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Archive > Volume 25 Issue 4 > 2018,25(4):389-393. DOI:10.3872/j.issn.1007-385X.2018.04.012 Prev Next
Basic Research
In vitro construction and amplification and primary functional analysis of anti- CD19 chimeric antigen receptor (CD19-CAR) modified T cells
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Abstract:
Objective: To establish a chimeric antigen receptor（CAR）modified T cells specifically targeting CD19 molecule (CD19- CAR-T cells) and to testify their in vitro killing effect on target cells. Methods: CD19-CAR fragments yielded by PCR were construct- ed into pCDH-GFP lentiviral vectors by molecular cloning technology. The packaged lentiviral particles were transducted into CD3 + T cells of donors. Transduction efficiency was measured by flow cytometry and PCR. The in vitro cytotoxicity of obtained CD19- CAR-T cells against CD19 + Ramos cells was tested by 7-AAD staining. Results: The amplification folds of CD3 + T cells increased to (78.8± 23.2) folds after in vitro culture for 10 days, and about (58.3±5.4)% cells expressing GFP. About (57.4±9.3)% CD19 + Ramos cells were specifically killed by the CD19-CAR-T cells in vitro at the E∶T ratio of 5∶1. Conclusion: This study successfully established an effective method for constructing and amplifying CD19-CAR-T cells in vitro, which showed profound efficiency and specific cyto- toxity against CD19 + Ramos cells. And this report might provide an experimental evidence for clinical treatment of CD19 + B cell neo- plasmas.
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Project Supported:
Project supported by the Foundation of the Jiangsu Social Development - Clinical Frontier Technology (No. BE2016809), and the Foundation of the Nanjing Science and Technology Development Plan (No. 201503011)
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