Fluorescence Imaging Research Articles

The Orthogonal α‐meso‐Linked bisBODIPY Photosensitizers Demonstrate Effective Two‐Photon Fluorescence Imaging and Photodynamic Therapy

AbstractHypoxic tumor microenvironments pose significant challenges to the clinical translation of cancer photodynamic therapy (PDT). While heavy atom‐free Type‐I boron dipyrromethenes (BODIPYs) photosensitizers can alleviate this challenge by reducing oxygen dependency, but they remain scarce. Herein, heavy‐atom‐free α,meso‐linked bisBODIPYs were designed and synthesized. These bisBODIPYs exhibit remarkable red‐shifted emission (λemmax ~670 nm), large Stokes shifts (~4480 cm−1) and they are capable of producing both superoxide anion (O2⋅−) and singlet oxygen (1O2) in solution and cells. Additionally, they offer a wide PDT treatment window, ranging from 0.26 to 83.5 μM. Furthermore, these bisBODIPYs also demonstrate superior two‐photon fluorescence, promising for surgical navigation and integrating diagnosis with treatment.

Alkyl anchor–modified artificial viral capsid budding outside-to-inside and inside-to-outside giant vesicles

ABSTRACT The budding of human immunodeficiency virus from an infected host cell is induced by the modification of structural proteins bearing long-chain fatty acids, followed by their anchoring to the cell membrane. Although many model budding systems using giant unilamellar vesicles (GUVs) induced by various stimuli have been developed, constructing an artificial viral budding system of GUVs using only synthesized molecules remains challenging. Herein, we report the construction of an artificial viral capsid budding system from a lipid bilayer of GUV. The C-terminus of the β-annulus peptide was modified using an octyl chain as an alkyl anchor via a disulfide bond. The self-assembly of the β-annulus peptide with an octyl chain formed an artificial viral capsid aggregate. The fluorescence imaging and transmission electron microscopy observations revealed that the addition of the tetramethylrhodamine (TMR)-labeled octyl chain–bearing β-annulus peptide to the outer aqueous phase of GUV induced the budding of the capsid-encapsulated daughter vesicle outside-to-inside the mother GUV. Conversely, the encapsulation of the TMR-labeled octyl chain–bearing β-annulus peptide in the inner aqueous phase of GUV induced the budding of the capsid-encapsulated daughter vesicle inside-to-outside the mother GUV. Contrarily, the addition of the TMR-labeled β-annulus peptide to GUV barely induced budding. It was demonstrated that the higher the membrane fluidity of GUV, the more likely budding would be induced by the addition of the alkyl anchor–modified artificial viral capsid. The simple virus-mimicking material developed in this study, which buds off through membrane anchoring, can provide physicochemical insights into the mechanisms of natural viral budding from cells.

Electrophoretic Microplate Protein Identification Based on Gold Staining of Molybdenum Disulfide Hydrogels.

Numerous high-performance nanotechnologies have been developed, but their practical applications are largely restricted by the nanomaterials' low stabilities and high operation complexity in aqueous substrates. Herein, we develop a simple and high-reliability hydrogel-based nanotechnology based on the in situ formation of Au nanoparticles in molybdenum disulfide (MoS2)-doped agarose (MoS2/AG) hydrogels for electrophoresis-integrated microplate protein recognition. After the incubation of MoS2/AG hydrogels in HAuCl4 solutions, MoS2 nanosheets spontaneously reduce Au ions, and the hydrogels are remarkably stained with the color of as-synthetic plasmonic Au hybrid nanomaterials (Au staining). Proteins can precisely mediate the morphologies and optical properties of Au/MoS2 heterostructures in the hydrogels. Consequently, Au staining-based protein recognition is exhibited, and hydrogels ensure the comparable stabilities and sensitivities of protein analysis. In comparison to the fluorescence imaging and dye staining, enhanced sensitivity and recognition performances of proteins are implemented by Au staining. In Au staining, exfoliated MoS2 semiconductors directly guide the oriented growth of plasmonic Au nanostructures in the presence of formaldehyde, showing environment-friendly features. The Au-stained hydrogels merge the synthesis and recognition applications of plasmonic Au nanomaterials. Significantly, the one-step incubation of the electrophoretic hydrogels leads to high simplicity of operation, largely challenging those multiple-step Ag staining routes which were performed with high complexity and formaldehyde toxicity. Due to its toxic-free, simple, and sensitive merits, the Au staining integrated with electrophoresis-based separation and microplate-based high-throughput measurements exhibits highly promising and improved practicality of those developing nanotechnologies and largely facilitates in-depth understanding of biological information.

