TransIT-AAViator® System Outperforms Competitor Reagents in Suspension AAV Cell Cultures with a GFP transgene: Viral Production 2.0 cells grown in Viral Production Medium were transfected with pAAV-WPRE-GFP (Aldevron), pAAV-Helper (Aldevron) and pAAV-RepCap (AAV9 and AAV8 Genemedi, AAV5 Aldevron) at a 1:1:1 plasmid concentration ratio with the following reagents: TransIT-AAViator® Transfection reagent (1.25:1,vol:wt), Polymer F (1:1) , or Polymer P (1:1). RevIT™-AAV Enhancer (1 µl/mL) was added at the time of transfection. All AAV serotypes were harvested at 72 hours post-transfection using chemical lysis. Genome titers were determined via dPCR using primers and a probe targeting the CMV promoter region. Total assembled capsids were determined using the AAV9, AAV8 or AAV5 Titration ELISA Kits (Progen), and the Percent Full Capsids were determined by dividing the number of genome copies by total assembled capsids for each condition. The error bars represent the standard deviation of duplicate wells.

TransIT-AAViator® System Outperforms Competitor Reagents in Suspension AAV Cell Cultures with an RPE65 transgene: Viral Production 2.0 cells grown in Viral Production Medium were were transfected with pAAV-WPRE-RPE65 (Aldevron), pAAV-Helper (Aldevron) and pAAV-RepCap (AAV9 and AAV8 Genemedi, AAV5 Aldevron) at a 1:1:1 plasmid concentration ratio with the following reagents: TransIT-AAViator® Transfection reagent (1.25:1,vol:wt), Polymer F (1:1) , or Polymer P (1:1). RevIT™-AAV Enhancer (1 µl/mL) was added at the time of transfection. All AAV serotypes were harvested at 72 hours post-transfection using chemical lysis. Genome titers were determined via dPCR using primers and a probe targeting the CMV promoter region. Total assembled capsids were determined using the AAV9, AAV8, or AAV5 Titration ELISA Kits (Progen), and the Percent Full Capsids were determined by dividing the number of genome copies by total assembled capsids for each condition. The error bars represent the standard deviation of duplicate wells.

High AAV Titers Across A Range of Plasmid DNA Doses: Viral Production 2.0 cells grown in Viral Production Medium were were transfected with pAAV-WPRE-GFP (Aldevron), pAAV-Helper (Aldevron) and pAAV-RepCap (AAV 9 Genemedi) at a 1:1:1 plasmid concentration ratio with TransIT-AAViator® Transfection reagent (1.25:1,vol:wt). RevIT™-AAV Enhancer (1 µl/mL) was added at the time of transfection. AAV was harvested at 72 hours post-transfection using chemical lysis. Genome titers were determined via dPCR using primers and a probe targeting the CMV promoter region. The error bars represent the standard deviation of duplicate wells.

RevIT™ allows more titer with less plasmid

RevIT™ AAV Enhancer enables using less plasmid DNA to achieve high titers and percent full capsids. Viral Production Cells 2.0 (3E6 cells/ml of culture, Gibco) cultured in Viral Production Medium (Gibco) in 6-well plates were transfected with TransIT-VirusGEN® Reagent (3 µl/ml of culture) and the indicated concentration of total plasmid DNA (pAAV-hrGFP and pHelper (Agilent), pALD-AAV5 (Aldevron)). If used, RevIT™ AAV Enhancer was added to the transfection mixture at 0.5 µl/ml of culture. AAV was harvested at 72 hours post-transfection. Genome copies were determined by dPCR using primers and a probe targeting the CMV promoter. Capsids were determined using the AAV5 Titration ELISA (Progen). The percent of full capsids is calculated by taking the percentage of genome copies to capsids. The error bars represent the range of duplicate wells.

RevIT™ enhances productivity at every scale

Consistent, high productivity is observed when using RevIT™ AAV Enhancer in both shake flasks and bioreactors. Viral Production Cells 2.0 (3E6 cells/ml of culture, Gibco) cultured in BalanCD HEK293 Medium (Irvine Scientific) supplemented with GlutaMAX (Gibco) were transfected with TransIT-VirusGEN® Reagent (3 µl/ml of culture) and the indicated concentration of total plasmid DNA (pALD-ITR-WPRE-GFP and pALD-HELP (Aldevron), AAV8 Rep-Cap plasmid (GeneMedi)). If used, RevIT™ AAV Enhancer was added to the transfection mixture at 1 µl/ml of culture. AAV was harvested at 72 hours post-transfection. Genome copies were determined by dPCR using primers and a probe targeting the CMV promoter. Capsids were determined using the AAV8 Titration ELISA (Progen). The percent of full capsids is calculated by taking the percentage of genome copies to capsids. The error bars represent the range of duplicate flasks for the 30-ml cultures; the error bars represent the range of duplicate samples pulled from the same bioreactor approximately 20 minutes apart for the 2-L cultures.

RevIT™ Boosts Any Transfection Reagent

While designed to perfect the TransIT-AAViator® system, RevIT™ also enhances AAV genome titers across multiple serotypes when used with *other* common polymer-based reagents like FectoVIR®-AAV and PEIpro®.

Data showing a consistent increase in Genome Copies (GC/mL) across AAV serotypes 2, 5, 8, and 9 when RevIT™ is added (+) compared to the reagent alone (-).

RevIT™: Effective in Different Culture Media

The performance boost from RevIT™ is maintained across different commercial culture media formulations, highlighting its robustness and versatility in diverse lab environments.

RevIT™ consistently increases AAV titers in both Viral Production Medium (VPM) and BalanCD HEK293 Medium across multiple AAV serotypes.

RevIT™: Achieve More with Less pDNA

A key advantage of RevIT™ is its ability to maintain high titers even with significantly reduced amounts of plasmid DNA. This allows for drastic cost savings on expensive plasmids without sacrificing AAV yield.

Data demonstrates that with RevIT™, plasmid DNA concentration can be reduced to as low as 0.75 µg/mL while still achieving higher titers than the standard protocol without the enhancer.

RevIT™: Flexible and Simple Protocol

RevIT™ seamlessly integrates into your existing workflow with a wide window for addition, providing flexibility without complicating your protocol.

RevIT™ is effective when added pre-transfection, post-transfection, or even directly into the transfection complex, simplifying scale-up and automation.