All animal experiments were conducted in accordance with the German Animal Welfare Law and approved by local authorities. To study the tumor homing of the αVβ3 binder, male BALB/c nude mice (11 weeks; Taconic M&B A/S, Lille Skensved, Denmark) were inoculated subcutaneously (s.c.) with 2 × 106 786-O RCC cells suspended in phosphate-buffered saline (PBS). The mice were fed with chlorophyll-free chow (Harlan, Horst, The Netherlands) 7 days prior to imaging to reduce the background fluorescence. When the 786-O tumors reached an average tumor size of 0.7 cm in diameter, the mice (n = 3 mice/group) were injected with 20 nmol of BAY810 (1.35 mg/kg), consisting of the αVβ3 binder coupled to IRDye® 800CW (LI-COR Bioscience, Bad Homburg, Germany), according to the manufacturer’s instructions, or BAY813 (1.35 mg/kg), a non-binding control conjugate of IRDye® 800CW. As a negative control, one mouse was injected with IRDye® 800CW carboxylate (0.84 mg/kg) in a sterile 0.9% NaCl aqueous solution (Sigma-Aldrich). Tumor accumulation of the IRDye® conjugates in mice was determined 8, 24, and 48 h post administration using the LI-COR Pearl® imaging system (LI-COR Bioscience, Bad Homburg, Germany).
Pearl imaging system
Manufactured by LI COR
Sourced in United States
The Pearl Imaging System is a fluorescence-based imager designed for life science research applications. It is capable of capturing high-resolution images of fluorescently labeled samples. The system utilizes multiple excitation and emission wavelengths to enable the detection of a variety of fluorescent probes.
Automatically generated - may contain errors
Lab products found in correlation
Lsm800 confocal microscope, by Zeiss (4 mentions)
500 mhz nmr spectrometer, by Agilent Technologies (2 mentions)
Agilent 1100 hplc system, by Thermo Fisher Scientific (2 mentions)
Kta pure system, by GE Healthcare (2 mentions)
Hypersil ods c18 column, by Thermo Fisher Scientific (2 mentions)
Superdex 200 increase column 10 300 gl, by Wyatt Technology (2 mentions)
Odyssey clx, by LI COR (2 mentions)
Irdye 800cw, by LI COR (1 mentions)
Rm2255 rotary microtome, by Leica camera (1 mentions)
Tbi 0310, by Precision Systems and Instrumentation (1 mentions)
Irdye 800cw nhs ester, by LI COR (1 mentions)
Image studio software, by LI COR (1 mentions)
10 protocols using pearl imaging system
1
Tumor Targeting of α₅β₃ Integrin Binder
2
Multimodal Characterization of Polymer-based Formulations
3
Biodistribution of Fluorescent Probe in Xenograft Mice
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All animal studies were UAB Occupational Health & Safety approved (Project #14–124) and carried out in accordance with the policies and guidelines set by the Institutional Animal Care and Use Committee (IACUC Animal Project Number #10159). Briefly[49 (link)], 6-week-old female athymic nude mice (Charles River; Hartford, CT) were stereotactically injected with 5 × 105 X1016 cells into the caudate putamen 1 week before biodistribution studies. Animals were sorted by bioluminescence imaging to have similar tumor burden between groups and then the two groups were randomized to the two dose levels but the investigators were not blinded. MED2-CY7 diluted in 100 μL of normal saline was injected via tail vein, and mice sacrificed 3 hours later. Major organs were collected and weighed before imaging on Pearl Imaging System (LI-COR Biosciences) at 700nm channel using uniform Z-plane thickness similar to previous work.[52 (link)] The brain was formalin fixed overnight before sectioning, gross fluorescent imaging, and paraffin embedding and mounting. Consecutive sections were H&E stained or fluorescence intensity determined using an Odyssey CLx (LI-COR Biosciences) at 700nm channel. Fluorescent intensity per mg of tissue was calculated in excel and graphed using Prism. For comparisons, the color scale was kept uniform for each group of images.
