Fast volumetric imaging with line-scan confocal microscopy by an electro-tunable lens

AbstractIn microscopic imaging of biological tissues, particularly real-time visualization of neuronal activities, rapid acquisition of volumetric images poses a prominent challenge. Typically, two-dimensional (2D) microscopy can be devised into an imaging system with 3D capability using any varifocal lens. Despite the conceptual simplicity, such an upgrade yet requires additional, complicated device components and suffers a reduced acquisition rate, which is critical to document neuronal dynamics properly. In this study, we implemented an electro-tunable lens (ETL) in the line-scan confocal microscopy, enabling the volumetric acquisition at the rate of 20 frames per second with the maximum volume of interest of 315 × 315 × 80 μm3. The axial extent of point-spread-function (PSF) was 17.6 ± 1.6 μm and 90.4 ± 2.1 μm with the ETL operating in either stationary or resonant mode, respectively, revealing significant depth elongation by the resonant mode ETL microscopy. We further demonstrated the utilities of the ETL system by volume imaging of cleared mouse brain ex vivo samples and in vivo brains. The current study foregrounds the successful application of resonant ETL for constructing a basis for a high-performance 3D line-scan confocal microscopy system, which will enhance our understanding of various dynamic biological processes.

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Performance of a novel protease-activated fluorescent imaging system for intraoperative detection of residual breast cancer during breast conserving surgery

Breast Cancer Research and Treatment ◽

10.1007/s10549-021-06106-w ◽

2021 ◽

Vol 187 (1) ◽

pp. 145-153

Author(s):

Conor R. Lanahan ◽

Bridget N. Kelly ◽

Michele A. Gadd ◽

Michelle C. Specht ◽

Carson L. Brown ◽

...

Keyword(s):

Breast Cancer ◽

Real Time ◽

Imaging System ◽

Ex Vivo ◽

Menopausal Status ◽

Fluorescent Imaging ◽

Cancer Subtypes ◽

Surgical Workflow ◽

Tumor Types

Abstract Purpose Safe breast cancer lumpectomies require microscopically clear margins. Real-time margin assessment options are limited, and 20–40% of lumpectomies have positive margins requiring re-excision. The LUM Imaging System previously showed excellent sensitivity and specificity for tumor detection during lumpectomy surgery. We explored its impact on surgical workflow and performance across patient and tumor types. Methods We performed IRB-approved, prospective, non-randomized studies in breast cancer lumpectomy procedures. The LUM Imaging System uses LUM015, a protease-activated fluorescent imaging agent that identifies residual tumor in the surgical cavity walls. Fluorescent cavity images were collected in real-time and analyzed using system software. Results Cavity and specimen images were obtained in 55 patients injected with LUM015 at 0.5 or 1.0 mg/kg and in 5 patients who did not receive LUM015. All tumor types were distinguished from normal tissue, with mean tumor:normal (T:N) signal ratios of 3.81–5.69. T:N ratios were 4.45 in non-dense and 4.00 in dense breasts (p = 0.59) and 3.52 in premenopausal and 4.59 in postmenopausal women (p = 0.19). Histopathology and tumor receptor testing were not affected by LUM015. Falsely positive readings were more likely when tumor was present < 2 mm from the adjacent specimen margin. LUM015 signal was stable in vivo at least 6.5 h post injection, and ex vivo at least 4 h post excision. Conclusions Intraoperative use of the LUM Imaging System detected all breast cancer subtypes with robust performance independent of menopausal status and breast density. There was no significant impact on histopathology or receptor evaluation.

