A total of 1.0 × 106 cells was labeled with XenoLight DiR (Perkin Elmer, Waltham, MA, USA). The staining procedure was performed according to the manufacturer’s instructions. Briefly, cells were treated with DiR for 30 min at 37 °C, centrifuged for 3 min at 400 × g at room temperature, and washed twice with PBS. In all cases, DiR-labeled cells were suspended in saline and then intravenously injected through the tail vein within 2 h of labeling. Before taking the fluorescence images, the hair of C57BL/6 mice was removed to avoid any interference in detecting the fluorescence signal. Fluorescence images were obtained at 1, 2, 4, and 6 h after injection using the Xenogen IVIS Spectrum system (Perkin Elmer). All images were acquired with excitation at 748 nm and emission at 780 nm.
Xenolight dir
Manufactured by PerkinElmer
Sourced in United States
The XenoLight DiR is a near-infrared fluorescent dye used as a labeling agent for in vivo imaging. It emits fluorescence at a wavelength of approximately 750 nm, which allows for deep tissue penetration and reduced autofluorescence. The dye can be used to label a variety of biological samples, including cells and tissues, for visualization and tracking purposes in preclinical research applications.
Automatically generated - may contain errors
Lab products found in correlation
Ivis spectrum, by PerkinElmer (2 mentions)
Human cd14 microbeads, by Miltenyi Biotec (2 mentions)
Fetal bovine serum (fbs), by Benchmark Scientific (2 mentions)
Dynabeads human t activator cd3 cd28, by Thermo Fisher Scientific (2 mentions)
Human pan t cell isolation kit, by Miltenyi Biotec (2 mentions)
Cd16 microbeads, by Miltenyi Biotec (2 mentions)
Pen strep, by Thermo Fisher Scientific (2 mentions)
Rpmi 1640, by Thermo Fisher Scientific (2 mentions)
Ficoll paque, by GE Healthcare (2 mentions)
L glutamine, by Thermo Fisher Scientific (2 mentions)
Recombinant il 2 ril 2, by Merck Group (2 mentions)
Lsm 880, by Zeiss (2 mentions)
Xenogen ivis spectrum system, by PerkinElmer (1 mentions)
Ivis spectrum in vivo imaging system, by PerkinElmer (1 mentions)
Ivis lumina xr system, by PerkinElmer (1 mentions)
Mannitol, by Merck Group (1 mentions)
Soybean phosphatidylcholine spc, by Merck Group (1 mentions)
Paclitaxel tax, by Merck Group (1 mentions)
Tween 80, by Merck Group (1 mentions)
Mw 3000, by Thermo Fisher Scientific (1 mentions)
15 protocols using xenolight dir
1
In Vivo Cell Labeling and Imaging
2
Fluorescent Labeling of UC-MSCs for Biodistribution
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Cultured UC-MSC suspensions were fluorescently labelled for 10 min with 10 µM carboxyfluorescein succinimidyl ester (CFSE) or 320 μg/mL XenoLight DiR (PerkinElmer) in DMEM for 20 min for in vivo biodistribution analysis using IVIS® Spectrum in vivo imaging system (PerkinElmer). Fluorescently labelled UC-MSC cultures were extensively washed before harvesting with trypsin. Fluorescently labelled UC-MSC suspension were identically washed twice through centrifugation before transplantation.
3
Cardiomyocyte Labeling and Myocardial Injection
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For the staining, 1 × 107 cardiomyocytes were resuspended in 1 ml DPBS and incubated for 20 min with 300 µM XenoLight DiR fluorescent dye (PerkinElmer, Waltham, MA, USA) dissolved in DMSO. Afterwards, cells were centrifuged at 200 × g for 5 min, washed twice with 5 ml DPBS and resuspended in 1 ml prewarmed CMM medium.
Porcine hearts were rinsed with 0.9% NaCl solution, sealed in plastic bags and warmed up to 37 °C in a water bath to simulate physiological body temperature. Before injection, hearts were cut 5 cm horizontally above the apex and the upper part was removed. This allowed better positioning and observation of the hearts by an vivo imaging system (IVIS Spectrum, Perkin Elmer). Then, 1 × 106 XenoLight DiR fluorescent dye-labeled cardiomyocytes resuspended in 100 µl CMM or 100 µl CMM without cells were injected at a 90° angle 2 cm above the apex into the myocardium either using the new hydrojet system (E60/E10, E80/E10) or a 27 G needle.