Carrier-Free Self-Assembled Nanomedicines for Promoting Apoptosis and Inhibiting Proliferation in Hepatocellular Carcinoma.

In order to improve the effectiveness of tumor treatment and reduce the toxic side effects of drugs, we formed carrier-free multifunctional nanoparticles (BI NPs) by noncovalent interaction of berberine hydrochloride and IR780. BI NPs possessed the synergistic effects of promoting apoptosis, inhibiting proliferation and metastasis of tumors, and phototherapeutic treatment. Dispersive and passive targeting ability retention (EPR) effects of BI NPs on tumor sites in vivo could be monitored by fluorescence imaging. In addition, BI NPs exhibited effective reactive oxygen species (ROS) generation and photothermal conversion capabilities, photodynamic therapy (PDT), and photothermal therapy (PTT). Importantly, BI NPs inhibit tumor suppression through the AMPK/PI3K/AKT signaling pathway to inhibit tumor proliferation and metastasis. BI NPs not only have efficient in vivo multimodal therapeutic effects but also have good biosafety and potential clinical applications.

Fluorescent-based micellar incorporated hydrogel materials for selective determination of long-chain aldehydes.

A highly sensitive micelle-induced sensory has been developed for detection of long-chain aldehydes as potential biomarkers of respiratory cancers. The micelle-like sensor was fabricated through the partial self-assembly of CTAB and S2 surfactants, containing a fluorescent hydrazine-functionalized dye (Naph-NH2). In principle, long-chain aldehydes with amphiphilic character act as the induced-fit surfactants to form well-entrapped micellar particles, as well as react with Naph-NH2 to form hydrazone derivatives resulting in fluorescent enhancement. The limit of detection (LOD) of micellar Naph-NH2/CTAB/S2 platform was calculated to be 64.09-80.98µM for detection of long-chain aldehydes, which showed fluorescent imaging in lung cancer cells (A549). This micellar sensory probe demonstrated practical applicability for long-chain aldehyde sensing in human blood samples with an accepted percent recovery of ~ 94.02-102.4%. Beyond Naph-NH2/CTAB/S2 sensor, the milcellar hybrid sensor was successfully developed by incorporating a micelle-like platform with supramolecular gel regarding to carboxylate-based gelators (Gel1), which showed a tenfold improvement in sensitivity. Expectedly, the determination of long-chain aldehydes through these sensing platforms holds significant promise for point-of-care cancer diagnosis and therapy.

Development of a plant-based surgical training model for fluorescence-guided cancer surgery.

Fluorescence-guided surgery (FGS) can help surgeons to discriminate tumor tissue from adjacent normal tissues using fluorescent tracers. We developed a surgical training model, manufactured using sustainable vegetable organic material with indocyanine green (ICG)-containing "tumor." Surgeons evaluated the model with both the closed-field and endoscopic fluorescence imaging devices and assessed its efficacy to identify residual tumor after enucleation using electrocautery. Strong correlations of fluorescence were obtained at all working distance (3, 5, 7, and 10 cm), showing the robustness of fluorescence signal for the closed-field and endoscopic fluorescence imaging devices. The higher fluorescence signals were obtained in the wound bed in the closed-field fluorescence imaging device and the residual tumor could be clearly identified by fluorescence endoscopy. Our FGS training model may provide experience for surgeons unfamiliar with optical surgery and subsequent tissue interactions. The model seemed particularly helpful in teaching surgeons the principles of FGS.

Ultrahigh-Speed 3D DNA Walker with Dual Self-Protected DNAzymes for Ultrasensitive Fluorescence Detection and Intracellular Imaging of microRNA.

