Production Of Disulfide-Rich Cow Knob Domain Antibodies: Picobodies™

Bovine CDR H3 knob domains, sized at 4-6 kDa, can access epitopes that conventional antibodies cannot reach, making them attractive candidates for inhaled therapeutics and precision targeting applications. The core challenge is achieving soluble, correctly folded product at meaningful titers when multiple disulfide bonds must form accurately.

Using a microbial expression platform with a broad toolbox of genetic elements, fusion partners, and engineered host strains, researchers screened candidates at 96-well scale and identified strains yielding up to 2 g/L soluble fusion protein. SDS-CGE provided rapid solubility assessment within 90 minutes per plate, while biolayer interferometry distinguished highly active clones from baseline within 30-60 minutes per plate. RP-UPLC and intact LC-MS confirmed expected cleavage of the fusion tag, and ELISA validated binding to the SARS-CoV-2 spike protein at an IC50 of 19 nM.

If you are evaluating expression systems for disulfide-rich miniprotein candidates, access the full case study below to learn more.