Purification of infectious adenovirus in two hours by ultracentrifugation and tangential flow filtration

Journal Article · Fri Jun 17 00:00:00 EDT 2005 · Biochemical and Biophysical Research Communications

DOI:https://doi.org/10.1016/j.bbrc.2005.03.227· OSTI ID:20710799

Ugai, Hideyo; Yamasaki, Takahito; Hirose, Megumi; Inabe, Kumiko; Kujime, Yukari; Terashima, Miho ^[1]; Liu, Bingbing ^[1]; Tang, Hong ^[1]; Zhao, Mujun ^[2]; Murata, Takehide ^[1]; Kimura, Makoto; Pan, Jianzhi ^[1]; Obata, Yuichi ^[3]; Hamada, Hirofumi ^[4]; Yokoyama, Kazunari K ^[1]

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1. Gene Engineering Division, BioResource Center, Tsukuba Institute, RIKEN - Institute of Physical and Chemical Research, 3-1-1 Koyadai, Tsukuba, Ibaraki 305-0074 (Japan)
2. Institute of Biochemistry and Cell Biology, Shanghai Institute for Biological Science, Chinese Academy of Sciences, Shanghai 20031 (China)
3. Department of Biological Systems, Tsukuba Institute, RIKEN - Institute of Physical and Chemical Research, 3-1-1 Koyadai, Tsukuba, Ibaraki 305-0074 (Japan)
4. Department of Molecular Medicine, Sapporo Medical University, Sapporo, Hokkaido 060-8556 (Japan)

Adenoviruses are excellent vectors for gene transfer and are used extensively for high-level expression of the products of transgenes in living cells. The development of simple and rapid methods for the purification of stable infectious recombinant adenoviruses (rAds) remains a challenge. We report here a method for the purification of infectious adenovirus type 5 (Ad5) that involves ultracentrifugation on a cesium chloride gradient at 604,000g for 15 min at 4 deg C and tangential flow filtration. The entire procedure requires less than two hours and infectious Ad5 can be recovered at levels higher than 64% of the number of plaque-forming units (pfu) in the initial crude preparation of viruses. We have obtained titers of infectious purified Ad5 of 1.35 x 10{sup 10} pfu/ml and a ratio of particle titer to infectious titer of seven. The method described here allows the rapid purification of rAds for studies of gene function in vivo and in vitro, as well as the rapid purification of Ad5.

• OSTI ID: 20710799
• Journal Information: Biochemical and Biophysical Research Communications, Vol. 331, Issue 4; Other Information: DOI: 10.1016/j.bbrc.2005.03.227; PII: S0006-291X(05)00752-7; Copyright (c) 2005 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved; Country of input: International Atomic Energy Agency (IAEA); ISSN 0006-291X
• Country of Publication: United States
• Language: English

Related Subjects

60 APPLIED LIFE SCIENCES
ADENOVIRUS
CESIUM CHLORIDES
FILTRATION
GENES
IN VITRO
IN VIVO
PURIFICATION
ULTRACENTRIFUGATION