Mice were intravenously injected with DiI-labeled LNP encapsulating a mixture of siRNA (siGL4:siCy5-green fluorescent protein [GFP] 1:1 ratio) at a dose of 0.5 mg/kg of both siRNAs. After 1 hour, the mice were sacrificed, and their liver tissues (0.5–1 cm2) were collected and immersed in staining solution (Dulbecco’s phosphate-buffered saline [D-PBS (-)]) containing 20 µg/mL FITC-labeled Isolectin B4 (Vector Laboratories, Burlingame, CA, USA) for staining blood vessels and 1 µg/mL Hoechst33342 (Dojindo, Kumamoto, Japan) for staining nuclei. Intrahepatic distribution of the LNPs was observed using confocal laser scanning microscopy (CLSM; Nikon A1, Nikon, Tokyo, Japan).
Fitc labeled isolectin b4
Manufactured by Vector Laboratories
Sourced in United States
FITC-labeled Isolectin B4 is a fluorescently labeled lectin derived from the plant Griffonia simplicifolia. It binds specifically to α-D-galactose and N-acetyl-α-D-galactosamine residues on cell surfaces.
Automatically generated - may contain errors
Lab products found in correlation
Fluorescence microscope, by Zeiss (2 mentions)
Hoechst 33342, by Dojindo Laboratories (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Beyotime (1 mentions)
Embedding medium, by Sakura Finetek (1 mentions)
Fluorescence mounting medium, by Agilent Technologies (1 mentions)
Image pro plus, by Media Cybernetics (1 mentions)
3 protocols using fitc labeled isolectin b4
1
In Vivo Intrahepatic LNP Tracking
2
Immunohistochemical Analysis of Retinal Endocan
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Eyeballs from control and OIR mice were dissected and rapidly frozen in embedding medium (Sakura Finetek, Torrance, CA). Retina sections (10‐μm thick) were thawed, air‐dried, and fixed in 4% paraformaldehyde at room temperature for 10 min. After blocking with 10% FBS in PBS for 1 hr, sections were incubated with goat anti‐mouse endocan Ab (R&D Systems) overnight at 4°C, followed by incubation with an Alexa Fluor 555‐conjugated donkey anti‐goat secondary Ab (1:500; Invitrogen) and FITC‐labeled isolectin B4 (1:50; Vector Laboratories Inc., Burlingame, CA) for 1 hr at room temperature. Sections were then rinsed in PBS and stained with DAPI (Beyotime Biotechnology, Shanghai, China) for 5 min. Images were captured using a fluorescence microscope (Carl Zeiss Microscopy, Thornwood, NY).
3
Inhibiting Oxygen-Induced Retinal Neovascularization
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C57BL/6 mice were exposed to 75% oxygen from postnatal day 7 (P7) with their nursing mother and returned to room air at P12. These mice were subsequently randomly divided into five groups. One eye received an intravitreal injection of 1 lL mouse endocan neutralizing antibody (NAb, R&D Systems, Minneapolis, MN, USA) at a concentration of either 0.01, 0.1, 0.5, or 1 lg/lL; the contralateral eye was treated with PBS. At P17, mice were euthanized and eyes were prepared for immunofluorescent flat-mounts with FITC-labeled isolectin B4 (1:50; Vector Laboratories, Inc., Burlingame, CA, USA) for 40 minutes, retinas were washed three times in PBS and flatmounted with fluorescence mounting medium (DAKO, Agilent Technologies, CA, USA) and sealed with a cover slip. Digital photographs were obtained with a fluorescence microscope (Carl Zeiss Microscopy, Thornwood, NY, USA) at 35 magnification, and images were merged into a single image to show the entire retina using the photomerge option of a raster graphics editor (Photoshop CS 6.0; Adobe Systems, San Jose, CA, USA). Imaging software (Image Pro Plus; Media Cybernetics, Inc., Rockville, MD, USA) was used to measure the area of retinal NV per retina by a blinded investigator.
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