Interest in the use of targeted microbubbles for ultrasound molecular imaging (USMI) has been growing in recent years as a safe and efficacious means of diagnosing tumor angiogenesis and assessing response to therapy. Of particular interest are cloaked microbubbles, which improve specificity by concealing the ligand from blood components until they reach the target vasculature, where the ligand can be transiently revealed for firm receptor-binding by ultrasound acoustic radiation force pulses. Herein, a bio-orthogonal “click” conjugation chemistry is introduced to decorate the surface of cloaked 4-5 μm diameter microbubbles as part of a sterile and reproducible production process. Azido-functionalized antagonists for the angiogenic biomarkers αVβ3 integrin (cRGD) and VEGFR2 (A7R) proteins were conjugated to bimodal-brush microbubbles via strain-promoted [3+2] azide-alkyne cycloaddition (SPAAC) click chemistry. Ligand conjugation was validated by epifluorescent microscopy, flow cytometry and Fourier-transform infrared spectroscopy. Sterility was validated by bacterial culture and endotoxin analysis. Additionally, clinically normal dogs receiving escalating microbubble doses were shown to experience no pathologic changes in physical examination, complete blood count, serum biochemistry profile or coagulation panel. This bio-orthogonal microbubble conjugation process for cloaked peptide ligands may be leveraged for future USMI studies of tumor angiogenesis for translation to pre-clinical and clinical applications.