Bioconjugation of biologically useful proteins is in great demand (e.g., conjugation to biotins, metal chelators, and drug carriers to target specific tissues for both in vitro and in vivo use). These conjugates provide widespread opportunities for various biological and biomedical applications. Evolving state-of-the-art protein conjugation strategies have led to the development of many affinity ligands, including for cancer imaging and diagnosis. However, to achieve the desirable protein conjugates, there are many challenges that remain to be addressed in order to obtain a reproducible procedure for all proteins and ligands. These include a control over the protein modification and the efficiency of the conjugation while retaining the original biological protein affinity postmodification. Here we present detailed conjugation methods for the human fibronectin tenth type III domain (FN3) protein scaffold for use in preclinical PET imaging. More specifically, this chapter provides detailed methods to produce a FN3 and a FN3-chelator-conjugate, its labeling with the radionuclide 64-Cu, and its use for noninvasive PET imaging in mice.
Keywords: 64-Cu; DOTA; FN3 protein; PET/CT; hCD20.