P6 mice were anesthetized by intraperitoneal injection of ketamine and xylazine (100 mg/kg body weight), the skin over the temporal vein was removed, and 15 μL of 1 mg/mL Alexa Fluor 568-conjugated isolectin GS-IB4 (Thermo Fisher Scientific, CAT#I21412) in PBS with 1 mM CaCl2 were injected into temporal vein. 10 min after the injection, the eyes were removed and fixed in 2% PFA/PBS for 2 hours at room temperature. Retinas were dissected, permeabilized with PBST, and stained as described above.
Alexa fluor 568 conjugated isolectin gs ib4
Manufactured by Thermo Fisher Scientific
Alexa Fluor 568-conjugated isolectin GS-IB4 is a fluorescently labeled lectin used for the detection and visualization of endothelial cells and microvascular structures. The Alexa Fluor 568 dye provides a bright red fluorescent signal.
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Lab products found in correlation
2 protocols using alexa fluor 568 conjugated isolectin gs ib4
1
Temporal Vein Labeling in Mice
2
Retinal Vessel Morphology Quantification
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Retinal vessels were visualized by staining flat mounts with Alexa Fluor 568-conjugated isolectin GS-IB4 (I21412; Thermo Fisher Scientific) for 1 day at 4°C prior to confocal microscopy. ImageJ software (National Institutes of Health, Bethesda, MD, USA) was used to determine the effects of rAAV2-shmTOR-GFP activity on the rat OIR model. Neovascularization was quantified by manually outlining the areas wherein neovascular tufts were formed and comparing their sum amount with the total area of the flat mount. Calculating the percentage of avascular areas was carried out in a similar manner. Vessel tortuosity was determined by drawing a straight line from the terminus of a retinal artery to where it begins toward the center of the retina. Then, after manually drawing a line that follows the path of said artery, the two values were compared with yield a vessel tortuosity ratio.
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