Gene Vector and Virus Core–  

Care and Handling of AAV

AAV
It is useful to think of AAV as a very large protein (MW= ~5,200,000).  As such it is relatively stable but should be treated with at least as much care as other proteins.  It is resistant to most proteases and a variety of chemical treatments that would inactive many viruses.  However, because it is so large, it has a large surface area and is more likely to stick to hydrophobic surfaces and aggregate than smaller proteins.

DON'T:

  1. Don’t expose to environmental extremes (pH, chelating agents like EDTA, temperature, organic solvents, protein denaturants, strong detergents, etc.) that denature proteins.
  2. Don’t introduce air into the sample by vortexing, blowing bubbles and similar operations (which results in protein denaturation).
  3. Don’t dry (which results in protein denaturation).
  4. Don’t freeze and thaw multiple times.  AAV is more stable than many viruses or proteins and can be frozen and thawed several times with minimal loss of activity but it is best to avoid.
  5. Don’t expose to “regular” plastics (especially polystyrene or similar very hydrophobic plastics) for prolonged periods in a liquid phase.  Most AAVs are very sticky and losses can occur if they are exposed to regular plastics (e.g., tubes, cell culture plates, pipette tips) if they are not frozen.  It is best to store thawed AAV in siliconized or low protein binding tubes and pipette it with similar pipette tips.  Pluronic F-68 used at 0.01%-0.1% in the formulation buffer will minimize sticking if regular plastics are used.
  6. Don’t’ dilute into low salt.  Some AAVs (e.g., AAV-2) aggregate in low salt and if the aggregates are large they will be non-infectious.

DO:

  1. Do aliquot and freeze at -80C for long term storage if the virus is not used within 1-2 weeks.

NOTE:

  1.  AAV appears to retain infectious activity for > 10 years when stored at -80C.

 

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