Herein, a dual self-protected DNAzyme-based 3D DNA walker (dSPD walker), composed of activated dual self-protected walking particles (ac-dSPWPs) and track particles (TPs), was constructed for ultrasensitive and ultrahigh-speed fluorescence detection and imaging of microRNAs (miRNAs) in living cells. Impressively, compared with the defect that "one" target miRNA only initiates "one" walking arm of the conventional single self-protected DNAzyme walker, the dSPD walker benefits from the secondary amplification and spatial confinement effect and could guide "one" target miRNA to generate "n" secondary targets, thereby initiating "n" nearby walking strands immediately, realizing the initial rate over one-magnitude-order faster than that of the conventional one. Moreover, in the process of relative motion between ac-dSPWPs and TPs, the ac-dSPWPs could cleave multiple substrate strands simultaneously to speed up movement and reduce the derailment rate, as well as combine with successive TPs to facilitate a large amount of continuous signal accumulation, achieving an ultrafast detection of miRNA-221 within 10 min in vitro and high sensitivity with a low detection limit of 0.84 pM. In addition, the DNA nanospheres obtained by the rolling circle amplification reaction can capture the Cy5 fluorescence dispersed in liquids, which achieves the high-contrast imaging of miRNA-221, resulting in further ultrasensitive imaging of miRNA-221 in cancer cells. The proposed strategy has made a bold innovation in the rapid and sensitive detection as well as intracellular imaging of low-abundance biomarkers, offering promising application in early diagnosis and relevant research of cancer and tumors.

Use of Ambient Light Compatible Fluorescence-Guided Surgical Technology for Objective Assessment of Flap Perfusion in Autologous Breast Reconstruction.

Decreased autologous flap vascular perfusion can lead to secondary procedures. Fluorescence angiography during surgery reduces the probability of repeat surgery but suffers from interpretation variability. Recently, the OnLume Avata System was developed, which evaluates real-time vascular perfusion in ambient light. This study aims to predict complications in autologous breast reconstruction using measures of relative intensity (RI) and relative area (RA). Patients undergoing autologous breast reconstruction underwent intraoperative tissue perfusion assessment using the OnLume Avata System. Post-hoc image annotation was completed by labeling areas of the flap interpreted to be "Well Perfused," "Questionably Perfused," and "Under Perfused." RIs and RAs were calculated for the marked areas. Primary complications of interest were overall complication rate, fat and mastectomy skin flap necrosis, and surgical revision. Logistic regression was applied to determine the odds of developing a complication based on RI and RA for each image. A total of 25 patients (45 flaps) were included. In total, 17 patients (68%) developed at least one complication. Patients who developed any complication (p = 0.02) or underwent a surgical revision for complications (p = 0.02) had statistically lower RI of under-perfused portions of the flap. Patients with greater areas of under-perfused flap had a significantly higher risk of developing fat necrosis (odds ratio [OR]: 5.71, p = 0.03) and required a revision operation (OR: 1.10, p = 0.01). Image-based interpretation using the OnLume Avata System correlated with the risk of developing postoperative complications that standard fluorescence imaging systems may not appreciate. This information can benefit surgeons to improve perfusion assessment and intraoperative decision-making.

A GSH-Responsive Prodrug with Simultaneous Triple-Activation Capacity for Photodynamic/Sonodynamic Combination Therapy with Inhibited Skin Phototoxicity.

Herein, a dual-sensitizer prodrug, named pro-THPC, has been designed to function as both a photosensitizer and a sonosensitizer prodrug for precise antitumor combination therapy with minimized skin phototoxicity. Pro-THPC could be activated by glutathione (GSH) to release the dual-sensitizer, THPC, which simultaneously switches on fluorescence emission and combined capabilities of photodynamic therapy (PDT) and sonodynamic therapy (SDT). Pro-THPC is further formulated into nanoparticles (NPs) for water dispersity to enable in vivo applications. In vivo fluorescence imaging shows that the pro-THPC NPs group exhibits a significantly higher tumor-to-normal tissue ratio (T/N) (T/N = 5.2 ± 0.55) compared to the "always on" THPC NPs group (T/N = 2.9 ± 0.47) and the pro-THPC NPs group co-administrated with GSH synthesis inhibitor (buthionine sulfoximine, BSO) (T/N = 3.2 ± 0.63). In addition, the generation of the designed dual-sensitizer's reactive oxygen species (ROS) is effectively confined within the tumor tissues due to the relatively strong correlation between ROS generation and fluorescence emission. In vivo studies further demonstrate the remarkable efficacy of the designed pro-THPC NPs to eradicate tumors through the combination of PDT and SDT while significantly reducing skin phototoxicity.