4
In Vivo Optical Imaging of HA Hydrogel and MSCs
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Serial in vivo optical imaging was performed to assess the localization of the HA hydrogel compound and the MSCs. MSCs were tagged with QtrackerTM (Invitrogen, Eugene, OR, USA, Cat#: Q25061MP), which enabled in vivo imaging of live cells, and HA hydrogels were tagged by covalently bonding the near-infrared marker Cy7.5 to the Ad-HA component of the HA hydrogel as previously described [8 (link),9 (link)]. The animals were imaged utilizing the Pearl® imaging system (Li-Cor, Lincoln, NE, USA) prior to injection, immediately postinjection and on days 3–7, 14, 21, and 28. Signal intensity measured in photons/pixel/second was analyzed (Pearl Impulse Software v2.0), normalized per animal to peak intensity, and averaged per cohort.
5
Nanobody-Photosensitizer Tumor Imaging
6
Comprehensive Analytical Techniques for Polymer Characterization
7
Nanobody-Photosensitizer Tumor Imaging
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Tumor specificity of 7D12–PS and 7D12-9G8–PS was explored by optical imaging in vivo and fluorescence imaging of tissue sections. At 1 h after injection of 7D12–PS, 7D12-9G8–PS and R2–PS (200 μg), mice were imaged (n = 2) with the Pearl imaging system (LI-COR), while kept under isoflurane anesthesia.
In parallel, other mice injected with each of the nanobody–PS conjugates (n= 1)had their tongues resected, fixed overnight in 4% formalin and embedded in paraffin blocks. Tissue was sectioned at 10 μm and fluorescence imaging was performed using the Odyssey. All images were acquired using the same settings. Further, sections were processed for hematoxylin and eosin (H&E) staining.
In parallel, other mice injected with each of the nanobody–PS conjugates (n= 1)had their tongues resected, fixed overnight in 4% formalin and embedded in paraffin blocks. Tissue was sectioned at 10 μm and fluorescence imaging was performed using the Odyssey. All images were acquired using the same settings. Further, sections were processed for hematoxylin and eosin (H&E) staining.
8
Biodistribution of Fluorescent Probe in Xenograft Mice
9
Quantifying Brain PD-L1 Expression via Fluorescence Imaging
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After MRI scans, pigs were sacrificed and their brains were harvested and fixed in 10% formalin for at least 48 h prior to cutting into 2 mm slices, obtained using a 3D-printed brain slicer matrix. The brains were cut horizontally, preserving the FUS-sonicated regions and nonsonicated contralateral regions on the same level. The fixed brain slices were imaged using the LI-COR Pearl imaging system with the 800 nm acquisition channel for 800CW-aPD-L1 imaging. All images were acquired under the same imaging parameters, including the exposure time. The fluorescence intensity of brain slices was quantified using the LI-COR Image Lite Software (version 5.2).
10
Anti-FnEDA Antibody Targeting in mCIA
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To compare the targeting of anti-FnEDA mAbs in disease vs. normal tissues, mCIA animals were enrolled between days 24 and 28 at the first clinical signs of disease and were used for imaging at peak inflammation on day 7 post-enrollment.
Conjugation of anti-FnEDA antibody with IRDye 800CW-NHS ester (Li-COR) was performed according to methods described in a previous section. For organ distribution, 5 mg/kg of labeled anti-FnEDA and anti-TeTx were administered through an i.v. injection to mCIA and naïve mice, 3 mice per group. Injected mice were euthanized at 72 h post-administration, perfused with saline and the organs were excised to acquire fluorescence images across different tissues using a LiCOR Pearl imaging system. Mean fluorescence was determined from using ImageStudio software (Li-COR) by subtracting the background, drawing a region of interest around the organs, and using a measure function to determine the mean value.
Conjugation of anti-FnEDA antibody with IRDye 800CW-NHS ester (Li-COR) was performed according to methods described in a previous section. For organ distribution, 5 mg/kg of labeled anti-FnEDA and anti-TeTx were administered through an i.v. injection to mCIA and naïve mice, 3 mice per group. Injected mice were euthanized at 72 h post-administration, perfused with saline and the organs were excised to acquire fluorescence images across different tissues using a LiCOR Pearl imaging system. Mean fluorescence was determined from using ImageStudio software (Li-COR) by subtracting the background, drawing a region of interest around the organs, and using a measure function to determine the mean value.
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