The critical growth factor density required to support neural lineage generation from mouse embryonic stem cells is assessed by constructing a surface density gradient of immobilized nerve growth factor (NGF) from a plasma polymer film base. A chemical surface gradient varying from high hydroxyl group density to high aldehyde group density is prepared through diffusion-controlled plasma polymerization of two monomers (ethanol and propionaldehyde) under a moving mask. NGF density gradients are then produced by reductive amination with the aldehyde groups on the plasma polymer surface. Mouse embryoid body derived (mEB) cell differentiation on the gradient surface is evaluated by immunofluorescence staining against Nestin. mEB cell density and the percentage of Nestin-positive cells increase with increasing NGF density up to a critical value corresponding to 52.9 ng cm-2, above which cell attachment and differentiation do not increase further. This gradient-based screening approach allows the growth factor surface densities to be optimized for biomaterials intended for cell differentiation or expansion, which is highly relevant to creating efficient manufacture processes for cell therapies.
- embryonic stem cells
- high throughput screening
- nerve growth factor gradients
- neural differentiation