Structural changes in the crystalline lens as a function of the postmortem interval assessed with two-photon imaging microscopy.

The properties and structure of the crystalline lens change as time after death passes. Some experiments have suggested that these might be used to estimate the postmortem interval (PMI). In this study, the organization and texture of the rabbit lens were objectively evaluated as a function of the PMI using two-photon excitation fluorescence (TPEF) imaging microscopy. Between 24 h and 72 h, the lens presented a highly organized structure, although the fiber delineation was progressively vanishing. At 96 h, this turned into a homogeneous pattern where fibers were hardly observed. This behaviour was similar for parameters providing information on tissue texture. On the other hand, the fiber density of the lens is linearly reduced with the PMI. On average, density at 24 h was approximately two-fold when compared to 96 h after death. The present results show that TPEF microscopy combined with different quantitative tools can be used to objectively monitor temporal changes in the lens fiber organization after death. This might help to estimate the PMI, which is one of the most complex problems in forensic science.

Assessing Pediatric Burn Wound Infection Using a Point-of-Care Fluorescence Imaging Device.

Wound infection is the most common complication among pediatric burn patients. When not treated promptly, burn wound infection may lead to delayed healing, failure of skin grafts, or death. Standard burn wound assessment includes inspection for visual signs and symptoms of infection (VSSI) and microbial sampling. To aid in the assessment of burn wound infection, the MolecuLight, a point-of-care autofluorescence imaging device, was introduced at our pediatric burn program in 2020. The MolecuLight uses violet light to illuminate the wound bed, causing clinically relevant quantities of 29 different species of bacteria (>104 CFU/g) to fluoresce in real time. The objectives of this study were to evaluate the role of the MolecuLight in the management of pediatric burn wounds and determine if the findings from the MolecuLight corresponded to VSSI and/or microbial sampling. A retrospective review of patients 0-18 years who had burn wounds assessed with the MolecuLight between November 1, 2020 and June 8, 2023 was conducted. Data were extracted from the medical records of 178 eligible patients with 218 wounds imaged with the device. Fluorescence corresponded with VSSI in 81% of wounds and microbial findings in 82% of wounds. MolecuLight fluorescence, in combination with VSSI, improved sensitivity for detecting wound infections by 39% and decreased specificity by 19% compared to visual signs and symptoms in isolation. Incorporation of the MolecuLight in standard burn wound assessments can improve the detection of infections, which may promote improved wound healing outcomes and antimicrobial stewardship.

A plasma membrane-localized transporter remobilizes aleurone layer magnesium for seed germination in rice.

The aleurone layer in cereal grains acts as a major reservoir of essential mineral nutrients, significantly influencing seed germination. However, the molecular mechanism underlying the redistribution of nutrients from the aleurone layer in the germinating seed is still not well understood. Here, in rice, we identified a plasma membrane (PM) localized magnesium transporter, MAGNESIUM RELEASE TRANSPORTER 3 (MGR3), is critical for seed germination. OsMGR3 is predominantly expressed in the aleurone layer cells of endosperm, facilitating magnesium remobilization during germination. Non-invasive Micro-test Technology assay data demonstrated that the loss-of-function of OsMGR3 restrained magnesium efflux from the aleurone layer. In the embryo/endosperm grafting experiment, we observed that the mutation of OsMGR3 in the aleurone layer suppressed the growth and differentiation of the embryo during germination. Furthermore, magnesium fluorescence imaging revealed the osmgr3 mutant seeds showed impaired exportation of aleurone layer-stored magnesium to the embryo, consequently delaying germination. Importantly, we discovered that disrupting OsMGR3 could inhibit pre-harvest sprouting without affecting rice yield and quality. Therefore, the magnesium efflux transporter OsMGR3 in the aleurone layer represents a promising genetic target for future agronomic trait improvement.

