Background
The development of optimized cultivation strategies for mesenchymal stem cells (MSC) is one major field in the area of tissue engineering and cell based therapies. The physiological tissue niche of stem cells is characterized by a complex set of structural, physical and chemical cues, which create a complex three-dimensional dynamic microenvironment. Mimicking these conditions in vitro is a promising strategy for expansion and differentiation of MSC. This also includes the application of physiological oxygen concentrations (e.g. 3-8 % for most tissues).Multiple cell biological effects of varying oxygen supply have been reported [1]. However, a lot of current research work is dedicated to the analysis of selected biomarkers and distinct signaling pathways, which carries the risk of overlooking unexpected biological effects. A more systemic approach involves the investigation of the entirety of proteins secreted by the cells - the secretome. However, secretome analysis is very challenging for most current cell cultures, because serum or other undefined protein-rich medium supplements are commonly used and create a strong background of high-abundance proteins in the samples. In order to exploit the potential of secretome analysis for MSC cultures a methodological toolbox was developed that provides access to the rather low abundant secretome in serum containing media.