Preparation of purified, sterilized, and stable adenovirus vectors using albumin.
Cesium chloride ultracentrifugation combined with dialysis is a standard method for preparing a high titer of purified recombinant adenovirus. However, it was found that sterilization of the filter membrane after dialysis led to a complete loss of adenovirus activity. A virus pellet was visualized after centrifugation shortly and electron microscopy revealed an aggregation of recombinant virus within the filter membrane following dialysis with 10% (V/V) glycerol in phosphate buffered saline. The entrapment of aggregated adenovirus by the filter membrane explains why adenovirus activity is lost following sterilization. The addition of 1% albumin prevented viral aggregation and allowed the purified virus to retain its activity after filter sterilization. Furthermore, viral activity was retained within the 1% albumin solution for at least 1 week at 37 degrees C and for 2 weeks at 4 degrees C, whereas viral activity within the albumin-free solution was quickly lost.
Hosokawa
Mami
M
Department of Medical Oncology and Clinical Research, National Shikoku Cancer Center, 13-Horinocuhi, Matsuyama, Ehime, Japan.
Hama
Seiji
S
Mandai
Koichi
K
Okuda
Kazuaki
K
Takashima
Shigemitsu
S
Tajiri
Hisao
H
Eguchi
Kenji
K
Heike
Yuji
Y
D000256:Adenoviridae / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D000818:Animals; D002460:Cell Line; D005822:Genetic Vectors / Q000302:isolation & purification* / Q000502:physiology; D006801:Humans; D011995:Recombination, Genetic; D012710:Serum Albumin, Bovine; D014776:Virus Cultivation / Q000379:methods
D012710:Serum Albumin, Bovine