EnderScope: a low-cost 3D printer-based scanning microscope for microplastic detection.

Low-cost and scalable technologies that allow people to measure microplastics in their local environment could facilitate a greater understanding of the global problem of marine microplastic pollution. A typical way to measure marine microplastic pollution involves imaging filtered seawater samples stained with a fluorescent dye to aid in the detection of microplastics. Although traditional fluorescence microscopy allows these particles to be manually counted and detected, this is a resource- and labour-intensive task. Here, we describe a novel, low-cost microscope for automated scanning and detection of microplastics in filtered seawater samples-the EnderScope. This microscope is based on the mechanics of a low-cost 3D printer (Creality Ender 3). The hotend of the printer is replaced with an optics module, allowing for the reliable and calibrated motion system of the 3D printer to be used for automated scanning over a large area (>20 × 20 cm). The EnderScope is capable of both reflected light and fluorescence imaging. In both configurations, we aimed to make the design as simple and cost-effective as possible, for example, by using low-cost LEDs for illumination and lighting gels as emission filters. We believe this tool is a cost-effective solution for microplastic measurement. This article is part of the Theo Murphy meeting issue 'Open, reproducible hardware for microscopy'.

Two-site microendoscopic imaging probe for simultaneous three-dimensional imaging at two anatomic locations in tissues.

Systems that can image in three dimensions at cellular resolution and across different locations within an organism may enable insights into complex biological processes, such as immune responses, for which a single location measurement may be insufficient. In this Letter, we describe an in vivo two-site imaging probe (TIP) that can simultaneously image two anatomic sites with a maximum separation of a few centimeters. The TIP consists of two identical bendable graded index (GRIN) lenses and is demonstrated by a two-photon two-color fluorescence imaging system. Each GRIN lens has a field of view of 162 × 162 × 170 µm3, a nominal numerical aperture of 0.5, a magnification of 0.7, and working distances of 0.2 mm in air for both ends. A blind linear unmixing algorithm is applied to suppress bleedthrough between channels. We use this system to successfully demonstrate two-site two-photon two-color imaging of two biomedically relevant samples, i.e., (1) a mixture of two autofluorescent anti-cancer drugs and (2) a live hybrid tumor consisting of two spectrally distinct fluorescent cell lines.

A Novel Multi-Effect Photosensitizer for Tumor Destruction via Multimodal Imaging Guided Synergistic Cancer Phototherapy.

How to ingeniously design multi-effect photosensitizers (PSs), including multimodal imaging and multi-channel therapy, is of great significance for highly spatiotemporal controllable precise phototherapy of malignant tumors. Herein, a novel multifunctional zinc(II) phthalocyanine-based planar micromolecule amphiphile (ZnPc 1) was successfully designed and synthesized, in which N atom with photoinduced electron transfer effect was introduced to enhance the near-infrared absorbance and nonradiative heat generation. After simple self-assembling into nanoparticles (NPs), ZnPc 1 NPs would exhibit enhanced multimodal imaging properties including fluorescence (FL) imaging (FLI) /photoacoustic (PA) imaging (PAI) /infrared (IR) thermal imaging, which was further used to guide the combined photodynamic therapy (PDT) and photothermal therapy (PTT). It was that under the self-guidance of the multimodal imaging, ZnPc 1 NPs could precisely pinpoint the tumor from the vertical and horizontal boundaries achieving highly efficient and accurate treatment of cancer. Accordingly, the integration of FL/PA/IR multimodal imaging and PDT/PTT synergistic therapy pathway into one ZnPc 1 could provide a blueprint for the next generation of phototherapy, which offered a new paradigm for the integration of diagnosis and treatment in tumor and a promising prospect for precise cancer therapy.

