A two-amino acid mutation in murine IgA enables downstream processing and purification on staphylococcal superantigen-like protein 7.

With few exceptions, all currently marketed antibody therapeutics are IgG molecules. One of the reasons that other antibody isotypes are less developed are the difficulties associated with their purification. While commercial chromatography affinity resins, like staphylococcal superantigen-like 7 (SSL7) protein-containing resin, allow purification of IgAs from many animal species, these are not useful for murine IgAs. Because the mouse model is predominantly used for preclinical evaluation of IgA-based therapeutics, there is a need to develop an effective purification method for mouse IgA. Here, we adapted the sequence of a mouse IgA by mutating two amino acid residues in the fragment crystallizable (Fc) sequence to facilitate its purification on SSL7 resin. The mutated IgA Fc (hereafter referred to as IgA*) was then genetically fused to the variable domain of a llama heavy chain-only antibody (VHH) directed against the fusion protein of human respiratory syncytial virus (HRSV), resulting in VHH-IgA*, and transiently produced in infiltrated Nicotiana benthamiana leaves. These plant-produced mouse VHH-IgA* fusions were enriched by SSL7 affinity chromatography and were found to be functional in ELISA and could neutralize RSV in vitro, suggesting no detrimental effect of the mutation on their antigen-binding properties. This approach for the purification of murine IgA will facilitate downstream processing steps when designing innovative murine IgA-based fusions.

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