Specific affinity depletion of cell adhesion molecules and growth factors from serum

Serum is a common component of most in vitro cell culture media, particular ly of primary cells. Studies of cellular responses to particular adhesion m olecules or growth factors are often confounded by the presence of these mo lecules in the serum supplement. We describe a combined affinity protocol f or removing vitronectin and fibronectin from serum. This protocol can also be used to purify these molecules. We also describe the removal of growth-p romoting elements using heparin-Sepharose. As vitronectin and fibronectin e ach bind to heparin, these molecules are removed first and the heparin-Seph arose depletion occurs last in the sequence. This protocol provides a detai led step-by-step guide to achieve quantitative depletion of serum in an opt imised format, with additional information on pitfalls and problems. It sho uld be of use to people who wish to accurately determine the relationship b etween cells, extracellular matrix molecules and growth factors. (C) 2001 E lsevier Science B.V. All rights reserved.