Targeted microbubbles are generated by post-labeling buried ligand microbubbles. According to embodiments, buried ligand microbubbles are created with steric brushes protecting functionalized polymer tethers. Ligands were attached to the functionalized tethers by diffusion of ligands of a small size through the steric barrier. The steric barrier was substantially capable of hindering access to the tethers by larger molecules. The embodiments disclosed include methods for creating microbubble batches that can be loaded with selected ligands, for titrating the targeting ligands thereby to reduce waste and cost, and for using resulting buried ligand molecules for medical purposes.
