Development of cell-based assay for detecting replication-competent adeno-associated virus by qPCR.

ABSTRACT: Recombinant, non-replicative adeno-associated virus (AAV) containing a therapeutic gene of interest (GOI) is widely used as a vector for gene therapy. One way to manufacture AAV is through triple-transfection of HEK293 cells, with plasmids containing the (1) GOI, (2) replication (rep2) and capsid (cap) sequences, and (3) adenovirus helper sequences. During the manufacturing of AAV, replication-competent AAV (rcAAV) could theoretically be generated via homologous and non-homologous recombination events. rcAAV contaminants could lead to reduced efficacy, or an adverse immunogenic response. Therefore, testing is required by regulatory agencies. However, there is a paucity of literature on this critical assay. Here, we have developed a sensitive, cell-based assay for detection of

SUBMITTER: Shevade A 

PROVIDER: S-EPMC12329522 | biostudies-literature | 2025 Sep

REPOSITORIES: biostudies-literature