Adeno-Associated Virus (AAV) Packaging
Recombinant adeno-associated virus (AAV) is a versatile and popular viral vector tool used for in vitro and in vivo gene delivery. AAVs have emerged as the most effective vehicles for gene therapy due to their ability to transduce a wide variety of mammalian cell types and their non-pathogenicity and low immunogenicity in host organisms.
VectorBuilder offers superior quality AAV packaging services to support your AAV-based gene therapy experiments. We have developed a series of proprietary technologies and reagents that have greatly improved recombinant AAV production protocols in terms of titer, purity, viability and consistency, especially for the AAV vector systems used in our vector cloning services. As a result, we have a growing base of highly satisfied customers who come back to us time after time for their AAV vector cloning and AAV packaging needs.
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How do I obtain AAV packaging services?
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Types of AAV offered
We offer second-generation recombinant AAV packaging for the serotypes* listed below. We can package both ssAAV (single-stranded AAV) and scAAV (self-complementary AAV).
Serotypes 1, 2, 3, 4, 5, 6, 6.2, 7, 8, 9, rh10, DJ, DJ/8, PHP.eB, PHP.S, AAV2-retro, AAV2-QuadYF and AAV2.7m8. The tables below list different AAV serotypes and their tissue tropism.
| Serotype | Tissue tropism |
|---|---|
| AAV1 | Smooth muscle, CNS, lung, retina, inner ear, pancreas, heart, liver |
| AAV2 | Smooth muscle, CNS, liver, kidney, retina, inner ear |
| AAV3 | Smooth muscle, liver, lung |
| AAV4 | CNS, retina, lung, kidney |
| AAV5 | Smooth muscle, CNS, lung, retina |
| AAV6 | Smooth muscle, heart, lung, adipose, liver |
| AAV6.2 | Lung, liver, inner ear |
| AAV7 | Smooth muscle, retina, CNS, liver |
| AAV8 | Smooth muscle, CNS, retina, inner ear, liver, pancreas, heart, kidney, adipose |
| AAV9 | Smooth muscle, lung, liver, heart, pancreas, CNS, retina, inner ear, testes, kidney |
| AAVrh10 | Smooth muscle, lung, liver, heart, pancreas, CNS, retina, kidney |
| AAV-DJ | Liver, heart, kidney, spleen |
| AAV-DJ/8 | Liver, brain, spleen |
| AAV-PHP.eB | CNS |
| AAV-PHP.S | PNS |
| AAV2-retro | Spinal nerves |
| AAV2-QuadYF | Endothelial cell |
| AAV2.7m8 | Retina, inner ear |
| Tissue type | Recommended AAV serotypes |
|---|---|
| Smooth muscle | AAV1, AAV2, AAV3, AAV5, AAV6, AAV7, AAV8, AAV9, AAVrh10 |
| CNS | AAV1, AAV2, AAV4, AAV5, AAV7, AAV8, AAV9, AAVrh10, AAV-PHP.eB |
| PNS | AAV-PHP.S |
| Brain | AAV1, AAV2, AAV5, AAV7, AAV8, AAV-DJ/8 |
| Retina | AAV1, AAV2, AAV4, AAV5, AAV7, AAV8, AAV9, AAVrh10, AAV2.7m8 |
| Inner ear | AAV1, AAV2, AAV6.2, AAV8, AAV9, AAV2.7m8 |
| Lung | AAV1, AAV3, AAV4, AAV5, AAV6, AAV6.2, AAV9, AAVrh10 |
| Liver | AAV1, AAV2, AAV3, AAV6, AAV6.2, AAV7, AAV8, AAV9, AAVrh10, AAV-DJ, AAV-DJ/8 |
| Pancreas | AAV1, AAV2, AAV6, AAV8, AAV9, AAVrh10 |
| Heart | AAV1,AAV4, AAV5, AAV6, AAV8, AAV9, AAVrh10, AAV-DJ |
| Kidney | AAV2, AAV4, AAV8, AAV9, AAVrh10, AAV-DJ, AAV-DJ/8 |
| Adipose | AAV6, AAV8, AAV9 |
| Testes | AAV2, AAV9 |
| Spleen | AAV-DJ, AAV-DJ/8 |
| Spinal nerves | AAV2-retro |
| Endothelial cells | AAV2-QuadYF |
* Please note that the ITRs carrying your gene of interest (GOI) is from the AAV2 genome. Different serotypes are distinguished by the capsid protein serotype.
