IgG purification from human plasma by membrane affinity chromatography with immobilized Cu(II) and Ni(II) : effect of quelators IDA, TREN and CM-Asp / Purificação de IgG a partir do plasma humano por cromatografia em membranas con ions Cu(II) e Ni(II) imobilizados : efeito dos agentes quelantes IDA, TREN e CM-Asp

AUTOR(ES)

Mariana Borsoi Ribeiro

DATA DE PUBLICAÇÃO

2006

RESUMO

The use of immunoglobulin G (lgG) in therapeutic applications has growing vastly and it is used in the treatment of a growing number of indications. Several chromatographic techniques have been investigated for IgG purification. In this work, we investigated the effect of the chelators iminodiacetic acid (IDA), carboxymethylaspartate acid (CM-Asp) and Tris-2(aminoethyl)amine (TREN) for IgG purification, from human plasma using Immobilized Metal-ion Affinity Chromatography (IMAC) technique using hollow fibers membranes as support to the chromatographic experiments. For that, adsorption experiments were done, using different buffers systems, on filtration module and on finely cut poly(ethylene) vinyl alcohol (PEV A) membranes containing chelators IDA, TREN and CM-Asp with the metallic ions nickel and copper. The adsorbent selectivity was verified through electrophoresis SDS-PAGE and nefelometric analysis. Best purification conditions were found with PEV A-CM-Asp-Ni(ll) filtration module in the presence of 25 mM Tris HCI pH 7.0 and elution by increasing Tris concentration. In this case, it was possible to obtain IgG with approximately 90% of purity. According to electrophoresis SDS-P AGE and nefelometric analysis, PEV A-CM-Asp-Ni(ll) filtration modules showed better selectivity and superior adsorption dynamic capacity for IgG than those obtained by finely cut membranes (medium adsorption capacity: 53.0 and 7.7 mg of IgG /g of dry membrane, respectively). Lagmüir model adjusted to adsorption isotherms at room temperature (250 C), showed high adsorption capacity for PEV A-IDA-Ni(ll) and PEV A-CM-Asp-Ni(ll) systems (204.6 and 302.3 mg/g dry of membrane, respectively) and showed dissociation constants characteristic of medium affinity systems (6.1 and 10.1 x 10-6 M, respectively). The thermodynamic parameters analysis obtained for PEV A-IDA-Ni(ll)-lgG system indicated the complexity of the interactions, and the existence of hydrophobic and electrostatics interaction besides the coordination bound. In this work, we could purify IgG from human plasma in a single stage, using the affinity chromatographic membranes technique, allowing in this way, the treatment of great volumes per unit of time, showing the high potentiality of this method in industrial processes

ASSUNTO(S)

imunoglobulinas g membranes cromatografia de afinidade membranas purification ions metalicos imunoglobulin g affinity chrmatography purificação metal ions

Documentos Relacionados

• Purification on monoclonal antibodies using IMAC in PEVA hollow fiber membranes: comparison of chelating agents IDA, CM-Asp and TREN
• The physico-chemical properties of 2-methoxyphenoxyacetates of Mn(II), Co(II), Ni(II) and Cu(II)
• The physico-chemical properties of 4-chlorophenoxyacetates of Mn(II), Co(II), Ni(II) and Cu(II)
• Caracterização magnética, térmica e espectral de 2,6-dimetóxibenzoatos de Co(II), Ni(II) e Cu(II)
• Efeito do íon metálico, do sistema tamponante e do sal (NaCl) na adsorção de IgG humana em fibras ocas derivatizadas com CM-Asp