Fetal Vascular Labeling via Intracardiac Lectin Injection

Prior to fetal harvest, we performed an in utero intracardiac injection of fluorescein-conjugated tomato-lectin (TL) for CDH fetuses in utero as previously described (Shah et al., 2011 (link)). An ultrasound microscope probe was used to guide the injection apparatus. Sub-microliter injection system (Nanofil-100, World Precision Instruments, Sarasota, FL, United States of America) was used to inject TL into the heart. Each fetal heart was injected with 40 μL of TL. The injected fluorescent-conjugated lectin was allowed to circulate for 10 min to ensure adequate binding to the vascular endothelial wall of the vasculature, after which the fetus was harvested and fixed in 4% PFA for immunofluorescent analyses.