Price and turnaround
Due to our technological advantage, our AAV packaging services have the highest quality in the industry and are also faster and more affordable than those from other suppliers. Prices and turnaround times for different scales of AAV packaging are shown below:
| Scale | Application | Typical Titer* | Minimum Titer** | Volume | Price (USD) | Turnaround*** |
|---|---|---|---|---|---|---|
| Pilot | Cell culture | >1012 GC/ml | >2x1011 GC/ml | 250 ul (10x25 ul), PBS buffer | $399 | 10-20 days |
| Medium | Cell culture | >1012 GC/ml | >2x1011 GC/ml | 1ml (10x100 ul), PBS buffer | $549 | 10-20 days |
| Large | Cell culture | >5x1012 GC/ml | >2x1012 GC/ml | 1ml (10x100 ul), PBS buffer | $999 | 10-20 days |
| Ultra-purified pilot | Cell culture & in vivo | >2x1013 GC/ml | >1013 GC/ml | 100 ul (4x25 ul), PBS buffer | $1149 | 20-30 days |
| Ultra-purified medium | Cell culture & in vivo | >2x1013 GC/ml | >1013 GC/ml | 500 ul (10x50 ul), PBS buffer | $1649 | 20-30 days |
| Ultra-purified large | Cell culture & in vivo | >2x1013 GC/ml | >1013 GC/ml | 1 ml (10x100 ul), PBS buffer | $2549 | 20-30 days |
GC = Genome copy
* Typical titer is achieved in >70% of projects.
** Minimum titer is guaranteed. It applies to vectors for which the region being packaged into virus (from 5’ ITR to 3’ ITR) is below the AAV cargo limit (4.7 kb). For sizes above this limit, the region packaged into the virus may be truncated and result in loss-of-function. Additionally, we may not be able to guarantee titer in the following situations:
- Serotypes 6 and DJ/8 which can only achieve 30-50% of the titer of the other serotypes.
- Vectors containing sequences that could adversely affect the packaging process such as toxic genes (e.g. proapoptotic genes), genes that compromise the integrity of packaging cells or virus (e.g. membrane proteins that cause cell aggregation), and sequences prone to rearrangements or secondary structures (e.g. repetitive or highly GC-rich sequences).
- Customer-supplied transfer plasmid containing undisclosed sequences or atypical ITRs that may introduce uncertainties to packaging efficiency.
- Customer-supplied rep-cap plasmid and/or helper plasmid.
*** Estimated turnaround is the time from production initiation to completion. It does not include waiting time for any customer-supplied materials (e.g. template DNA or viral vectors), QC of such materials, and transit time for shipping final deliverables to the customer.
