Affinity capture of a biotinylated retrovirus on macroporous monolithic adsorbents: towards a rapid single-step purification process

Biotechnol Bioeng. 2005 Mar 30;89(7):783-7. doi: 10.1002/bit.20382.

Abstract

A streptavidin derivitised macroporous monolith was developed to enable single-step capture of chemically biotinylated Moloney Murine Leukaemia Virus (MoMuLV) from crude, unclarified cell culture supernatant. Monoliths were prepared by aqueous cryopolymerisation of acrylamide with N,N''-methylene-bis (acrylamide) and glycidyl methacrylate (Arvidsson et al. [2003] J Chrom A 986:275-290). Streptavidin was immobilised to the epoxy functionalised monoliths. Particulate-containing cell culture supernatant was passed through the monolith without preclarification of the feedstock and adsorption capacities of 2 x 10(5) cfu/ml of adsorbent were demonstrated (cf. Fractogel streptavidin, at 3.9 x 10(5) cfu/ml of adsorbent). The specific titre of the recovered fraction was increased by 425-fold; however, recoveries of less than 8% were achieved. Adsorption of nonbiotinylated MoMuLV on the streptavidin-coated monolith was not observed.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acrylamides / chemistry
  • Adsorption
  • Animals
  • Biotinylation*
  • Chromatography, Affinity / methods*
  • Culture Media
  • Feasibility Studies
  • HeLa Cells
  • Humans
  • Methacrylates / chemistry
  • Mice
  • Moloney murine leukemia virus / isolation & purification
  • Moloney murine leukemia virus / metabolism
  • NIH 3T3 Cells
  • Nanostructures
  • Polymers
  • Retroviridae / isolation & purification*
  • Retroviridae / metabolism*
  • Streptavidin / metabolism
  • Water / chemistry

Substances

  • Acrylamides
  • Culture Media
  • Methacrylates
  • Polymers
  • Water
  • Streptavidin