Hyperspectral microscopy is an advanced visualization technique that combines hyperspectral imaging with state-of-the-art optics and computer software to enable the rapid identification of materials at the micro- and nanoscales. Achieving this level of resolution has traditionally required time-consuming and costly electron microscopy techniques. While hyperspectral microscopy has already been applied to the analysis of bulk materials and biologicals, it shows extraordinary promise as an analytical tool to locate individual nanoparticles and aggregates in complex samples through rapid optical and spectroscopic identification. This technique can be used to not only screen for the presence of nanomaterials, but also to locate, identify, and characterize them. It could also be used to identify a subset of samples that would then move on for further analysis via other advanced metrology. This review will describe the science and origins of hyperspectral microscopy, examine current and emerging applications in life science, and examine potential applications of this technology that could improve research efficiency or lead to novel discoveries.
Nanomaterials are increasingly prevalent throughout industry, manufacturing, and biomedical research. The need for tools and techniques that aid in the identification, localization, and characterization of nanoscale materials in biological samples is on the rise. Currently available methods, such as electron microscopy, tend to be resource-intensive, making their use prohibitive for much of the research community. Enhanced darkfield microscopy complemented with a hyperspectral imaging system may provide a solution to this bottleneck by enabling rapid and less expensive characterization of nanoparticles in histological samples. This method allows for high-contrast nanoscale imaging as well as nanomaterial identification. For this technique, histological tissue samples are prepared as they would be for light-based microscopy. First, positive control samples are analyzed to generate the reference spectra that will enable the detection of a material of interest in the sample. Negative controls without the material of interest are also analyzed in order to improve specificity (reduce false positives). Samples can then be imaged and analyzed using methods and software for hyperspectral microscopy or matched against these reference spectra in order to provide maps of the location of materials of interest in a sample. The technique is particularly well-suited for materials with highly unique reflectance spectra, such as noble metals, but is also applicable to other materials, such as semi-metallic oxides. This technique provides information that is difficult to acquire from histological samples without the use of electron microscopy techniques, which may provide higher sensitivity and resolution, but are vastly more resource-intensive and time-consuming than light microscopy.
Objective: To compare natural vs. hormone replacement treatment (HRT) for cryo-thaw embryo transfer cycles in patients with frozen embryos from previous ART. Design and Settings: Retrospective chart review of 164 patients (242 cycles) who underwent natural or HRT cryo-thaw embryo transfer between January 2002 and April 2005. Main Outcome Measures: Pregnancy rates per transfer in natural and HRT cryo-thaw cycles. Results: The pregnancy rate per transfer was higher with natural cycles (36.76% vs. 22.99%, P = 0.0298). There was no statistical difference in mean age, endometrial thickness, and average embryo quality in successful cycles. Mean endometrial thickness prior to transfer was greater in natural cycles (9.95 vs. 8.89 mm, P < 0.001). Mean serum estradiol levels were higher in the HRT group prior to transfer (526.1 vs. 103.8 pg/ml, P < 0.001), and were found to be lower in women who achieved pregnancy (337.1 vs. 433.3 pg/ml, P = 0.0136). Conclusion: Hormone replacement in preparation for cryo-thaw transfer of embryos was found to be associated with decreased pregnancy rates in comparison to natural cycle cryo-thaw transfer. Greater endometrial thickness was achieved with lower serum estradiol levels in patients undergoing natural cycles, suggesting Financial disclosure: none of the above authors had any financial interests with commercial companies.
