Efficient whole brain transduction by systemic infusion of minimally purified AAV-PHP.eB.

Konno, Ayumu; Hirai, Hirokazu

BACKGROUND Adeno-associated virus (AAV) vectors have excellent properties as gene transfer vehicles. The recent development of AAV-PHP.eB, highly BBB-permeable capsid variant of AAV serotype 9, has opened up systemic application for whole brain transduction. To attain high transduction efficacy, much efforts have been paid to purify AAV vectors using gradient centrifugation or column chromatography. These methods are time-consuming, cost substantially and require expensive equipment. NEW METHOD We propose a simple purification method for the production of systemically applicable AAV-PHP.eB targeting the brain. The new method, which we named minimal purification (MP) method, requires only 2 steps: removal of cell debris using a syringe filter and concentration using a disposable ultrafiltration device. RESULTS The MP method yielded 2 times more AAV-PHP.eB than the standard ultracentrifuge purification (UCP) method. Intravenous injection of AAV-PHP.eB prepared using the MP method caused robust whole brain transduction without overt toxicity on the liver and kidney. Moreover, we found almost no difference in cellular density and morphology of brain microglia between control mice and mice treated systemically with the MP viral solution, suggesting no influence of the viral injection on brain immunity. COMPARISON WITH EXISTING METHODS The new method, which requires only a benchtop centrifuge and takes only 2 - 4 h to obtain a ready-to-use viral solution, is much less expensive than the existing UCP method, and can avoid cumbersome and time-consuming purification processes. CONCLUSIONS This simplified method further expands the use of AAV vectors in the neuroscience community.