Fragments of protein A eluted during protein A affinity chromatography

Carter-Franklin, Jayme N.; Victa, Corazon; McDonald, Paul C.; Fahrner, Robert

doi:10.1016/j.chroma.2007.06.012

Cited by 77 publications

(42 citation statements)

References 19 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…One factor is the amount of Protein A that is leached with each cycle. A recent study by Carter-Franklin et al 36 demonstrated that intact Protein A leaches when loading either purified antibody or unpurified antibody in harvested cell culture fluid and additional fragments of Protein A leach when loading unpurified antibody in harvested cell culture fluid. The amount of leached Protein A can be reduced by addition of EDTA in harvested cell culture fluid to inhibit proteinases.…”

Section: Chromatographic Processesmentioning

confidence: 99%

Recovery and purification process development for monoclonal antibody production

Liu

Winter

et al. 2010

mAbs

463

411

View full text Add to dashboard Cite

Section: Chromatographic Processesmentioning

confidence: 99%

Recovery and purification process development for monoclonal antibody production

Liu

Winter

et al. 2010

mAbs

463

411

View full text Add to dashboard Cite

“…However, it suffers from problems regarding ligand stability and cost, since protein A resin is very expensive and readily amenable to chemical and biological degradation. 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

View full text Add to dashboard Cite

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.

show abstract

“…Immobilized protein A, G and L possessing affinity for type 1-4 of IgGs can be used for the isolation of MAbs from ascites or culture supernatant (Table l) (Roque et al, 2005a(Roque et al, , 2007Hahn et al, 2006;Carter-Franklin et al, 2007;Pavlovic et al, 2007). Protein A is a cell wall protein of S. aureus, which binds selectively to Fc region of IgG but cannot form a complex with human IgG 3 .…”

Section: Purification Of Antibodies On Immobilized Proteins a G And Lmentioning

confidence: 99%

Separation of antigens and antibodies by immunoaffinity chromatography

Sheng

Kong

2012

Pharmaceutical Biology

View full text Add to dashboard Cite

Fragments of protein A eluted during protein A affinity chromatography

Cited by 77 publications

References 19 publications

Recovery and purification process development for monoclonal antibody production

Recovery and purification process development for monoclonal antibody production

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

Separation of antigens and antibodies by immunoaffinity chromatography

Contact Info

Product

Resources

About