Comprehensive Immunostaining of Retinal Vasculature

Retinal vasculature and CNV lesions were immunostained with directly conjugated Alexa Fluor 488-Isolectin B4, Alexa Fluor 594-Isolectin B4, or Alexa Fluor 647-Isolectin B4 (Sigma). Endothelial cell junctions and phosphorylated VE-cadherin were stained with anti-VE-cadherin antibody (1:100; BD Rat 555289) and affinity purified rabbit antibodies against VE-cadherin pY658 and pY685 (Orsenigo et al., 2012 (link)). Phosphorylated c-Src was assessed using anti-phospho-Src (Tyr418) Antibody (1:100; Invitrogen Rabbit 44–660G). Secondary antibodies used were Alexa488 anti-Rat (1:500; Invitrogen Donkey A-21208) and Alexa555 anti-Rabbit (1:500; Donkey A-31572). Inflammatory cells were stained with anti-CD45 (1:300; BD Biosciences Goat 553076) and anti-CD68 (1:300; BioRad Rat MCA1957). Secondary antibodies used were Alexa488 anti-Rat (1:500; Invitrogen Donkey A-21208). Alexa555 anti-Goat (1:500; Invitrogen Donkey A-21432).