Quantitation of AAV in a dual-vector system using SV-AUC.

IF 3.7 3区 医学 Q2 CHEMISTRY, MEDICINAL
Alexander E Yarawsky, Carlo Ciatto, Peter Slade, Natalya Figueroa, John W Burgner, Michael DeLion, Lake N Paul
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引用次数: 0

Abstract

Sedimentation velocity analytical ultracentrifugation (SV-AUC) has become the "gold standard" for characterization of the empty, partial, and full capsids of gene therapy products (e.g., AAV and Adenovirus vectors). Other techniques, such SEC-MALS, TEM, and mass photometry, are commonly used for capsid quantitation, however, the resolving power of these techniques is lacking. In this body of work, SV-AUC was implemented in the characterization of a dual-vector AAV system where the difference in packaged genomes was ∼400 nucleotides. SV-AUC instrument parameters and analysis were optimized to accurately quantitate both AAV vectors with less than 8% error and highly correlated linearity (R2 > 0.99) as compared to ddPCR. The results of this work highlight the resolution and accuracy of dual-vector capsid quantitation by SV-AUC and demonstrate the use of the powerful Bayesian analysis implemented in the SEDFIT analysis software.

使用 SV-AUC 对双载体系统中的 AAV 进行定量。
沉降速度分析超速离心法(SV-AUC)已成为表征基因治疗产品(如 AAV 和腺病毒载体)空、部分和完整囊壳的 "黄金标准"。其他技术,如 SEC-MALS、TEM 和质量光度法,通常也用于囊体定量,但这些技术的分辨能力不足。在这项工作中,SV-AUC 被用于表征双载体 AAV 系统,该系统的包装基因组相差 400 个核苷酸。对 SV-AUC 仪器参数和分析进行了优化,与 ddPCR 相比,SV-AUC 能准确定量两种 AAV 载体,误差小于 8%,线性相关度高(R2 > 0.99)。这项工作的结果凸显了 SV-AUC 定量双载体包囊的分辨率和准确性,并展示了 SEDFIT 分析软件中强大的贝叶斯分析功能的应用。
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来源期刊
CiteScore
7.30
自引率
13.20%
发文量
367
审稿时长
33 days
期刊介绍: The Journal of Pharmaceutical Sciences will publish original research papers, original research notes, invited topical reviews (including Minireviews), and editorial commentary and news. The area of focus shall be concepts in basic pharmaceutical science and such topics as chemical processing of pharmaceuticals, including crystallization, lyophilization, chemical stability of drugs, pharmacokinetics, biopharmaceutics, pharmacodynamics, pro-drug developments, metabolic disposition of bioactive agents, dosage form design, protein-peptide chemistry and biotechnology specifically as these relate to pharmaceutical technology, and targeted drug delivery.
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