Detecting cell adhesion molecules in intact lung using quantum dot conjugates targeted to endothelial cells

Abstract

Quantum dots (QD) coupled with fluorescence imaging techniques are a powerful tool for detecting proteins in vivo. Here we used dual QD labeling to detect cellular adhesion molecules, CAM (endothelial marker, PECAM and vascular cell adhesion molecule, VCAM) expression in a LPS model of lung inflammation. QDs were functionalized via an adaptor protein and linked to anti‐PECAM and anti‐VCAM Abs to create QD655‐PECAM (QDP) and QD525‐VCAM (QDV), respectively. We established probe specificity in vitro in a PECAM expressing cell line and in LPS treated endothelial cells (for VCAM). For in situ experiments, mice lungs were removed at 24h after LPS instillation, and perfused with medium containing QDV and QDP. For in vivo lung studies, QDs were injected into the jugular vein at 24h after LPS treatment and lungs were removed and processed after overnight fixation. Imaged sections from both studies showed that LPS treated mice had 8–10 fold higher QDV signal, compared to naïve lungs, which significantly colocalized with QDP, indicating increased VCAM expression in pulmonary endothelium. Thus, QD labeling for intravital microscopy will be useful for detection and localization of proteins in pulmonary tissue.