lentivirus titering

How to titer retrovirus/lentivirus

A number of methods have been developed for determining the lentiviral/retroviral titers such as using ELISA to assess the amount of p24 antigen, enzyme assay to quantify reverse transcriptase activity, and real time RT-PCR to count viral RNA genome. However, these methods do not differentiate non-infectious from infectious virus particles that are present in the virus preparation; therefore, these measurements rely on some correlation factors to derive a titer, which can be inaccurate.

Because an accurate virus titer is critical for most experimental purposes, Allele Biotech’ pre-packaged viruses are measured by the number of genomic integration events to determine the infectious titer. This functional titering is done by first transducing TE671cells (selected for consistent transduction efficiency) followed by measuring the viral genome integrated into the cell genome with QPCR. The number of cellular genome copies is also measured, and the copy number of viruses is divided by that of cells to obtain active virus titer.
How to titer lentivirus retrovirus

A plot of quantitative PCR (Q-PCR) titering of virus preparation is provided above.
TE671 cells (1×105) were transduced with 2×10-3ml of a virus sample to be titered. Cells were harvested and DNA extracted for QPCR. The titer is determined as followed, transduction unit per ml (TU/ml)=N [X1/(X2/2)]/2×10-3ml, where N is the cell number, X1 is the copy number of integrated virus DNA, and X2 is the copy number of GAPDH DNA.