Porcine hearts were rinsed with 0.9% NaCl solution, sealed in plastic bags and warmed up to 37 °C in a water bath to simulate physiological body temperature. Before injection, hearts were cut 5 cm horizontally above the apex and the upper part was removed. This allowed better positioning and observation of the hearts by an vivo imaging system (IVIS Spectrum, Perkin Elmer). Then, 1 × 106 XenoLight DiR fluorescent dye-labeled cardiomyocytes resuspended in 100 µl CMM or 100 µl CMM without cells were injected at a 90° angle 2 cm above the apex into the myocardium either using the new hydrojet system (E60/E10, E80/E10) or a 27 G needle.
4
Tracking Mesenchymal Stem Cells with DiR Dye
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To allow in vivo tracking of MSCs, BM-MSCs and MenSCs (both at passage 7) were stained with XenoLight DiR, a near-IR lipophilic membrane dye (PerkinElmer, Waltham, MA, USA). MSCs were incubated with 320 μg/ml of DiR for 30 min at 37°C according to the manufacturer's protocol.
The DiR-labeled MSCs were spinned down for 5 min at 1000 rpm, and cell pellets were resuspended in PBS. This procedure was repeated twice to ensure complete removal of any unbound dye.
The DiR-labeled MSCs were spinned down for 5 min at 1000 rpm, and cell pellets were resuspended in PBS. This procedure was repeated twice to ensure complete removal of any unbound dye.
5
Isolation and Activation of Human T Cells
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Peripheral blood was obtained from healthy donors through an IRB-approved program at the University of Maryland, Baltimore. Peripheral blood mononuclear cells (PBMCs) were harvested from peripheral blood by Ficoll–Paque (GE Healthcare) gradient centrifugation.
T cells from PBMCs were isolated using human Pan T Cell Isolation Kits (Miltenyi Biotec) according to the manufacturer’s protocol. T cells were further cultured with T-cell medium, which was composed of RPMI 1640 (Life Technologies), 2 mMl -glutamine (Life Technologies), 10% FBS (Benchmark), and 100 U/ml pen/strep (Life Technologies)] supplemented with recombinant IL-2 (rIL-2) (30 U/ml) (Millipore Sigma). Human T-Activator CD3/CD28 Dynabeads™ (Thermo Fisher Scientific) were used for T cell expansion and activation according to the manufacturer’s protocol. Activated T cells were incubated at 37 °C and 5% CO2 for 7 days before injection into mice.
Before injection into mice, T cells were labeled by incubation with 320 μg/mL Xenolight Dir (PerkinElmer) for 30 min and subsequently washed twice with chilled PBS.
CD14+ cells and CD16+ cells were isolated by using human CD14 microbeads (Miltenyi Biotec) and CD16 microbeads (Miltenyi Biotec), respectively, according to the manufacturer’s protocol.
T cells from PBMCs were isolated using human Pan T Cell Isolation Kits (Miltenyi Biotec) according to the manufacturer’s protocol. T cells were further cultured with T-cell medium, which was composed of RPMI 1640 (Life Technologies), 2 mM
Before injection into mice, T cells were labeled by incubation with 320 μg/mL Xenolight Dir (PerkinElmer) for 30 min and subsequently washed twice with chilled PBS.
CD14+ cells and CD16+ cells were isolated by using human CD14 microbeads (Miltenyi Biotec) and CD16 microbeads (Miltenyi Biotec), respectively, according to the manufacturer’s protocol.
6
In Vivo Tracking of Magnetized MSCs
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Magnetized MSCs were fluorescently labeled with XenoLight DiR (PerkinElmer, Inc., Waltham, Massachusetts), a lipophilic near‐infrared fluorescent dye (absorption/emission: 748/780 nm). DiR was initially dissolved in ethanol, and this solution then mixed with PBS. MSCs were incubated with 32 μg/mL DiR solution in a 37°C incubator for 30 minutes. After incubation, the cells were centrifuged and washed with PBS to remove free dye, in accordance with the manufacturer's instructions.
