Purification of animal immunoglobulin G (IgG) using peptoid affinity ligands

Abstract

The availability of highly pure animal antibodies is critical in the production of diagnostic tools and biosensors. The peptoid PL16, previously isolated from an ensemble of peptoid variants of the IgG‐binding peptide HWRGWV, was utilized in this work as affinity ligand on WorkBeads resin for the purification of immunoglobulin G (IgG) from a variety of mammalian sources and chicken immunoglobulin Y (IgY). The chromatographic protocol initially optimized for murine serum and ascites was subsequently employed for processing rabbit, goat and sheep, donkey, llama, and chicken sera. The PL16‐WorkBeads resin proved able to recover all antibody targets with values of yield between 50 and 90%, and purity consistently above 90%. Notably, PL16 not only binds a broader spectrum of animal immunoglobulins than the reference ligands Protein A and G, but it also binds equally well with all their subclasses. Unlike the protein ligands, in fact, PL16 afforded excellent values of yield and purity of mammalian polyclonal IgG, namely murine (47 and 94%), rabbit (66.5 and 91.7%), caprine IgG (63 and 91–95%), donkey, and llama (93 and 97%), as well as chicken IgY (42 and 92%). Of notice, it is also the ability of PL16 to target monomeric IgG without binding aggregated IgG; when challenged with a mixture of monomeric and aggregated murine IgG, PL16 eluted <3% of fed aggregates, against 11–13% eluted by Protein A and G. Collectively, these results prove the potential of the proposed peptoid ligand for large‐scale purification of animal immunoglobulins.

Purification of polyclonal Immunoglobulin G from human serum using LigaGuardTM and LigaTrapTM adsorbents

“…IgG purification from human Ig-rich paste using LigaTrap TM resin LigaTrap TM resin has been developed by our team for purifying γ-globulins from polyclonal and monoclonal sources. This affinity adsorbent has been used by purifying (i) pIgG from a variety of mammalian sources, including the human, rodent (mouse, rat, and rabbit), caprine (goat and sheep), equine (horse and donkey), camelid (camel and llama); (ii) human monoclonal IgG, IgA, and IgM from Chinese Hamster Ovary (CHO) and Human Embryonic Kidney (HEK) cell culture supernatants [ 33,38 ]; and(iii) avian IgY [ 39 ]. Compared to Protein A/G-based adsorbents, the LigaTrap TM resin features comparable binding capacity and selectivity, improved lifetime [ 32 ], and a substantially lower cost, making it ideal for the chromatographic extraction of pIgG from large volumes of pooled plasma.…”

“…Human IgG resin and LigaGuard TM resin. The peptoid ligand PL-16 was synthesized and conjugated to WorkBeads 40 ACT resin as described in prior work (note: the conjugation strategy and the value of peptoid density on WorkBeads resin are proprietary information of LigaTrap Technologies LLC) [ 32,33 ]. The resulting LigaTrap TM Human IgG resin was rinsed in water and stored in 20% v/v aqueous methanol for long-term storage.…”

“…In this study, a LigaGuard TM adsorbent tailored for IVIg purification from human plasma has been developed, whose peptide ligands bind all non-Ig plasma proteins. Our group has also developed a Ig-targeting peptide- [ 28 ] and peptoid-based [ [29][30][31] ] adsorbents (LigaTrap TM ), which have been demonstrated by purifying pIgG from different mammalian plasma as well as therapeutic mAbs from industrial cell culture fluids.…”

Bio-instructive materials on-demand – combinatorial chemistry of peptoids, foldamers, and beyond

“…1,3,135,136 The applications of these peptoid-based materials span molecular recognition and sensing, 135,[162][163][164][165] membranes, 31,166,167 antifouling coatings, [168][169][170][171][172] the assembly and arrangement of nanoparticles [173][174][175][176][177][178] and chromatography. [179][180][181][182] Some of these unique materials emerged with the help of combinatorial approaches. In a combinatorial Ugi-4CR step-growth polymerization, Stiernet et al assembled peptoid-based polymers.…”

Peptide Affinity Chromatography Applied to Therapeutic Antibodies Purification

“…Unlike protein A, HWRGWV could bind all subclasses of human IgG including IgG 3 . Afterwards, Reese et al ( 2020 ), used a peptoid variant of the peptide HWRGWV to bind IgG from several mammalians as well as chicken immunoglobulin IgY.…”