Production and Purification of a Human Dll1(ECD)IgGFc Fusion Protein in CHOSFS Cells
Notch signalling regulates arterial differentiation during embryonic development. The Notch ligand Delta-like 1 (Dll1) is known to be an essential regulator of postnatal arteriosclerosis (Limbourg et al. 2007). Clarification of its role within cardiac repair may lead to new ways of improving cardiovascular regeneration through targeted manipulation of cardiovascular differentiation. Since the soluble form leads to inactivation (Nichols et al. 2007), a fusion protein of the extracellular domain of the Notch ligand Delta-like 1 and the Fc region of IgG has been selected to be recombinantly produced in mammalian cells. CHOSFS cells were used as the expression system in this work. They were chemically transfected and cultured in serum-free medium under selective conditions. High-producing clones were gained by single-cell cloning via FACS and via dilution method. For FPLC-purification protein G as well as protein A columns were used. The activity was verified using a luciferase assay utilizing human umbilical cord vein endothelial cells (HUVEC).
KeywordsCell Culture Supernatant Notch Ligand Spinner Flask Single Cell Cloning Flow Cytometry Measurement
This work was performed within the activities of the JRG “large scale cultivation” in the DFG cluster of excellence “Rebirth” (from regenerative biology to reconstructive therapy).