Use of a modified zirconia support in the separation of immunoproteins

Anuradha Subramanian, Sabyasachi Sarkar

Research output: Contribution to journalArticlepeer-review

20 Scopus citations

Abstract

Zirconia beads (25-38 μm in diameter) were modified with N,N,N′,N′-ethylenediaminetetramethylenephosphonic acid to generate a zirconia based pseudoaffinity support, further referred to as r_PEZ. The influence of pH, salt concentration and temperature on the binding of human immunoglobulin G (hIgG) to r_PEZ was studied. Temperature had no significant impact on the maximum binding capacity (Qmax), and the equilibrium-binding constant (Kd), whereas pH and the salt concentration had a noticeable impact on both Qmax and Kd. The Qmax value of 55 mg hIgG/ml of bead was obtained at a pH of 5.5 and found to decrease with an increase of pH. The modified zirconia support allowed the separation of immunoglobulins (IgG, IgA and IgM) from untreated human serum. Elution was possible under mild conditions with a step salt gradient. Overall protein recoveries in the range of 109-125% were obtained with human serum. Human IgG, human IgA, and human IgM yields of 29.50±6.3, 3.22±0.7, and 6.84±0.7%, respectively, were obtained at a linear velocity of 4.32 cm/min. Purity of products, obtained from a single chromatographic step was estimated to be greater than 89.0±2.6%. The utility of r_PEZ in the selective removal of immunoglobulins, as in immunoadsorption was discussed.

Original languageEnglish (US)
Pages (from-to)179-187
Number of pages9
JournalJournal of Chromatography A
Volume944
Issue number1-2
DOIs
StatePublished - Jan 25 2002

Keywords

  • Albumin
  • Immunoglobulins
  • Proteins
  • Pseudoaffinity separations
  • Stationary phases, LC
  • Zirconia

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Organic Chemistry

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