Molecular Epidemiology Investigation of Bovine Norovirus in Southern Taiwan and Preparation and Application of the Monoclonal Antibodies of Capsid Protein

• Main Authors: Guan-Cheng Chen, 陳冠呈
• Other Authors: Ming-Hui Liao
• Language: zh-TW
• Published: 2011
• Online Access: http://ndltd.ncl.edu.tw/handle/09341424008461343668

碩士 === 國立屏東科技大學 === 生物科技研究所 === 99 === Noroviruses ( NoV ), belong to the family Caliciviridae and have been identified in human beings and in several animal species including the bovine. NoV is the major cause of nonbacterial gastroenteritis in humans. The bovine norovirus was first isolated from the diarrhoea stools of newborn calves. In this experiment, we collected 531 bovine fecal samples in May 2009 to April 2010 from Kaohsiung and Pingtung. Reverse transcription-polymerase chain reaction ( RT-PCR ) was used to detect the presence of bovine NoV. A positive ratio of 3.0 % ( 16 out of 531 samples ) for Bovine NoV infection was found in the fecal samples. Due to the lack of cell culture or animal models for NoV multiply. We used the technology of protein expression. The capsid protein gene of NoV was cloned into prokaryotic expression vector pET32a. The purified recombinant capsid protein was used to immunized BALB/c mice. The anti-serum of recombinant protein can recognize the NoV capsid protein by Western blot assay. The myeloma cell NS-1 fused with spleen cell from immunized BALB/c mice to produce hybridoma cells. Antibodies secreted from hybridoma cells were screened by ELISA, Western-blot and Immuno-dot. There were 5 monoclonal antibodies obtained in this study. They can be applied to diagnostic NoV in the future.