OHSU

Recommended Protocols

OHSU Biosafety Manual

 

Screening for Replication Competent Virus

Recombination events or contamination from wildtype virus can result in the presence of replication competent virus (RCV) in a population of replication deficient viral vectors. The Institutional Biosafety Committee (IBC) recommends testing for the presence of RCV in viral stocks to be used for in vivo experiments (also see vector chart).

Testing methods must be communicated to the IBC upon review of proposals involving the in vivo use of viral vectors.

Adenoviral Vectors

The IBC's preferred method of testing for the presence of Replication Competent Adenovirus (RCA) is by plaque assay. A protocol for plaque purification and testing for RCA on non-E1 expressing cells is available here: Plaque Assay. A PCR protocol is also available for follow-up testing: Ad PCR. Also see Zhu, et al, Human Gene Therapy 10:113-121 (1999) for additional PCR primers and a discussion on this topic (available online from the OHSU Library).

Retroviral Vectors

The IBC preferred protocol for testing for the presence of RCV for murine retroviral vectors is available here: RCR.

For detecting RCV in lentiviral preps for 3rd generation lentiviral vectors, see Dull et al, "A Third Generation Lentivirus Vector with a Conditional Packaging System." J. Viro., 1998. p8463-8471 (available online from the OHSU Library).