While engineered nanomaterials (ENMs) are increasingly incorporated into industrial processes and consumer products, the potential biological effects and health outcomes of exposure remain unknown. Novel advanced direct visualization techniques that require less time, cost, and resource investment than electron microscopy (EM) are needed for identifying and locating ENMs in biological samples. Hyperspectral imaging (HSI) combines spectrophotometry and imaging, using advanced optics and algorithms to capture a spectrum from 400 to 1000 nm at each pixel in an enhanced dark-field microscopic (EDFM) image. HSI-EDFM can be used to confirm the identity of the materials of interest in a sample and generate an image "mapping" their presence and location in a sample. Hyperspectral mapping is particularly important for biological samples, where ENM morphology is visually indistinct from surrounding tissue structures. While use of HSI (without mapping) is increasing, no studies to date have compared results from hyperspectral mapping with conventional methods. Thus, the objective of this study was to utilize EDFM-HSI to locate, identify, and map metal oxide ENMs in ex vivo histological porcine skin tissues, a toxicological model of cutaneous exposure, and compare findings with those of Raman spectroscopy (RS), energy-dispersive X-ray spectroscopy (EDS), and scanning electron microscopy (SEM). Results demonstrate that EDFM-HSI mapping is capable of locating and identifying ENMs in tissue, as confirmed by conventional methods. This study serves as initial confirmation of EDFM-HSI mapping as a novel and higher throughput technique for ENM identification in biological samples, and serves as the basis for further protocol development utilizing EDFM-HSI for semiquantitation of ENMs.
A concerted effort is being made to insert Prevention through Design principles into discussions of sustainability, occupational safety and health, and green chemistry related to nanotechnology. Prevention through Design is a set of principles that includes solutions to design out potential hazards in nanomanufacturing including the design of nanomaterials, and strategies to eliminate exposures and minimize risks that may be related to the manufacturing processes and equipment at various stages of the lifecycle of an engineered nanomaterial.
Hyperspectral imaging (HSI) and mapping are increasingly used for visualization and identification of nanoparticles (NPs) in a variety of matrices, including aqueous suspensions and biological samples. Reference spectral libraries (RSLs) contain hyperspectral data collected from materials of known composition and are used to detect the known materials in experimental samples through a one-to-one pixel "mapping" process. In some HSI studies, RSLs created from raw NPs were used to map NPs in experimental samples in a different matrix; for example, RSLs created from NPs in suspension to map NPs in biological tissue. Others have utilized RSLs created from NPs in the same matrix. However, few studies have systematically compared hyperspectral data as a function of the matrix in which the NPs are found and its impact on mapping results. The objective of this study is to compare RSLs created from metal oxide NPs in aqueous suspensions to RSLs created from the same NPs in rat tissues following in vivo inhalation exposure, and to investigate the differences in mapping that result from the use of each RSL. Results demonstrate that the spectral profiles of these NPs are matrix dependent: RSLs created from NPs in positive control tissues mapped to experimental tissues more appropriately than RSLs created from NPs in suspension. Aqueous suspension RSLs mapped 0-602 out of 500,424 pixels per tissue image while tissue RSLs mapped 689-18,435 pixels for the same images. This study underscores the need for appropriate positive controls for the creation of RSLs for mapping NPs in experimental samples.
Abstract:As the scope of nanotechnology applications in medicine evolves, it is important to simultaneously recognize and advance contributions germane to public health. A wide range of innovations in nanomedicine stand to impact nearly every medical specialty and unveil novel ways to improve the quality and extend the duration of life -these gains can be measured at both individual and population levels. For example, heart disease and cancer combined make up approximately half of all deaths in the United States per year, and already, advances in nanomedicine demonstrate great potential to reduce rates of morbidity and mortality due to these diseases. Meanwhile, public health applications of nanomedicine such as rapid and portable diagnostics and more effective vaccinations have the potential to revolutionize global health. Research driven by innovators across disciplines such as engineering, biology, medicine, and public health should collaborate in order to achieve maximal potential impact in health for individuals and populations. In turn, knowledge gaps regarding the potential health and safety implications of exposure to engineered nanomaterials must be continuously addressed and actively researched. Dynamic, proactive, and socially responsible research will drive nanomedicine as it plays an increasingly integral and transformative role in medicine and public health in the 21st century.
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