An immunomagnetic cell separation system based on a retroviral vector containing a chimeric, recombinant human-murine CD4 gene

摘要

Primary B- and T-lymphocytes and neoplastic cells of B- and T-cell origin are particularly hard to transfect with plasmids. Thus, functional and molecular studies in immunology and oncohematology require selection of cells after plasmid transduction, assuring obtaining homogeneous cell populations expressing plasmid-delivered molecules. The selection is usually achieved by incorporation of an antibiotic resistance gene or fluorescent protein into a plasmid and subsequent antibiotic selection or fluorescence-activated cell sorting (FACS). However, these cell sorting methods have significant drawbacks that limit their utilization. An ideal system should be fast, cheap and ensure efficient selection of transfected cells. In an attempt to deliver a sorting system meeting these requirements, we have generated a plasmid for retroviral gene delivery and expression, containing a recombinant human-murine CD4 gene, with a truncated intracellular domain to prevent undesired signaling events. After retroviral infection, cells expressing hmCD4 are selected by immunomagnetic anti-mCD4 beads. The system offers a very robust, efficient, fast and cost-saving cell separation solution. The vector is available for the academic community upon request.