An activatable azophenyl fluorescent probe for hypoxic fluorescence imaging in living cells.

Cellular hypoxia is a common pathological process in various diseases. Detecting cellular hypoxia is of great scientific significance for early diagnosis of tumors. The hypoxia fluorescence probe analysis method can efficiently and conveniently evaluate the hypoxia status in tumor cells. These probes are covalently linked by hypoxic recognition groups and organic fluorescent molecules. Currently, the fluorescent molecules used in these probes often exhibit the aggregation-caused quenching effect, which is not conducive to fluorescence imaging in water. Herein, an activatable hypoxia fluorescence probe was constructed by covalently linking aggregation-induced emission luminogens to the hypoxic recognition group azobenzene. It does not emit fluorescence in solution and in solid state under light excitation due to the presence of photosensitive azo bonds. It can be cleaved by intracellular azoreductase into fluorescent amino derivatives with aggregation-induced emission characteristic. As the concentration of oxygen in cells decreases, its fluorescence intensity increases, making it suitable for fluorescence imaging to detect hypoxic environment in live cancer cells. This work broadens the molecular design approach for activatable hypoxia fluorescent probes.

IR-780 Dye-based Targeting of Cancer-associated Fibroblasts Improves Cancer Immunotherapy by Increasing Intra-tumoral T Lymphocytes Infiltration.

Immune-checkpoint inhibitors (ICIs) against programmed death (PD)-1/PD-L1 pathway immunotherapy have been demonstrated to be effective in only a subset of patients with cancer, while the rest may exhibit low response or may develop drug resistance after initially responding. Previous studies have indicated that extensive collagen-rich stroma secreted by cancer-associated fibroblasts (CAFs) within the tumor microenvironment is one of the key obstructions of the immunotherapy for some tumors by decreasing the infiltrating cytotoxic T cells. However, there is still a lack of effective therapeutic strategies to control the extracellular matrix by targeting CAFs. The enhanced uptake of IR-780 by CAFs was assessed by using in vivo or ex vivo nearinfrared fluorescence imaging, confocal NIR fluorescent imaging, and CAFs isolation testing. The fibrotic phenotype down-regulation effects and in vitro CAFs killing effect of IR-780 were tested by qPCR, western blot, and flow cytometry. The in vivo therapeutic enhancement of anti-PD-L1 by IR-780 was evaluated on EMT6 and MC38 subcutaneous xenograft mice models. IR-780 has been demonstrated to be preferentially taken up by CAFs and accumulate in the mitochondria. Further results identified low-dose IR-780 to downregulate the fibrotic phenotype, while high-dose IR-780 could directly kill both CAFs and EMT6 cells in vitro. Moreover, IR-780 significantly inhibited extracellular matrix (ECM) protein deposition in the peri-tumoral stroma on subcutaneous EMT6 and MC38 xenografts, which increased the proportion of tumor-infiltrating lymphocytes (TILs) in the deep tumor and further promoted anti-PD-L1 therapeutic efficacy. This work provides a unique strategy for the inhibition of ECM protein deposition in the tumor microenvironment by targeted regulating of CAFs, which destroys the T cell barrier and further promotes tumor response to PD-L1 monoclonal antibody. IR-780 has been proposed as a potential therapeutic small-molecule adjuvant to promote the effect of immunotherapy.

A handheld device for intra-cavity and ex vivo fluorescence imaging of breast conserving surgery margins with 5-aminolevulinic acid