In vivo tracking of MSCs was then performed using an IVIS Lumina XR system (Caliper Life Sciences/PerkinElmer, Hopkinton, Massachusetts). The filters were configured at 710 nm for excitation and 760 nm for emission. Fluorescence images were acquired immediately, 24 hours, and 48 hours after cell transplantation. At 48 hours, mice were euthanized by cervical dislocation under anesthesia and ex vivo imaging of their lungs was performed. Fluorescent signal intensities (expressed as average radiance values relative to silicotic mice without MSCs) were measured and analyzed in Living Image 4.3.1 software. Two independent experiments were performed in triplicate.
In vivo tracking of MSCs was then performed using an IVIS Lumina XR system (Caliper Life Sciences/PerkinElmer, Hopkinton, Massachusetts). The filters were configured at 710 nm for excitation and 760 nm for emission. Fluorescence images were acquired immediately, 24 hours, and 48 hours after cell transplantation. At 48 hours, mice were euthanized by cervical dislocation under anesthesia and ex vivo imaging of their lungs was performed. Fluorescent signal intensities (expressed as average radiance values relative to silicotic mice without MSCs) were measured and analyzed in Living Image 4.3.1 software. Two independent experiments were performed in triplicate.
7
In Vivo Tracking of Labeled hucMSCs
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hucMSCs were labeled with the XenoLight DiR (1,1′-dioctadecyltetramethyl-indotricarbocyanine iodide; “DiR,” Perkin Elmer, Catalog#125964) for 25 min at 37 °C at a concentration of 15 μM in PBS. Following labeling, cells were washed twice in 0.9% saline solution to remove unbound dye before i.v. injection. Mice were imaged with a NightOWL in vivo imaging system.
8
Preclinical Evaluation of Modified NK Cells
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Nonobese diabetic/severe-combined immunodeficiency/IL-2Rγcnull (NSG) mice (The Jackson Laboratory, Bar Harbor, ME, USA) were maintained and used in the current study. Animal experiments were performed according to protocols reviewed and approved by the Institutional Animal Care and Use Committee (IACUC), the Biological Resource Centre (BRC), and the Agency for Science, Technology and Research (A*STAR), Singapore (IACUC protocol number 181324). Mouse xenograft models were generated by s.c. injection of 5 × 106 FaDu tumor cells in the left flank in male NSG mice (8–10 weeks) or i.p. injection of 1 × 107 SKOV3-Luc tumor cells in female NSG mice (8–10 weeks). For in vivo migration experiments, NK cells (5 × 106) were labeled with Xenolight DiR (Perkin Elmer, OH, USA) and injected via the tail vein (i.v.) into tumor-bearing mice. Biodistribution of NK cells was examined by DIR imaging at time points indicated and using the IVIS imaging system (Xenogen). The SKOV3-Luc xenograft model was used to evaluate the in vivo antitumor efficacy of CXCR1-modified NK cells or NKG2D CAR+ CXCR1+ NK cells.
9
LHRH-Targeted Lipid Nanoparticles for Cancer Therapy
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Precirol ATO 5 was generously provided by Gattefossé USA (Paramus, NJ). Soybean phosphatidylcholine (SPC), Paclitaxel (TAX), Squalene, Tween-80, Mannitol were purchased from Sigma Aldrich (St. Louis, MO). DSPE-PEG (1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)-2000] were obtained from Avanti Polar Lipids (Alabaster, AL). XenoLight DiR obtained from Perkin Elmer (Akron, OH). A modified synthetic analog of Luteinizing Hormone-Releasing Hormone (LHRH) decapeptide (Gln-His-Trp-Ser-Tyr-DLys(D-Cys)-Leu-Arg-Pro) was synthesized according to our design by American Peptide Company, Inc. (Sunnyvale, CA). The sequence of native LHRH peptide, which is similar in human, mouse, and rat, was modified to provide a reactive amino group only on the side chain of a lysine residue, which replaced Gly at the position 6 to yield the superactive, degradation-resistant-Lys-6-des-Gly-10-Pro-9-ethylamide LHRH analog 10 (link)-13 (link). The synthesized sequence of LHRH peptide is highly efficient for targeting of drug delivery systems specifically to the cancer tumors 9 (link), 10 (link), 14 (link)-17 (link) and Cys residue do not influence the recognition process. The remmining materials were obtained from Sigma Aldrich (St. Louis, MO).
10
Isolation and Activation of Human T Cells
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