Immobilization of growth factors on solid supports for the modulation of stem cell fate.

Surface- and matrix-bound signals modulate stem cell fate in vivo and in vitro. This protocol enables the immobilization of a wide range of biomolecules that contain primary amino groups to different types of solid carriers, including glass substrates and standard polystyrene well plates. We describe how thin polymer coatings of poly(octadecene-alt-maleic anhydride) can be used to covalently attach growth factors directly, or through poly(ethylene glycol) spacers, to solid supports at defined concentrations. Surface-immobilized growth factors can be presented over a wide range of concentrations (5-150 ng cm(-2)), as we have previously shown for leukemia inhibitory factor and stem cell factor. Cell activation can be achieved in the presence of adhesion-promoting extracellular matrix proteins. Depending on the methods used, the overall procedure takes 1.5-3 d. In general, the approach can be used to investigate the effect of defined amounts of immobilized growth factors on stem cells and on the maintenance, growth and differentiation of other cell types.

Functional immobilization of signaling proteins enables control of stem cell fate.

The mode of ligand presentation has a fundamental role in organizing cell fate throughout development. We report a rapid and simple approach for immobilizing signaling ligands to maleic anhydride copolymer thin-film coatings, enabling stable signaling ligand presentation at interfaces at defined concentrations. We demonstrate the utility of this platform technology using leukemia inhibitory factor (LIF) and stem cell factor (SCF). Immobilized LIF supported mouse embryonic stem cell (mESC) pluripotency for at least 2 weeks in the absence of added diffusible LIF. Immobilized LIF activated signal transducer and activator of transcription 3 (STAT3) and mitogen-activated protein kinase (MAPK) signaling in a dose-dependent manner. The introduced method allows for the robust investigation of cell fate responses from interface-immobilized ligands.