Abstract
Aim:
To perform a site-specific conjugation of Fab′ fragments of a mouse monoclonal antibody(MoAb) B43(of IgG1 subtype) to a bifunctional chelator 6-[p- (bromoacetamido) benzyl]-1,4,8,11-tetraazacyclotetradecane-N,N′,N″,N′″-tetraacetic acid (BAT) via the thiol groups in the hinge distal to the antigenbinding site of the Fab′.
Methods:
B43 was cleaved using a simple 2-step method. First, stable F(ab′)2 was produced by pepsin treatment. Fab′ with free thiol in the hinge region was then obtained by cysteine reduction of F(ab′)2. Second, a site-specific conjugation of Fab′ to thiol-specific BAT was performed in a one-step reaction.
Results:
The Fab′ fragment had approximately 1.8 free thiol groups per molecule after cysteine reduction. The conjugation efficiency and the chemical yield were approximately 1.28 moles chelator/Fab′ and 74% of the initial concentration of Fab′, respectively. The F(ab′)2, Fab′ and Fab′-BAT all maintained reasonable antigen-binding properties. 67Cu labeling of the conjugate under standard conditions did not impair the immunoreactivity of Fab′-BAT.
Conclusion:
This is a simple and efficient method for producing immunoreactive conjugates of Fab′- BAT, which can be used to make radiometal-labeled conjugates for further diagnostic and therapeutic applications.
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Li, J., Wang, Xh., Wang, Xm. et al. Site-specific conjugation of bifunctional chelator BAT to mouse IgG1 Fab' fragment. Acta Pharmacol Sin 27, 237–241 (2006). https://doi.org/10.1111/j.1745-7254.2006.00242.x
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DOI: https://doi.org/10.1111/j.1745-7254.2006.00242.x
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