Systematic Comparison and Validation of Quantitative Real-Time PCR Methods for the Quantitation of Adeno-Associated Viral Products

Overview

Journal Hum Gene Ther Methods

Publisher Mary Ann Liebert

Specialties Genetics
Pharmacology

Date 2015 May 9

PMID 25953194

Citations 33

Authors

Natalie Jayne Werling

Stifani Satkunanathan

Robin Thorpe

Yuan Zhao

Affiliations

Soon will be listed here.

Abstract

Adeno-associated viral (AAV) vectors show great promise for gene therapy because of their excellent safety profile; however, development of robust dose-determining assays for AAV has presented a significant challenge. With the ultimate goal of future harmonization and standardization of AAV dose determination assays, we systematically analyzed the influence of key variables, including sample preparation procedure, the choice of primers, and real-time quantitative PCR (qPCR) target sequences and calibration DNA conformation on the qPCR quantitation of AAV products. Our results emphasize the importance of designing qPCR primers and conducting sample preparation and demonstrate the need for extensive characterization, vigorous control, and use of reference materials in clinical dose determination.

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