High-level β-globin expression and preferred intragenic integration after lentiviral transduction of human cord blood stem cells
J. Clin. Invest. Suzan Imren, et al. 114:953 doi:10.1172/JCI21838 [Go to this article.]

Figure 3
Lineage-specific expression of human β^A-T87Q -globin protein in human erythroid cells generated in vitro. (A) Photomicrograph of Wright-Giemsa_stained slide preparation of cells from erythroid suspension cultures initiated 14 days previously with cells obtained immediately after exposure to lentivirus. (B) FACS analysis of the cells shown in A after staining with antibodies specific to human glycophorin-A. (C) Representative HPLC profile of a cell lysate obtained from a 14-day culture of cord blood cells set up immediately after infection. Peaks representing differently eluting β-globin proteins are indicated. (D) Isolation by FACS of human CD45/71⁺ cells from bone marrow (BM) aspirates of transplanted mice to initiate CFC or suspension cultures that support terminal human erythroid cell differentiation. (E) Representative HPLC profile of a cell lysate from a 14-day suspension culture of human erythroid cells initiated with cells obtained from a mouse that received a transplant of transduced cord blood cells 11 weeks previously. (F) RT-PCR analysis of RNA extracted from cells shown in A (right panels) and human CD19/20⁺ B-lymphoid cells isolated by FACS (left panels) from a marrow sample of a repopulated mouse 19 weeks after transplant. The β^A-T87Q-globin transcript was detected in erythroblasts even when diluted 32 times (top right panel), whereas no signal was seen in the extract of B-lymphoid cells (top left panel). Triangles indicate decreasing concentration.