A survey of ex vivo/in vitro transduction efficiency of mammalian primary cells and cell lines with Nine natural adeno-associated virus (AAV1-9) and one engineered adeno-associated virus serotype

Abstract

BackgroundThe ability to deliver a gene of interest into a specific cell type is an essential aspect of biomedical research. Viruses can be a useful tool for this delivery, particularly in difficult to transfect cell types. Adeno-associated virus (AAV) is a useful gene transfer vector because of its ability to mediate efficient gene transduction in numerous dividing and quiescent cell types, without inducing any known pathogenicity. There are now a number of natural for that designed AAV serotypes that each has a differential ability to infect a variety of cell types. Although transduction studies have been completed, the bulk of the studies have been done in vivo, and there has never been a comprehensive study of transduction ex vivo/in vitro.MethodsEach cell type was infected with each serotype at a multiplicity of infection of 100,000 viral genomes/cell and transduction was analyzed by flow cytometry + .ResultsWe found that AAV1 and AAV6 have the greatest ability to transduce a wide range of cell types, however, for particular cell types, there are specific serotypes that provide optimal transduction.ConclusionsIn this work, we describe the transduction efficiency of ten different AAV serotypes in thirty-four different mammalian cell lines and primary cell types. Although these results may not be universal due to numerous factors such as, culture conditions and/ or cell growth rates and cell heterogeneity, these results provide an important and unique resource for investigators who use AAV as an ex vivo gene delivery vector or who work with cells that are difficult to transfect.

Robust Lentiviral Gene Delivery But Limited Transduction Capacity of Commonly Used Adeno-Associated Viral Serotypes in Xenotransplanted Human Skin

“…54 Previous studies using rAAV vectors in vitro for transduction of primary human keratinocytes have reported poor efficacy of the commonly used serotypes, 28,48 although serotypes 1, 2, and 6 may be suitable for this purpose. [25][26][27]55 Using our panel of single-stranded rAAV-CMV-FLuc vectors for in vitro transduction of primary cells of human origin, including keratinocytes, we could partially support earlier claims (Supplementary Fig. S4).…”

“…AAV-mediated gene transfer into murine skin in vivo has been described, 23,24 but primary human keratinocytes seem refractory to transduction in vitro by AAV-based vectors. [25][26][27][28] Reports on using rAAV for gene transfer to human skin in vivo are not currently available. Therefore, we compared the gene transfer capability of a selection of frequently used AAV serotypes (AAV1, 2, 5, 6, 8, or 9) in xenotransplanted human skin by intradermally administering AAV vectors expressing a cytomegalovirus (CMV)-driven firefly luciferase gene.…”

Long-Term Episomal Transgene Expression from Mitotically Stable Integration-Deficient Lentiviral Vectors

“…The in vitro transduction capacity of AAV vectors is generally lower than that of IDLV vectors (Ellis et al, 2013), and here the single-stranded vectors yielded a reasonable 15% and 16% eGFP + cells 5 days after transduction for ssAAV-CG and ssAAV-CGm, respectively (Fig. 3a).…”

Use of Adeno-Associated Virus to Enrich Cardiomyocytes Derived from Human Stem Cells

“…2,3 A comprehensive in vitro survey of AAV transduction efficiency on various mammalian cell types has been conducted. 4 Though previous study has proven the feasibility of AAV to transduce stem cell differentiated cardiomyocytes on a small scale, 5 an extensive optimization of AAV on stem cell-derived cardiomyocytes has not been reported. Here we compared the transduction efficiency of seven commonly used AAV serotypes in low-purity induced pluripotent stem cell (iPSC) differentiated cardiomyocytes, and all tested serotypes demonstrated preferential cardiomyocytes transduction in comparison to noncardiomyocytes, with AAV1 showing the highest cardiac transduction efficiency.…”