earth observation application in vegetation and carbon dioxide emission monitoring. However, such sensors are still far from consumers' access due to their prohibitive price tag. ...
With advancements in computer processing power and deep learning techniques, hyperspectral imaging is continually being explored for improved sensing applications in various fields. However, the high cost associated with such imaging platforms impedes their widespread use in spite of the availability of the needed processing power. In this paper, we develop a novel theoretical framework required for an open source ultra-low-cost hyperspectral imaging platform based on the line scan method suitable for remote sensing applications. Then, we demonstrate the design and fabrication of an open source platform using consumer-grade commercial off-the-shelf components that are both affordable and easily accessible to researchers and users. At the heart of the optical system is a consumer-grade spectroscope along with a basic galvanometer mirror that is widely used in laser scanning devices. The utilized pushbroom scanning method provides a very high spectral resolution of 2.8 nm, as tested against commercial spectral sensors. Since the resolution is limited by the slit width of the spectroscope, we also provide a deconvolution method for the line scan in order to improve the monochromatic spatial resolution. Finally, we provide a cost-effective testing method for the hyperspectral imaging platform where the results validate both the spectral and spatial performances of the platform.
... Various HSI methods have been developed, primarily based on scanning, to acquire a hyperspectral cube [15]. The typical method is the push-broom HSI, where a dispersed image of a slit is acquired column-by-column [16,17]. ...
Hyperspectral imaging (HSI) has become a valuable tool in sample characterization in various scientific fields. While many approaches have been tested, specific applications and technology usually lead to only a narrow part of the spectrum being studied. We demonstrate the use of a broadband HSI setup based on compressed sensing capable of capturing data in visible (VIS), near-infrared (NIR), and short-wave infrared (SWIR) spectral regions. Using a tested design, we developed a dual configuration and tested its performance on a set of samples demonstrating spatial resolution and spectral reconstruction. Samples showing a potential use of the setup in optical defect detection are also tested. The setup showcases a dual single-pixel camera configuration capable of combining various detectors with a shared spatial modulation, further improving data efficiency and providing an affordable instrument from broadband spectral studies.
... The combination of optical spectroscopy and imaging is rapidly developing in many fields [1][2][3][4]. Microscopes are combined with spectrometers to create images where both spatial and spectral information are available for richer contrast. Typically, a confocal microscope is combined with a spectrometer that utilizes a diffraction grating or an etalon interferometer; the confocal microscope scans the object being imaged pixel by pixel and the spectrometer analyzes the spectral information at each pixel [4,5]. ...
Spectral imaging techniques extract spectral information using dispersive elements in combination with optical microscopes. For rapid acquisition, multiplexing spectral information along one dimension of imaged pixels has been demonstrated in hyperspectral imaging, as well as in Raman and Brillouin imaging. Full-field spectroscopy, i.e., multiplexing where imaged pixels are collected in 2D simultaneously while spectral analysis is performed sequentially, can increase spectral imaging speed, but so far has been attained at low spectral resolutions. Here, we extend 2D multiplexing to high spectral resolutions of ∼80 MHz (∼0.0001 nm) using high-throughput spectral discrimination based on atomic transitions.
... number of dyes that can be excited by a single laser beam so that each dye is distinguishable from the others is eight 33 . This upper limit is not determined by spectral resolution (i.e., number of colors) but by the widths of the fluorescence spectra (≥ 50 nm) and the magnitudes of the Stokes shifts (≤ 200 nm) of the dyes when fluorescence resonance energy transfer (FRET) 10 is used. However, the number of colors must be greater than or equal to the number of dyes in order to unmix the spectral overlaps of the dyes 31,32 . ...
An ultra-small (54 × 58 × 8.5 mm) and large aperture (1 × 7 mm) nine-color spectrometer—using an array of ten dichroic mirrors “biparted” as two layers—was developed and used for snapshot spectral imaging. Incident-light flux with a cross section smaller than the aperture size is split into nine color fluxes with 20-nm-width contiguous wavelength bands and central wavelengths of 530, 550, 570, 590, 610, 630, 650, 670, and 690 nm. Images of the nine color fluxes are simultaneously and efficiently measured by an image sensor. Unlike a conventional dichroic-mirror array, the developed dichroic-mirror array has a unique biparted configuration that not only increases the number of colors that can be measured simultaneously but also improves the image resolution of each color flux. The developed nine-color spectrometer was used for four-capillary-array electrophoresis. Eight dyes concurrently migrating in each capillary were simultaneously quantified by nine-color laser-induced fluorescence detection. Since the nine-color spectrometer is not only ultra-small and inexpensive but also has high light throughput and sufficient spectral resolution for most spectral-imaging applications, it has the potential to be widely used in various fields.
