Cytotechnology

, Volume 19, Issue 1, pp 63–72 | Cite as

Separation of growth-stimulating activity of BSA fraction V from the bulk of albumin using Heparin Sepharose Chromatography

  • Joanne Keenan
  • Gerald Doherty
  • Martin Clynes
Article

Abstract

Bovine serum albumin (BSA) is a potential source of biological contamination in cell culture medium. The aim of this work was to attempt to replace BSA in low serum and serum-free medium (SFM). BSA fraction V was subjected to a variety of processes in order to determine if the growth promoting activity observed for NRK cells could be extracted from the BSA molecule. These included solvent extractions, diafiltration, reverse phase HPLC and affinity chromatography using heparin sepharose. Solvent extraction and diafiltration failed to remove the activity from the BSA. Affinity chromatography using heparin sepharose indicated that all of the activity observed with BSA was retained in the 0.5 M NaCl fraction and was associated with less than 3% of the original protein. The major protein band in the 0.5 M NaCl fraction had the same apparent molecular weight as albumin (as seen by SDS-PAGE and analytical reverse phase HPLC). Unlike the untreated BSA, the 0.5 M NaCl fraction was partially susceptible to proteolytic digestion and to variations in pH.

Key words

BSA fraction V bovine serum albumin NRK heparin sepharose chromatography 

Abbreviations

HS

heparin sepharose

DHS

donor horse serum

SFM

serum free-medium

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Copyright information

© Kluwer Academic Publishers 1996

Authors and Affiliations

  • Joanne Keenan
    • 1
  • Gerald Doherty
    • 1
  • Martin Clynes
    • 1
  1. 1.National Cell and Tissue CentreDublin City UniversityGlasnevinIreland

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