Deliverables
If you choose pilot, medium or large scale AAV packaging, each order includes the following deliverables:
- Aliquots of custom AAV at selected scale
- Free: Aliquots of control virus*
If you choose ultra-purified pilot or ultra-purified medium scale AAV packaging (in vivo grade), each order includes the following deliverables:
- Aliquots of custom AAV at selected scale
- "Add-on"purchase (optional): Aliquots of ultra-purified control virus*
* The control AAV vector is designed to match the biological application of the custom virus and to be used for testing AAV transduction. For example, if the custom virus overexpresses a gene, then the control virus provided will be EGFP control AAV (AAV overexpressing EGFP), and if the custom virus expresses an shRNA against a gene, then the control virus provided will express a scramble shRNA. Detailed information for the various control viruses is shown below:
| Vector System | Control Vector Name | Control Vector ID | Custom Virus Packaging Scale | Corresponding Control Virus Amount | Price (USD) |
| AAV gene expression system | pAAV[Exp]-CMV>EGFP:WPRE | VB190926-1395dab | Pilot | >2x1011 GC/ml, 100 ul in PBS | Free |
| Medium | >2x1011 GC/ml, 2x100 ul in PBS | Free | |||
| Large | >2x1011 GC/ml, 2x100 ul in PBS | Free | |||
| Ultra-purified pilot | >1013 GC/ml, 4x25 ul in PBS | $390 | |||
| Ultra-purified medium | >1013 GC/ml, 10x50 ul in PBS | $550 | |||
| Ultra-purified large | >1013 GC/ml, 10x100 ul in PBS | $850 | |||
| AAV U6-based shRNA knockdown system | pAAV[shRNA]-EGFP-U6> Scramble_shRNA |
VB180117-1020znr | Pilot | >2x1011 GC/ml, 100 ul in PBS | Free |
| Medium | >2x1011 GC/ml, 2x100 ul in PBS | Free | |||
| Large | >2x1011 GC/ml, 2x100 ul in PBS | Free | |||
| Ultra-purified pilot | >1013 GC/ml, 4x25 ul in PBS | $390 | |||
| Ultra-purified medium | >1013 GC/ml, 10x50 ul in PBS | $550 | |||
| Ultra-purified large | >1013 GC/ml, 10x100 ul in PBS | $850 | |||
| AAV miR30-based shRNA knockdown system | pAAV[miR30]-CMV>EGFP:Scramble_miR30-shRNA:WPRE | VB190813-1134mnj | Pilot | >2x1011 GC/ml, 100 ul in PBS | Free |
| Medium | >2x1011 GC/ml, 2x100 ul in PBS | Free | |||
| Large | >2x1011 GC/ml, 2x100 ul in PBS | Free | |||
| Ultra-purified pilot | >1013 GC/ml, 4x25 ul in PBS | $390 | |||
| Ultra-purified medium | >1013 GC/ml, 10x50 ul in PBS | $550 | |||
| Ultra-purified large | >1013 GC/ml, 10x100 ul in PBS | $850 | |||
| AAV CRISPR system | pAAV[Exp]-CMV>EGFP:WPRE | VB190926-1395dab | Pilot | >2x1011 GC/ml, 100 ul in PBS | Free |
| Medium | >2x1011 GC/ml, 2x100 ul in PBS | Free | |||
| Large | >2x1011 GC/ml, 2x100 ul in PBS | Free | |||
| Ultra-purified pilot | >1013 GC/ml, 4x25 ul in PBS | $390 | |||
| Ultra-purified medium | >1013 GC/ml, 10x50 ul in PBS | $550 | |||
| Ultra-purified large | >1013 GC/ml, 10x100 ul in PBS | $850 |
Production, purification and titration
For our recombinant AAV manufacturing, the transfer plasmid carrying the gene of interest (GOI) is co-transfected with our proprietary Rep-cap plasmid and helper plasmid encoding adenovirus genes (E4, E2A and VA) that mediate AAV replication into packaging cells. After a short incubation period, viral particles are harvested from cell lysates and concentrated by PEG precipitation. For ultra-purified AAV (in vivo grade), viral particles are further purified and concentrated by cesium chloride (CsCl) gradient ultracentrifugation. We use a qPCR-based approach to measure AAV titer.
Read more about our titration method
Quality control
For each AAV produced by VectorBuilder, quality control includes titer measurement, sterility testing for bacteria and fungi, and mycoplasma detection. If the transfer vector encodes a fluorescent protein, we would transduce cells with virus and detect corresponding fluorescence. Additionally, for ultra-purified AAV, we routinely sample virus quality by SDS-PAGE analysis and endotoxin assay.
Customer-supplied vectors
If customer-supplied AAV plasmids are used, please send them to us following the Materials Submission Guidelines. Please strictly follow our guidelines to set up shipment to avoid any delay or damage of materials. All customer-supplied materials undergo mandatory QC by VectorBuilder which may incur a $100 surcharge for each item. Please note that production may not be initiated until customer-supplied materials pass QC.
Questions
Please refer to our Vector FAQ for commonly asked technical questions. To get additional information about what we can offer, please contact VectorBuilder's service team following the contact information here.
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