Small molecule fluorophore and copolymer RGD peptide conjugates for ex vivo two-photon fluorescence tumor vasculature imaging

We report the use of small molecule and block copolymer RGD peptide conjugates for deep ex vivo imaging of tumor vasculature in "whole" excised tumors using two-photon fluorescence microscopy (2PFM). The fluorescent probes were administered to mice via tail-vein injection, after which the tumors were excised, fixed, and imaged without further sample preparation. Both RGD conjugates demonstrated specific targeting to tumor blood vessels, and this selectivity imparted excellent contrast in 2PFM micrographs that captured high-resolution 3-D images of the tumor vasculature up to depths of 830 μm in Lewis Lung Carcinoma (LLC) tumors. 2PFM ex vivo fluorescence micrographs clearly revealed tumor vessels, while differences in the sensitivity of tumor vessel imaging were apparent between the small molecule and block copolymer conjugates. Both the small molecule and polymer-based two-photon absorbing probe conjugate are valuable for deep tissue tumor microvasculature imaging.