Tweet
Biotechnol Rep (Amst). 2014 May 17;3:108-116. doi: 10.1016/j.btre.2014.05.001. eCollection 2014 Sep.
Stable lentiviral transformation of CHO cells for the expression of the hemagglutinin H5 of avian influenza virus in suspension culture.
Pose AG1, Morell NO1, Matos DA1, Rodr?guez ER1, Rodr?guez ES1, Cordero LR2, Molt? MPR1, Ramos EMG1, Guti?rrez A?1,2, P?rez LG1, G?mez JN2, Redondo AV2, Nordelo CB1.
Author information
Abstract
Avian influenza virus H5N1 has caused extensive damage worldwide among poultry and humans. Effective expression systems are needed for the production of viral proteins required for monitoring this devastating disease. The present study deals with the establishment of a stable expression system for the hemagglutinin H5 (HAH5) of avian influenza virus using CHO cells in suspension culture transduced with a recombinant lentiviral vector. The synthetic gene coding the HAH5 protein was inserted in a lentiviral vector with the aim of performing a stable transduction of CHO cells. After the selection of recombinant clones, the one with the highest expression level was adapted to suspension culture and the HAH5 protein was purified by immunoaffinity chromatography from the culture supernatant. There were no significant differences when this protein, purified or direct from the culture supernatant of CHO or SiHa cells, was utilized in an immunologic assay using positive and negative sera as reference. It was also demonstrated that the HAH5 protein in its purified form is able to bind anti-HAH5 antibodies generated with proper and non-proper folded proteins. The results demonstrate that the CHO cell line stably transduced with a lentiviral vector coding the sequence of the HAH5 protein and cultured in suspension can be a suitable expression system to obtain this protein for diagnostic purpose in a consistent and reliable manner.
KEYWORDS:
CHO cells; Hemagglutinin; Lentiviral vector; Suspension culture
PMID: 28626654 PMCID: PMC5466102 DOI: 10.1016/j.btre.2014.05.001
Free full text
Stable lentiviral transformation of CHO cells for the expression of the hemagglutinin H5 of avian influenza virus in suspension culture.
Pose AG1, Morell NO1, Matos DA1, Rodr?guez ER1, Rodr?guez ES1, Cordero LR2, Molt? MPR1, Ramos EMG1, Guti?rrez A?1,2, P?rez LG1, G?mez JN2, Redondo AV2, Nordelo CB1.
Author information
Abstract
Avian influenza virus H5N1 has caused extensive damage worldwide among poultry and humans. Effective expression systems are needed for the production of viral proteins required for monitoring this devastating disease. The present study deals with the establishment of a stable expression system for the hemagglutinin H5 (HAH5) of avian influenza virus using CHO cells in suspension culture transduced with a recombinant lentiviral vector. The synthetic gene coding the HAH5 protein was inserted in a lentiviral vector with the aim of performing a stable transduction of CHO cells. After the selection of recombinant clones, the one with the highest expression level was adapted to suspension culture and the HAH5 protein was purified by immunoaffinity chromatography from the culture supernatant. There were no significant differences when this protein, purified or direct from the culture supernatant of CHO or SiHa cells, was utilized in an immunologic assay using positive and negative sera as reference. It was also demonstrated that the HAH5 protein in its purified form is able to bind anti-HAH5 antibodies generated with proper and non-proper folded proteins. The results demonstrate that the CHO cell line stably transduced with a lentiviral vector coding the sequence of the HAH5 protein and cultured in suspension can be a suitable expression system to obtain this protein for diagnostic purpose in a consistent and reliable manner.
KEYWORDS:
CHO cells; Hemagglutinin; Lentiviral vector; Suspension culture
PMID: 28626654 PMCID: PMC5466102 DOI: 10.1016/j.btre.2014.05.001
Free full text