Addgene’s AAV is distributed as 100 µL of viral preparation at a titers of 2e12 - 2e13 vg/mL. Minimum titers are reported on each items' material page. Actual titers are reported with each shipment.
The price for each 100 µL aliquot is $350 plus shipping. As a non-profit organization, we aim to provide AAV preparations to the research community not to drive a profit, but largely to recover our production costs. To ensure that researchers receive high quality viral preparations, our viral production includes substantial quality control steps performed in-house at Addgene. These quality control steps are described below, and are performed on each lot of virus that we distribute. Quality control data for each lot of virus is available from Addgene upon request.
All titering is performed on AAV preparations that have been stored at -80°C and thawed. This ensures that any loss of titer associated with the recipient's initial thaw will be accounted for in our reported titers.
AAV particles are titered by real-time qPCR or by digital droplet PCR. Titers are reported as the average of at least two independent measurements. Titer values are determined by comparison to a standard curve of a plasmid sample of known concentration. In some cases, the qPCR assay and corresponding titer determinations are also validated using AAV Reference Material (ATCC).
Confirmation of Transfer Plasmid
The final AAV preparation undergoes PCR with primers targeting the transfer plasmid used in the initial transfection. Most of our primers are targeting the promoter region and/or the transgene. For DIO or FLEX plasmids, we also validate the orientation of the cassette using 2 or more primer pairs. PCR products are analyzed by gel electrophoresis to ensure the correct banding pattern and sizes.
When possible, the final viral preparation is analyzed by next-generation sequencing. Sequencing results are then analyzed for the presence of the viral genome, as well as for the absence of any contaminants. If/when next-generation sequencing results are obtained, the results are posted on the material page for each viral catalog item for which this validation was performed.
Purity of AAV preparations is assayed by comparing the relative stoichiometric ratios of the viral capsid proteins VP1, VP2 and VP3. Samples of viral preparations are subjected to polyacrylamide gel electrophoresis (PAGE) followed by silver staining and the molecular weight and relative intensity of the viral capsid proteins are analyzed. The abundance of viral capsid proteins as a fraction of total protein present in the sample is also determined and used to determine purity of the AAV preparation.
Whenever possible, viruses are tested in vitro for reporter gene expression to validate transduction. Fluorescence microscope images showing reporter or transgene expression are posted on the material page for each viral catalog item for which this validation was performed.
A note about MTAs and virus associated DNA that is required with viral service:
Each viral service request will also include virus associated DNA ($30), which is a sample of the purified DNA from the original plasmid that was used to make the virus. In order for Addgene to provide viral services to you, we must ensure that you have the right to use the original plasmid. Distribution of the original plasmid is important because it is the actual material being referenced in the Material Transfer Agreement (MTA) and its inclusion is what gives you permission to use the plasmid. For each unique plasmid, only a single order of virus associated DNA is required when requesting a viral preparation. In other words, if you request two or more viral preparations of the same plasmid, only one order of virus associated DNA (and a single $30 charge) would be required.