A Quantitative Dot Blot Assay for AAV Titration and Its Use for Functional Assessment of the Adeno-associated Virus Assembly-activating Proteins

This method can determine both purified and the non-purified AV vac titers and assess the ability for the AAV AAP to promote assembly of cognate and heterologous AV serotype-capsis. The main advantage of this technique is that it is a straightforward inexpensive method that avoids the drawbacks of other AAV titration methods. Though this method has commonly been used for AAV vector quantitation, it can also be applied to address various fundamental questions about AAV biology as shown in this protocol.

On day zero, after culturing HEK 293 cells to 90%confluency, prepare the transfection reagent by mixing the plasmids in 96 microliters of PBS without calcium or magnesium in sterile 1.5 milliliter micro-centrifuge tubes. As indicated in this table, the total amount of DNA is two micrograms per well. Add four microliters of PEI solution to the PBS plasmid DNA mix.