Cystine knot peptides, also know as knottins, are an emerging class of biotherapeutic molecules that can be engineered to bind to a diverse range of targets. Much like therapeutic antibodies, there is a need to develop critical reagents for pharmacokinetic (PK) and immunogenicity assays to support preclinical development, though this new class of molecules can present different challenges and considerations for reagent discovery. To this end, monoclonal and polyclonal reagent antibodies against a cystine-knot fusion protein (CKFP) were generated and used for PK and ADA assay development.
The final optimized anti-knottin (peptide) PK assay gave sensitivity down to 15.6 ng/mL CKFP, while the optimized anti-fusion protein PK assay gave sensitivity down to 0.469 ng/ mL CKFP.
In the ADA Bridging ELISA, the anti-knottin peptide antibody gave higher sensitivity (below 100 ng/mL) compared to the antifusion protein antibody (below 200 ng/mL).
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Joshua K. Lowitz, Michael Trang, Catherine Vo, Glen Lin, Jennifer Somera, Rick Chang, Shannon Jones-Iatauro, Ronald Gamatero, and John S. Kenney - Antibody Solutions, Sunnyvale, CA, USA