Novel hydrophobic interaction chromatography matrix for specific isolation and simple elution of immunoglobulins (A, G, and M) from porcine serum

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• 作者：Gabriela Ramos-Clamont ; Maria del Carmen Candia-Plata ; Roberto Guzman Zamudio and Luz Vazquez-Moreno
• 关键词：Blood derivatives ; Hydrophobic interaction chromatography ; Highly acetylated agarose ; Porcine immunoglobulins ; T-gel
• 刊名：Journal of Chromatography A
• 出版年：2006
• 出版时间：28 July 2006
• 年：2006
• 卷：1122
• 期：1-2
• 页码：28-34
• 全文大小：184 K

文摘

A new, highly acetylated agarose matrix (HA-Sepharose) was synthesized and used as a hydrophobic interaction chromatography (HIC) medium to specifically isolate immunoglobulins (Igs) from porcine serum. Recovery of Igs was in a single step and under mild conditions. HA-Sepharose adsorption was studied in terms of salt, gel acetylation time, flow rate, and protein concentration on the loading buffer. At 0.5 M Na₂SO₄, control with unmodified Sepharose retained a small fraction (0.70 mg/mL of matrix) of serum albumin. On the contrary HA-Sepharose retained primary Igs (IgA, IgG, and 53 % of IgM) as revealed by sodium dodecyl sulphate 10 % polyacrylamide gel electrophoresis (SDS-PAGE), quantitative radial immunodiffusion and immunodetection. At a flow rate of 1 mL/min, the HA-Sepharose column capacity (3.9 mg/mL of matrix) was similar to the reported capacity for the commercial thiophilic T-gel. However, HA-Sepharose showed higher recovery of IgA and IgM than the T-gel in the same salt conditions, clearly an advantage in terms of immunoglobulin recovery strategies. Acetylation changed the matrix adsorption from albumin to immunoglobulins; thus, the highly acetylated gel rendered recoveries of Igs from unprocessed porcine serum practically free of albumin.