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

Gabriela Ramos-Clamont, Maria del Carmen Candia-Plata, Roberto Guzman Zamudio, Luz Vazquez-Moreno

Research output: Contribution to journalArticle

12 Scopus citations

Abstract

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.

Original languageEnglish (US)
Pages (from-to)28-34
Number of pages7
JournalJournal of Chromatography A
Volume1122
Issue number1-2
DOIs
StatePublished - Jul 28 2006

Keywords

  • Blood derivatives
  • Highly acetylated agarose
  • Hydrophobic interaction chromatography
  • Porcine immunoglobulins
  • T-gel

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Organic Chemistry

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