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Applications of Vibrational Spectroscopy for Analysis of Connective Tissues

Molecules ◽

10.3390/molecules26040922 ◽

2021 ◽

Vol 26 (4) ◽

pp. 922

Author(s):

William Querido ◽

Shital Kandel ◽

Nancy Pleshko

Keyword(s):

Raman Spectroscopy ◽

Vibrational Spectroscopy ◽

Near Infrared ◽

Ex Vivo ◽

Biological Tissues ◽

Label Free ◽

Connective Tissues ◽

Bone Properties ◽

Composition And Structure

Advances in vibrational spectroscopy have propelled new insights into the molecular composition and structure of biological tissues. In this review, we discuss common modalities and techniques of vibrational spectroscopy, and present key examples to illustrate how they have been applied to enrich the assessment of connective tissues. In particular, we focus on applications of Fourier transform infrared (FTIR), near infrared (NIR) and Raman spectroscopy to assess cartilage and bone properties. We present strengths and limitations of each approach and discuss how the combination of spectrometers with microscopes (hyperspectral imaging) and fiber optic probes have greatly advanced their biomedical applications. We show how these modalities may be used to evaluate virtually any type of sample (ex vivo, in situ or in vivo) and how “spectral fingerprints” can be interpreted to quantify outcomes related to tissue composition and quality. We highlight the unparalleled advantage of vibrational spectroscopy as a label-free and often nondestructive approach to assess properties of the extracellular matrix (ECM) associated with normal, developing, aging, pathological and treated tissues. We believe this review will assist readers not only in better understanding applications of FTIR, NIR and Raman spectroscopy, but also in implementing these approaches for their own research projects.

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A Tumbling Magnetic Microrobot System for Biomedical Applications

Micromachines ◽

10.3390/mi11090861 ◽

2020 ◽

Vol 11 (9) ◽

pp. 861

Author(s):

Elizabeth E. Niedert ◽

Chenghao Bi ◽

Georges Adam ◽

Elly Lambert ◽

Luis Solorio ◽

...

Keyword(s):

Ultrasound Imaging ◽

Biomedical Applications ◽

Imaging System ◽

Ex Vivo ◽

Negative Control ◽

Circular Path ◽

Rotational Axis ◽

Significant Difference

A microrobot system comprising an untethered tumbling magnetic microrobot, a two-degree-of-freedom rotating permanent magnet, and an ultrasound imaging system has been developed for in vitro and in vivo biomedical applications. The microrobot tumbles end-over-end in a net forward motion due to applied magnetic torque from the rotating magnet. By turning the rotational axis of the magnet, two-dimensional directional control is possible and the microrobot was steered along various trajectories, including a circular path and P-shaped path. The microrobot is capable of moving over the unstructured terrain within a murine colon in in vitro, in situ, and in vivo conditions, as well as a porcine colon in ex vivo conditions. High-frequency ultrasound imaging allows for real-time determination of the microrobot’s position while it is optically occluded by animal tissue. When coated with a fluorescein payload, the microrobot was shown to release the majority of the payload over a 1-h time period in phosphate-buffered saline. Cytotoxicity tests demonstrated that the microrobot’s constituent materials, SU-8 and polydimethylsiloxane (PDMS), did not show a statistically significant difference in toxicity to murine fibroblasts from the negative control, even when the materials were doped with magnetic neodymium microparticles. The microrobot system’s capabilities make it promising for targeted drug delivery and other in vivo biomedical applications.

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Mesenchymal stem cells accelerated growth and metastasis of neuroblastoma and preferentially homed towards both primary and metastatic loci in orthotopic neuroblastoma model

BMC Cancer ◽

10.1186/s12885-021-08090-2 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Jiao-Le Yu ◽

