Evaluation of commercial chromatographic adsorbents for the direct capture of polyclonal rabbit antibodies from clarified antiserum

Bak, Hanne; Thomas, Owen R.T.

doi:10.1016/j.jchromb.2006.07.003

Cited by 46 publications

(32 citation statements)

References 37 publications

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“…A second generation mimetic resin, Mabsorbent A2P, had a higher DBC compared to the original A1P, 10.4 versus 8.2 mg/mL CV, though both were significantly lower than the other resins tested in this group. This observation is consistent with other studies performed by Bak and Thomas (2007) and Ghose et al (2006).…”

Section: Protein a Mimetic And Mixed-mode Interaction Resins-evaluatsupporting

confidence: 94%

“…As an alternative to protein A, resins with immobilized biomimetic ligands (Li et al, 1998;Roque et al, 2004;Teng et al, 1999) and mixed mode interaction resins (Boschetti et al, 2000;Schwartz et al, 2001;Subramanian, 2005) have become commercially available in recent years. Although Protein A resins are the leading choice for purification of MAbs, examples of non-affinity-based purification schemes for monoclonal antibodies have been discussed in the literature (Boschetti, 2001;Follman and Fahrner, 2004;Gagnon, 1996;Bak et al, 2007;Ghose et al, 2006;Lim et al, 1998;Vancan et al, 2002). Researchers have also investigated the possibility of increasing the productivity of MAb purification by combining primary recovery with protein A through the use of expanded bed chromatography (Ohashi et al, 2002;Thommes et al, 1996).…”

Section: Introductionmentioning

confidence: 99%

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Maximizing productivity of chromatography steps for purification of monoclonal antibodies

Tugçu

Roush

Göklen

2007

Biotech & Bioengineering

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The large scale production of monoclonal antibodies presents a challenge to design efficient and cost effective downstream purification processes. We explored a two stage resin screening approach to identify the best candidates to be utilized for the platform purification of monoclonal antibodies. The study focused on commercially available affinity resins including Protein A, mimetic and mixed-mode interaction resins as well as ion exchangers used in polishing steps. An initial screening using pure proteins was followed by a final screening where selected resins were utilized for the purification of MAbs in complex mixtures. Initial screenings aimed to measure the theoretical upper limit for dynamic binding capacity (DBC) at 1% breakthrough and productivity. We confirmed that DBC of affinity, mimetic and mixed-mode resins was a strong function of the linear velocity used for loading. Productivities >27 g/(L-h), were obtained for rProtein A FF, Mabselect and Prosep rA Ultra at 2 min residence time. For the cation exchangers, we identified UNOsphere S and Fractogel SO(3) as the best candidates for our purification based on DBC. For anion exchangers operated in flowthrough mode, Q Sepharose XL and UNOsphere Q were selected from the initial screening based on DBC and resolution of IgG from BSA. Finally, a three step purification scheme was implemented using the selected affinity and ion exchangers for the purification of IgG from complex feedstocks. We found that Mabselect followed by UNOsphere Q and UNOsphere S provided the best purification scheme for our applications based on productivity.

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Section: Protein a Mimetic And Mixed-mode Interaction Resins-evaluatsupporting

confidence: 94%

Section: Introductionmentioning

confidence: 99%

Maximizing productivity of chromatography steps for purification of monoclonal antibodies

Tugçu

Roush

Göklen

2007

Biotech & Bioengineering

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“…In addition, protein A leaching is a serious concern (Carter-Franklin et al, 2007), since protein A is known to cause severe immunogenic responses in humans (Terman and Bertram, 1985;. Non-affinity-based purification schemes for mAbs have also been the focus of much discussion in the literature (Li et al, 1998;Teng et al, 1999;Boschetti, 2001;Vancan et al, 2002;Follman and Fahrner, 2004;Ghose et al, 2006;Bak and Thomas, 2007;Ramessar et al, 2008).…”

