Description
Background: The viral system includes vectors developed from retrovirus (RV), adenovirus (AdV), adeno-associated virus (AAV), lentivirus (LV), and herpes simplex virus (HSV). AAV belongs to the family of Parvoviridae, a group of viruses among the smallest of single-stranded and non-enveloped DNA viruses. There are eight different AAV serotypes reported to date. Recombinant AAV-2 is the most common serotype used in gene delivery, and can be produced at high titers with a helper virus or AAV Helper Free System. AAV-2 can infect both dividing and non-dividing cells and can be maintained in the human host cell, creating the potential for long-term gene transfer. Because AAV-2 is a naturally defective virus, requiring provision of several factors in trans for productive infection, it is considered the safest viral vector to use. Recently a new vector, AAV-DJ, was developed using DNA family shuffling to create a hybrid capsid from 8 different AAV serotypes, resulting in a vector with significantly higher in vitro infection rates across a variety of cells and tissues. Recombinant AAV-2 and AAV-DJ vectors can be purified by CsCl gradient ultracentrifugation, iodixanol discontinuous gradient ultracentrifugation, and high-performance liquid chromatography (HPLC). The most popular technique, CsCl ultracentrifugation, is time consuming process which may result in poor recovery and quality (nonviral protein contamination and a high ratio of genome copies versus infectious units). ViraBind AAV Purification Mega Kit does not involve ultracentrifugation. AAV-2 or AAV-DJ is first purified from viral supernatant with a single-step column, then further purified/concentrated with a centrifugal concentrator (see Assay Principle below). The entire procedure takes about 3 hours. Each preparation can handle up to 10 x 150 mm dishes, resulting in 100 muL of highly purified AAV-2 (Figure 1) or AAV-DJ with low ratio of genome copies versus infectious units. ViraBind AAV Purification Mega Kit provides an efficient system for quick purification of AAV-2 or AAV-DJ with high recovery (>60%). The highly purified and concentrated viruses can be used in primary cell infections and in vivo applications