Shing Chan ◽

Marcus Kwong-Lam Fung ◽

Godfrey Chi-Fung Chan

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Primary Tumor ◽

Imaging System ◽

Ex Vivo ◽

Tumor Formation ◽

Human Neuroblastoma Cell ◽

Fluorescence Signal ◽

Human Neuroblastoma

Abstract Background Majority of neuroblastoma patients develop metastatic disease at diagnosis and their prognosis is poor with current therapeutic approach. Major challenges are how to tackle the mechanisms responsible for tumorigenesis and metastasis. Human mesenchymal stem cells (hMSCs) may be actively involved in the constitution of cancer microenvironment. Methods An orthotopic neuroblastoma murine model was utilized to mimic the clinical scenario. Human neuroblastoma cell line SK-N-LP was transfected with luciferase gene, which were inoculated with/without hMSCs into the adrenal area of SCID-beige mice. The growth and metastasis of neuroblastoma was observed by using Xenogen IVIS 100 in vivo imaging and evaluating gross tumors ex vivo. The homing of hMSCs towards tumor was analyzed by tracing fluorescence signal tagged on hMSCs using CRI Maestro™ imaging system. Results hMSCs mixed with neuroblastoma cells significantly accelerated tumor growth and apparently enhanced metastasis of neuroblastoma in vivo. hMSCs could be recruited by primary tumor and also become part of the tumor microenvironment in the metastatic lesion. The metastatic potential was consistently reduced in lung and tumor when hMSCs were pre-treated with stromal cell derived factor-1 (SDF-1) blocker, AMD3100, suggesting that the SDF-1/CXCR4 axis was one of the prime movers in the metastatic process. Conclusions hMSCs accelerated and facilitated tumor formation, growth and metastasis. Furthermore, the homing propensity of hMSCs towards both primary tumor and metastatic loci can also provide new therapeutic insights in utilizing bio-engineered hMSCs as vehicles for targeted anti-cancer therapy.

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In Vivo Confocal Microscopy Evaluation in Patients with Keratoconus

Journal of Clinical Medicine ◽

10.3390/jcm11020393 ◽

2022 ◽

Vol 11 (2) ◽

pp. 393

Author(s):

Alvin Wei Jun Teo ◽

Hassan Mansoor ◽

Nigel Sim ◽

Molly Tzu-Yu Lin ◽

Yu-Chi Liu

Keyword(s):

Confocal Microscopy ◽

Nerve Regeneration ◽

Nerve Fibre ◽

Ex Vivo ◽

Fibre Length ◽

In Vivo Confocal Microscopy ◽

Corneal Sensitivity ◽

Mechanical Removal ◽

Cellular Changes

Keratoconus is the most common primary corneal ectasia characterized by progressive focal thinning. Patients experience increased irregular astigmatism, decreased visual acuity and corneal sensitivity. Corneal collagen crosslinking (CXL), a minimally invasive procedure, is effective in halting disease progression. Historically, keratoconus research was confined to ex vivo settings. In vivo confocal microscopy (IVCM) has been used to examine the corneal microstructure clinically. In this review, we discuss keratoconus cellular changes evaluated by IVCM before and after CXL. Cellular changes before CXL include decreased keratocyte and nerve densities, disorganized subbasal nerves with thickening, increased nerve tortuosity and shortened nerve fibre length. Repopulation of keratocytes occurs up to 1 year post procedure. IVCM also correlates corneal nerve status to functional corneal sensitivity. Immediately after CXL, there is reduced nerve density and keratocyte absence due to mechanical removal of the epithelium and CXL effect. Nerve regeneration begins after 1 month, with nerve fibre densities recovering to pre-operative levels between 6 months to 1 year and remains stable up to 5 years. Nerves remain tortuous and nerve densities are reduced. Corneal sensitivity is reduced immediately postoperatively but recovers with nerve regeneration. Our article provides comprehensive review on the use of IVCM imaging in keratoconus patients.

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Rationally Designing Simple Rod-Like Amphiphilic NIR-Emissive AIE Probes for Precise In-Vivo Detection of Aβ Fibrils/Plaques at A Super-Early Stage

10.26434/chemrxiv.13699222.v1 ◽

2021 ◽

Author(s):

Yipu Wang ◽

Dong Mei ◽

Xinyi Zhang ◽

Da-Hui Qu ◽

Ju Mei ◽

...

