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Optimization of Radiolabeling methods of His-tagged single-chain antibody fragments (scFvs) with technetium tricarbonyl 99mTc(CO)3 as a molecular imaging agent
* 1 , 2 , * 3
1  Department of Nuclear Pharmacy, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, Iran
2  Protein Technology Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
3  Department of Pharmaceutical Chemistry, School of Pharmacy, Shahid Beheshti University of Medical Sciences, Tehran, Iran
Academic Editor: Maria Emília Sousa


Single-chain antibody fragments (scFvs) are considered more valuable agents for clinical imaging compared with parent antibodies due to their rapid tumor uptake and high tumor-to-background ratios at early times.

A number of studies reported that biomolecules can be radiolabeled with 99mTc in a high yield using indirect labeling techniques or non-site-specific conjugations, but these approaches can reduce their biological activities. His-tags are specific sites for radiolabeling of scFvs with 99mTc-tricarbonyl ([99mTc(CO)3]+). According to the reported studies, radiolabeling efficiency of His-tag-containing biomolecules varies unpredictably and depends on a series of various factors including the structure of the biomolecule and the conditions of radiolabeling.

Therefore, in this study, we tried to radiolabel two His-tagged scFvs with precursor complex of 99mTc(CO)3 and evaluate their radiolabeling efficiencies in different conditions including type of buffer, volume, ionic strength, pH, the concentration of antibody, and activity, temperature, and time using Thin-layer chromatography (TLC) and gamma counter.

His-tagged scFvs were radiolabeled with [99mTc(CO)3] + in 100% radiochemical purity at a range of 480-600 µCi/µg for 2 h at 50°C. We demonstrated that the radiochemical purity of radiolabeled His-tagged scFvs increases with higher [99mTc(CO)3]+ activity, and antibody concentrations in a smaller volume, increasing the temperature and pH of the reaction medium. Moreover, the radiolabeled His-tagged scFvs showed high stability for 24 h in the reaction medium. In the present study, the optimal and efficient radiolabelling of His-tagged scFvs successfully obtained that they can be used as potential agents for in vivo imaging.

Keywords: His-tagged scFvs, 99mTc tricarbonyl, Radiolabelling, Molecular imaging