The AAVanced™ Concentration Reagent enables one-step rAAV particle isolation from packaging cell media. Specifically optimised for the precipitation of most AAV serotypes from culture medium and based on an established nanoparticle technology, this reagent significantly reduces the complexity and time required for rAAV virus production. rAAV particles produced using this reagent have been tested successfully for multiple serotypes of AAV with no observed cytotoxic effects making it ideal for challenging in vitro and in vivo applications.
Benefits of using AAVanced™ Concentration Reagent
• Easy to use, single reagent
• One step purification
• Isolate rAAV directly from media
• No cell lysates or centrifugation
• Saves time and cost-effective
• Compatible with all serotypes
Easiest protocol to isolate rAAV particles
Advantage 1: AAVanced™ Concentration Reagent produces robust virions
Recombinant rAAV was packaged with a PGK-GFP expression shuttle vector. The rAAV particles were isolated either by the traditional cell lysis freeze-thaw with centifugation method or with the AAVanced concentration reagent from the packaging cell media. Equal amounts of rAAV were then added to HEK293 cells to test for infectivity based on GFP expression. Representative images of GFP expressing cells 10 days after infection.
Advantage 2: AAVanced™ rAAV isolation works with multiple serotypes in vitro
Recombinant AAV was packaged in parallel with three different serotypes: AAV-2, AAV-DJ and AAV-5. The exact same shuttle rAAV vector with a PGK promoter expressing GFP was used for all tests. The media was collected after 48 hours and the rAAV particles in the media were isolated using the AAVanced Concentration Reagent. Each rAAV particle pellet was resuspended in 100ml sterile PBS and then 1ml of the rAAV suspensions were added to three different cell lines (HEK293T, HT1080 and CHO) to validate transduction efficiencies. The infected cells were imaged for GFP expression six days after addition of the isolated rAAV particles
Advantage 3: AAVanced™ rAAV particles work in vivo
Shown in the upper right panels are representative images of hippocampal and midbrain sections from six week old C57 mice injected with AAV virus (ITR-PGK-GFP-ITR) concentrated using AAVanced Concentration Reagent. Approximately 1.5 ul of concentrated AAV virus (AAV-2 serotype) was delivered to hippocampal and midbrain regions by stereotaxic injection. Three weeks after virus injection, the animals were perfused with paraformaldehyde, fixed, and brains specimen were sectioned to 40 micron slices before visualization for GFP fluorescence.
Shown in the lower right panels are representative images of sections of mouse CA3 neurons in hippocampus from two month old C57 mice injected with AAV-5 virus (ITR-PGK-GFP-ITR) concentrated using AAVanced Concentration Reagent. Approximately 1.5 ul of concentrated rAAV-5 virus was delivered to cortical regions by stereotaxic injection. Two weeks after virus injection, animals were perfused by paraformaldehyde, fixed and brains were sectioned to 70 micron slices before visualization for GFP fluorescence.
Material available for download
AAVanced™ Concentration Reagent manual
Note: product availability depends on country - see product detail page.
|Details||Cat number & supplier||Size||Price|
|AAVancedTM Concentration Reagent (100 ml aliquot) AAV100A-1 · System Biosciences||
|AAVancedTM Concentration Reagent (250 ml aliquot) AAV110A-1 · System Biosciences||