My lab's vector of choice is AAV, with nearly every experiment requiring AAV. Before joining my lab, I had never worked with AAV, so naturally I had to package some virus for my first experiment. It was a bit intimidating, but I had my lab’s protocols and some great co-workers to help me out. Even with these tools, I found myself writing AAV production tricks into the margins of my protocol. While these tips weren’t critical to the experiment, they definitely made my life easier!  In this post, I’ll share some AAV production, purification, and titration tips, while also summarizing the basic steps and analyses needed for packaging AAV.

While lentiviral vectors are popular gene delivery tools, producing lentivirus, can pose certain challenges. Whether choosing a system that is the best fit for the experiment, trying to produce virus of a usable titer, or fine-tuning selection and expression in your target cell line, researchers often find themselves faced with a roadblock. In this post, we will provide an overview of some of the common challenges associated with producing and using lentivirus and offer some tips and tricks for overcoming these hurdles.

Many of us take comfort in the fact that it’s often not quantity, but quality that really matters. Well, it turns out this isn’t the case for using AAV. When it comes to infecting cells, titer, the amount of virus used, really does matter. (*psst*, quality definitely also matters).

Check out this post for a refresher on AAV titers

Remember the game show “The $25,000 Pyramid” where one player tries to get the other to guess a category by listing off things that fall into that category? Okay, let’s play. I’ll list the examples and you try to guess the category:

ELISA...
qPCR...
Digital droplet PCR...
DNA dot blot...
Transduction assay...
SDS-PAGE...
Electron microscopy…

Any guesses?