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Reference standards for accurate validation and optimization of assays that determine integrated Lentiviral vector copy number in transduced cells

Published

Author(s)

Barbara S. Paugh, Lajos Baranyi, Andre Roy, Hua-Jun He, Lindsay Harris, Kenneth D. Cole, Caroline Raimund, Patricia S. Langan, Moria Artlip, Srikanta Jana, Rimas J. Orentas, Sheng Lin-Gibson, Winfried Krueger, Boro Dropuli?

Abstract

Lentiviral vectors (LV) have emerged as a robust technology for therapeutic gene delivery into human cells as advanced medicinal products. As these products are increasingly commercialized, there are concomitant demands for their characterization to ensure safety, efficacy and consistency. Assays and standards are essential for accurately measuring parameters for such product characterization. A critical parameter is the vector copy number (VCN) which measures the genetic dose of a transgene present in gene-modified cells. Here we describe a set of clonal Jurkat cell lines with defined copy numbers of a reference lentiviral vector integrated into their genomes. Genomic DNA was characterized for copy number, genomic integrity and integration coordinates and showed uniform performance across independent quantitative PCR assays and stability during continuous long-term culture, demonstrating sustained renewability of the reference standard material. DNA from the Jurkat VCN standards would be useful for control of quantitative PCR assays for VCN determination in LV gene-modified cellular products and clinical samples.
Citation
Scientific Reports
Volume
11

Keywords

Lentiviral vectors (LV), Vector copy number (VCN), Gene therapy, Reference Materials, Quantitative PCR, Digital PCR, Lentiviral integration site, Jurkat cell.

Citation

Paugh, B. , Baranyi, L. , Roy, A. , He, H. , Harris, L. , Cole, K. , Raimund, C. , Langan, P. , Artlip, M. , Jana, S. , Orentas, R. , Lin-Gibson, S. , Krueger, W. and Dropuli?, B. (2021), Reference standards for accurate validation and optimization of assays that determine integrated Lentiviral vector copy number in transduced cells, Scientific Reports, [online], https://doi.org/10.1038/s41598-020-79698-w (Accessed October 3, 2024)

Issues

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Created July 11, 2021, Updated October 31, 2021