Vascular ultrasound examinations were performed on all rats before and after operation, luminal diameter and blood flow were measured using a small animal ultrasound scanner with a linear array transducer at 4 weeks after operation. CM-Dil-labeled hucMSCs observed by a fluorescence microscope (Olympus, Japan) was used to examine the homing of transplanted hucMSCs to the site of the vein grafts 3 days after transplantation. Endothelial regeneration was evaluated by CD34 immunofluorescence 14 days after vein grafting as we have previously described [13 (link)]. All the remaining animals were then humanely killed and the vein grafts samples were harvested. The therapeutic effects of hucMSCs were evaluated by subsequent histological and immunohistochemical examination.
Cm dil
Manufactured by Olympus
Sourced in Japan
CM-Dil is a lipophilic, carbocyanine dye used for the fluorescent labeling of cell membranes. It is commonly used in cell tracking and lineage studies.
Automatically generated - may contain errors
Lab products found in correlation
Cm dil, by Thermo Fisher Scientific (3 mentions)
4 6 diamidino 2 phenylindole (dapi), by Olympus (2 mentions)
Uorescence microscope, by Olympus (2 mentions)
Fluorescence microscope, by Olympus (1 mentions)
F1000, by Olympus (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Vector Laboratories (1 mentions)
Formalin solution, by Merck Group (1 mentions)
Fibronectin, by Merck Group (1 mentions)
Cold methanol, by Merck Group (1 mentions)
Ix 73 inverted microscopy, by Olympus (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
Cellsense dimension, by Olympus (1 mentions)
Celltracker cm dil, by Thermo Fisher Scientific (1 mentions)
Ix71 22fl p, by Olympus (1 mentions)
7 protocols using cm dil
1
Vascular Ultrasound and Stem Cell Homing
2
In Vivo Tracking of Labeled Stem Cells
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Human uMSCs were labeled with CM-Dil according to the manufacturer's instructions (Invitrogen, USA). The labeling efficiency was examined under fluorescent microscopy and the viability of labeled cells was confirmed by their ability to grow in vitro. After labeling, cells were washed with PBS and 3×106 cells were injected into the STZ-treated rats as above. Rats were euthanized at the indicated times and paraffin sections (4-µm thickness) of selected tissues were stained with DAPI (Vectashield, USA) for 5 min at room temperature. The CM-Dil and DAPI staining was visualized and acquired by the fluorescent microscopy (Olympus f1000, Japan).
3
Exosome Internalization Dynamics in Cells
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To observe the internalization of exosomes, BMSC-Exos and MC3T3-E1 cells were labeled by fluorescence dye CM-Dio (Thermo Fisher Scientific Inc., Waltham, MA, USA) and CM-Dil (Thermo Fisher Scientific Inc., Waltham, MA, USA), respectively, as previously described [50 (link)]. Briefly, 20 μg exosomes was diluted with 100 μL PBS and incubated with CM-Dio in the dark for 30 min, washed with PBS, ultracentrifuged at 120,000× g for 70 min to remove nonbinding dye, then resuspended in PBS. MC3T3-E1 cells were incubated with CM-Dil in the dark for 30 min and centrifuged at 300× g for 5 min to remove nonbinding dye. The CM-Dio labeled exosomes were incubated with CM-Dil-labeled MC3T3-E1 cells at 37 °C for 48 h. After the incubation, a fluorescence microscope (BX53, Olympus, Tokyo, Japan) was used to collect and analyze the fluorescence images.
4
Evaluating Engineered hNSCs Migration to Metastatic CRC
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To examine whether engineered hNSCs can migrate to lymph node–derived metastatic colorectal adenocarcinoma, SW-620 cells (1×105 cells per well) and human dermal fibroblast cells (a control, 1×105 cells per well) were plated in a 24-well plate containing 1% CD-FBS phenol free DMEM medium with incubation for 24 hours. The bottom surface of transwell plates (0.4 μm, BD Biosciences, Dickinson, Franklin Lakes, NJ) coated the with fibronectin (250 μg/mL, Sigma-Aldrich) was placed in the 24-well plates and CM-Dil (Invitrogen Life Technologies) pre-stained engineered hNSCs were plated in the upper chambers of the transwell plates at a density of 1×105 cells per well in the same condition medium and cultured for 24 hours at 37°C. After washing the lower chamber, the cells were fixed with 10% formalin solution (Sigma-Aldrich) for 10 minutes and permeabilized using 100% cold-methanol (Sigma-Aldrich) for 10 minutes. Then, DAPI (4',6-diamidino-2-phenylindole, Invitrogen Life Technologies) was added to the lower chamber at 300 nM and the plates were incubated for 10 minutes at 37°C followed by washing with PBS. Cells stained with CM-Dil and DAPI were examined by fluorescence microscopy (IX-73 Inverted Microscopy, Olympus, Tokyo, Japan) and counted by Cell Sense Dimension.
5
Labeling ADSCs with CM-Dil Fluorescent Dye
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ADSCs were washed with PBS and incubated with 2 μM Cell Tracker CM-Dil (Molecular Probes Inc., Eugene, OR) at 37°C for 5 min and then for an additional 15 min at 4°C. The cells were washed with PBS and suspended in PBS at 2 × 107 cells/mL. CM-Dil-labeled ADSCs were observed under a fluorescence microscope (Olympus, Tokyo, Japan; model IX7122FL/P) at an excitation wavelength of 553 nm.
6
Vascular Regeneration with hucMSCs
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Vascular ultrasound examinations were performed on all rats before and after operation, luminal diameter and blood ow were measured using a small animal ultrasound scanner with a linear array transducer at 4 weeks after operation. CM-Dil labeled hucMSCs observed by a uorescence microscope (Olympus, Japan) was used to examine the homing of transplanted hucMSCs to the site of the vein grafts 3 days after transplantation. Endothelial regeneration was evaluated by CD34 immuno uorescence 14 days after vein grafting as we have previously described 13 .All the remaining animals were then humanely killed and the vein grafts samples were harvested. The therapeutic effects of hucMSCs was evaluated by subsequent histological and immunohistochemical examination.
7
Vascular Regeneration with hucMSCs
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Vascular ultrasound examinations were performed on all rats before and after operation, luminal diameter and blood ow were measured using a small animal ultrasound scanner with a linear array transducer at 4 weeks after operation. CM-Dil labeled hucMSCs observed by a uorescence microscope (Olympus, Japan) was used to examine the homing of transplanted hucMSCs to the site of the vein grafts 3 days after transplantation. Endothelial regeneration was evaluated by CD34 immuno uorescence 14 days after vein grafting as we have previously described 13 .All the remaining animals were then humanely killed and the vein grafts samples were harvested. The therapeutic effects of hucMSCs was evaluated by subsequent histological and immunohistochemical examination.
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