BackgroundVisualization of cancer during breast conserving surgery (BCS) remains challenging; the BCS reoperation rate is reported to be 20-70% of patients. An urgent clinical need exists for real-time intraoperative visualization of breast carcinomas during BCS. We previously demonstrated the ability of a prototype imaging device to identify breast carcinoma in excised surgical specimens following 5-aminolevulinic acid (5-ALA) administration. However, this prototype device was not designed to image the surgical cavity for remaining carcinoma after the excised lumpectomy specimen is removed. A new handheld fluorescence (FL) imaging prototype device, designed to image both excised specimens and within the surgical cavity, was assessed in a clinical trial to evaluate its clinical utility for first-in-human, real-time intraoperative imaging during index BCS.ResultsThe imaging device combines consumer-grade imaging sensory technology with miniature light-emitting diodes (LEDs) and multiband optical filtering to capture high-resolution white light (WL) and FL digital images and videos. The technology allows for visualization of protoporphyrin IX (PpIX), which fluoresces red when excited by violet-blue light. To date, n=17\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$n = 17$$\\end{document} patients have received 20mgkg\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$20\\frac{\ ext {mg}}{\ extrm{kg}}$$\\end{document} bodyweight (BW) 5-ALA orally 2-4 h before imaging to facilitate the accumulation of PpIX within tumour cells. Tissue types were identified based on their colour appearance. Breast tumours in sectioned lumpectomies appeared red, which contrasted against the green connective tissues and orange-brown adipose tissues. In addition, ductal carcinoma in situ (DCIS) that was missed during intraoperative standard of care was identified at the surgical margin at <1 mm depth. In addition, artifacts due to the surgical drape, illumination, and blood within the surgical cavity were discovered.ConclusionsThis study has demonstrated the detection of a grossly occult positive margin intraoperatively. Artifacts from imaging within the surgical cavity have been identified, and potential mitigations have been proposed.Trial RegistrationClinicalTrials.gov Identifier: NCT01837225 (Trial start date is September 2010. It was registered to ClinicalTrials.gov retrospectively on April 23, 2013, then later updated on April 9, 2020, to reflect the introduction of the new imaging device.)

Si-rhodamines with fine-tuned reactivities for tracking formaldehyde in glioblastoma

Formaldehyde (FA), an essential signaling molecule and an emerging neurotoxin in the brain, regulates various physiological and pathological processes. The abnormal metabolism of FA is associated with extensive brain diseases, including glioblastoma. However, FA probes with systematically fine-tuned reactivities are still very challenging. A group of silicon-substituted rhodamine-based FA probes was designed with fine-tuned structures and FA activities for tracking FA in vivo. With systematic structural manipulation, we have quantitatively revealed the chemical origins of the distinct FA reactivities in probes. Moreover, we successfully tracked the dynamic of FA in live cells, organs, and live animals by the designed probe. The vivid upregulation of FA in heterotopic xenografted and orthotopic glioblastoma by fluorescence imaging with the designed FA probe has proved that FA can be a potential molecular marker for glioblastoma.

A Novel Dual-labeled Peptide for Multimodal Imaging of EGFR with L858R Mutation.

The development of molecular imaging agents targeting epidermal growth factor receptor (EGFR) with L858R mutation may help with the selection of non-small cell lung carcinoma (NSCLCL) patients who may benefit from EFGR tyrosine kinase inhibitor (TKI) therapy. In this study, we developed 99mTc STHHYYP-GHEG-ECGK-tetramethylrhodamine (STHHYYP-ECGK-TAMRA) to target EGFR with L858R mutation in NSCLC tumors and verified its probability as a molecular imaging agent. Fmoc solid-phase peptide synthesis was used to synthesize STHHYYP-ECGKTAMRA. 99mTc labelled STHHYYP-ECGK-TAMRA was prepared. Gamma imaging, fluorescent imaging and biodistribution were performed in murine models bearing NCI-H1975 and NCI-H1650 tumors. The binding affinity value (Kd) of 99mTc STHHYYP-ECGK-TAMRA was estimated to be 130.6 ± 29.2 nM in NCI-H1975 cells. The gamma camera images showed a substantial uptake of 99mTc STHHYYP-ECGK-TAMRA in the NCI-H1975 tumor. The % injected dose/gram of the NCI-H1975 tumor tissue was 2.77 ± 0.70 and 3.48 ± 1.01 at 1 and 3 h, respectively. Specific binding of 99mTc STHHYYP-ECGK-TAMRA to L858R-mutated EGFRpositive NCI-H1975 cells and tumors was demonstrated in in vivo and in vitro studies. The results suggest that 99mTc STHHYYP-ECGK-TAMRA is a good candidate agent for dualmodality imaging targeting EGFR with L858R mutation.