Fitc labeled concanavalin a con a lectin

Retinal leukostasis was assayed by labeling the leukocytes adherent to retinal endothelium.20 (link) At 24 hours after IR procedure, mice were deeply anesthetized by intraperitoneal injection of a mixture of 100 mg/kg ketamine hydrochloride and 10 mg/kg xylazine hydrochloride and the chest cavity was carefully opened. The perfusion cannula was inserted into the aorta and drainage was achieved by opening the right atrium. PBS was perfused to wash out nonadherent blood cells. Then, FITC-labeled concanavalin A (Con A) lectin (40 μg/mL in PBS, pH 7.4; Vector Laboratories, Burlingame, CA, USA) was perfused to label the adherent leukocytes and vasculature. Next, PBS was perfused to remove the residual unbound Con A. Finally, eyeballs were removed and fixed with 4% PFA. Retinas were dissected and stained with anti-CD45 antibody. The total number of the adherent leukocytes per retina and leukocytes infiltrated into the retina were counted under fluorescence microscopy.