Keyword(s):

High Performance ◽

Ex Vivo ◽

Early Stage ◽

Signal To Noise Ratio ◽

High Signal ◽

In Vivo Detection ◽

Different Strains ◽

Aβ Fibrils

With increase of social aging, Alzheimer's disease (AD) has been one of the serious diseases threatening human health. The occurrence of Aβ fibrils or plaques is recognized as the hallmark of AD. Currently, optical imaging has stood out to be a promising technique for the imaging of Aβ fibrils/plaques and the diagnosis of AD. However, restricted by their poor blood-brain barrier (BBB) penetrability, short-wavelength excitation and emission, and aggregation-caused quenching (ACQ) effect, the clinically used gold-standard optical probes such as <a>thioflavin</a> T (ThT) and thioflavin S (ThS), are not effective enough in the early diagnosis of AD in vivo. Herein, we put forward an “all-in-one” design principle and demonstrate its feasibility in developing high-performance fluorescent probes which are specific to Aβ fibrils/plaques and promising for super-early in-vivo diagnosis of AD. As a proof of concept, a simple rod-like amphiphilic NIR fluorescent AIEgen, i.e., AIE-CNPy-AD, is developed by taking the specificity, BBB penetration ability, deep-tissue penetration capacity, high signal-to-noise ratio (SNR) into consideration. AIE-CNPy-AD is constituted by connecting the electron-donating and accepting moieties through single bonds and tagging with a propanesulfonate tail, giving rise to the NIR fluorescence, aggregation-induced emission (AIE) effect, amphiphilicity, and rod-like structure, which in turn result in high binding-affinity and excellent specificity to Aβ fibrils/plaques, satisfactory ability to penetrate BBB and deep tissues, ultrahigh SNR and sensitivity, and high-fidelity imaging capability. In-vitro, ex-vivo, and in-vivo <a>identifying of Aβ fibrils/plaques</a> in different strains of mice indicate that AIE-CNPy-AD holds the universality to the detection of Aβ fibrils/plaques. It is noteworthy that AIE-CNPy-AD is even able to trace the small and sparsely distributed Aβ fibrils/plaques in very young AD model mice such as 4-month-old APP/PS1 mice which are reported to be the youngest mice to have Aβ deposits in brains, suggesting its great potential in diagnosis and intervention of AD at a super-early stage.

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Proof of concept of a multimodal intravital molecular imaging system for tumour transpathology investigation

European Journal of Nuclear Medicine and Molecular Imaging ◽

10.1007/s00259-021-05574-y ◽

2021 ◽

Author(s):

Zhen Liu ◽

Tao Cheng ◽

Stephan Düwel ◽

Ziying Jian ◽

Geoffrey J. Topping ◽

...

Keyword(s):

Molecular Imaging ◽

Imaging System ◽

Ex Vivo ◽

Fiducial Markers ◽

Proof Of Concept ◽

Registration Accuracy ◽

Microscopic Imaging ◽

Window Chamber

Abstract Background Transpathology highlights the interpretation of the underlying physiology behind molecular imaging. However, it remains challenging due to the discrepancies between in vivo and in vitro measurements and difficulties of precise co-registration between trans-scaled images. This study aims to develop a multimodal intravital molecular imaging (MIMI) system as a tool for in vivo tumour transpathology investigation. Methods The proposed MIMI system integrates high-resolution positron imaging, magnetic resonance imaging (MRI) and microscopic imaging on a dorsal skin window chamber on an athymic nude rat. The window chamber frame was designed to be compatible with multimodal imaging and its fiducial markers were customized for precise physical alignment among modalities. The co-registration accuracy was evaluated based on phantoms with thin catheters. For proof of concept, tumour models of the human colorectal adenocarcinoma cell line HT-29 were imaged. The tissue within the window chamber was sectioned, fixed and haematoxylin–eosin (HE) stained for comparison with multimodal in vivo imaging. Results The final MIMI system had a maximum field of view (FOV) of 18 mm × 18 mm. Using the fiducial markers and the tubing phantom, the co-registration errors are 0.18 ± 0.27 mm between MRI and positron imaging, 0.19 ± 0.22 mm between positron imaging and microscopic imaging and 0.15 ± 0.27 mm between MRI and microscopic imaging. A pilot test demonstrated that the MIMI system provides an integrative visualization of the tumour anatomy, vasculatures and metabolism of the in vivo tumour microenvironment, which was consistent with ex vivo pathology. Conclusions The established multimodal intravital imaging system provided a co-registered in vivo platform for trans-scale and transparent investigation of the underlying pathology behind imaging, which has the potential to enhance the translation of molecular imaging.