Section: Introductionmentioning

confidence: 99%

Combinatorial de novo design and application of a biomimetic affinity ligand for the purification of human anti‐HIV mAb 4E10 from transgenic tobacco

Platis

Maltezos

K‐C.

et al. 2009

J of Molecular Recognition

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Monoclonal anti-HIV antibody 4E10 (mAb 4E10) is one of the most broadly neutralizing antibodies against HIV, directed against a specific epitope on envelope protein gp41. In the present study, a combinatorial de novo design approach was used for the development of a biomimetic ligand for the affinity purification of mAb 4E10 from tobacco transgenic extract in a single chromatographic step. The biomimetic ligand (4E10lig) was based on a L-Phe/beta-Ala bi-substituted 1,3,5-triazine (Trz) scaffold (beta-Ala-Trz-L-Phe, 4E10lig) which potentially mimics the more pronounced electrostatic and hydrophobic interactions of mAb 4E10-binding sequence determined by screening of a random peptide library. This library was comprised of Escherichia coli cells harboring a plasmid (pFlitrx) engineered to express a fusion protein containing random dodecapeptides that were inserted into the active loop of thioredoxin, which itself was inserted into the dispensable region of the flagellin gene. Adsorption equilibrium studies with this biomimetic ligand and mAb 4E10 determined a dissociation constant (K(D)) of 0.41 +/- 0.05 microM. Molecular modeling studies of the biomimetic ligand revealed that it can potentially occupy the same binding site as the natural binding core peptide epitope. The biomimetic affinity adsorbent was exploited in the development of a facile mAb 4E10 purification protocol, affording mAb 4E10 of high purity (approximately 95%) with good overall yield (60-80%). Analysis of the antibody preparation by SDS-PAGE, enzyme-linked immunosorbent assays (ELISA), and western blot showed that the mAb 4E10 was fully active and free of degraded variants, polyphenols, and alkaloids.

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“…The quality of the packing was evaluated by residence time distribution analysis employing 1% acetone as tracer [20]. Biomass deposition studies were done by injecting a 4 ml biomass pulse (OD at 600 nm ≈ 0.8 AU).…”

Section: Bio-colloid Deposition Experimentsmentioning

confidence: 99%

Surface energetics to assess biomass attachment onto hydrophobic interaction adsorbents in expanded beds

Vennapusa

Tarı

Cabrera

et al. 2009

Biochemical Engineering Journal

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a b s t r a c tCell-to-support interaction and cell-to-cell aggregation phenomena have been studied in a model system composed of intact yeast cells and Phenyl-Streamline adsorbents. Biomass components and beaded adsorbents were characterized by contact angle determinations with three diagnostic liquids and zeta potential measurements. Subsequently, free energy of interaction vs. distance profiles between interacting surfaces was calculated in the aqueous media provided by operating mobile phases. The effect of pH and ammonium sulphate concentration within the normal operating ranges was evaluated. Calculation indicated that moderate interaction between cell particles and adsorbent beads can develop in the presence of salt. Cell-to-cell aggregation was suspected to occur at high salt concentration and neutral pH. Predictions based on the application of the XDLVO approach were confirmed by independent experimental methods like biomass deposition experiments and laser diffraction spectroscopy. Understanding biomass attachment onto hydrophobic supports can help in alleviating process limitations normally encountered during expanded bed adsorption of bioproducts.

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Evaluation of commercial chromatographic adsorbents for the direct capture of polyclonal rabbit antibodies from clarified antiserum

Cited by 46 publications

References 37 publications

Maximizing productivity of chromatography steps for purification of monoclonal antibodies

Maximizing productivity of chromatography steps for purification of monoclonal antibodies

Combinatorial de novo design and application of a biomimetic affinity ligand for the purification of human anti‐HIV mAb 4E10 from transgenic tobacco

Surface energetics to assess biomass attachment onto hydrophobic interaction adsorbents in expanded beds

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