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Radiolabeling of methanol extracts of yarrow (Achillea millefolium l) in rats

Acta Cirurgica Brasileira ◽

10.1590/s0102-86502012000500003 ◽

2012 ◽

Vol 27 (5) ◽

pp. 294-300 ◽

Cited By ~ 9

Author(s):

Betul Cekic ◽

Ayfer Yurt Kilcar ◽

Fazilet Zumrut Biber Muftuler ◽

Perihan Unak ◽

Emin Ilker Medine

Keyword(s):

High Performance ◽

Imaging System ◽

New Drugs ◽

High Yield ◽

Albino Rats ◽

Imaging Studies ◽

Biodistribution Studies ◽

In Vivo Imaging System ◽

Wistar Albino Rats

PURPOSE: Current study is focused on extraction with methanol, purification, labeling with 131I using iodogen method of the yarrow plant and investigating in vivo biological activity using biodistribution and imaging studies on healthy animal models. The aim of the study is to contribute plant extracts to discover new drugs in the diagnosis and treatment of several diseases. METHODS: Nine female and nine male healthy Wistar albino rats, which were approximately 100-150 g in weight, were used for biodistribution studies. For imaging studies four healthy male Balb-C mice were used. Quality control studies were done utilizing thin layer radio chromatography (TLRC) and high performance liquid chromatography (HPLC) methods. For biodistribution studies, 131I radiolabeled Peak 7 (131I-Peak 7) was sterilized and injected into the tail veil of rats and imaging studies were obtained using Kodak FX PRO in vivo Imaging System. RESULTS: The radiolabeling yield of each purified the bioactive extracts of the yarrow plant, seven peaks was between 79 and 92%. The highest radiolabeling yield was calculated for 131I radiolabeled seventh peak (131I-Peak 7) (92.78±5.04, n=5). For this reason the biodistribution and imaging studies were done for 131I-Peak 7. That's why; these studies with Peak 7 were carried out. CONCLUSION: Peak 7 was radiolabeled with 131I in high yield for using imaging and therapeutic studies in nuclear medical applications.

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Development and Optimization of a Fluorescent Imaging System to Detect Amyloid-β Proteins: Phantom Study

Biomedical Engineering and Computational Biology ◽

10.1177/1179597218781081 ◽

2018 ◽

Vol 9 ◽

pp. 117959721878108 ◽

Cited By ~ 2

Author(s):

David Tes ◽

Karl Kratkiewicz ◽

Ahmed Aber ◽

Luke Horton ◽

Mohsin Zafar ◽

...

Keyword(s):

Alzheimer Disease ◽

Imaging System ◽

Ex Vivo ◽

Amyloid Β ◽

The United States ◽

Fluorescent Imaging ◽

Fluorescent Properties ◽

In Vivo Experiments ◽

The Brain

Alzheimer disease is the most common form of dementia, affecting more than 5 million people in the United States. During the progression of Alzheimer disease, a particular protein begins to accumulate in the brain and also in extensions of the brain, ie, the retina. This protein, amyloid-β (Aβ), exhibits fluorescent properties. The purpose of this research article is to explore the implications of designing a fluorescent imaging system able to detect Aβ proteins in the retina. We designed and implemented a fluorescent imaging system with a range of applications that can be reconfigured on a fluorophore to fluorophore basis and tested its feasibility and capabilities using Cy5 and CRANAD-2 imaging probes. The results indicate a promising potential for the imaging system to be used to study the Aβ biomarker. A performance evaluation involving ex vivo and in vivo experiments is planned for future study.

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