TY - JOUR
T1 - The Bottlenecks of Preparing Virus Particles by Size Exclusion for Antibody Generation
AU - Lee, Chi-Hsin
AU - Huang, Peng-Nien
AU - Mwale, Pharaoh Fellow
AU - Wang, Wei-Chu
AU - Leu, Sy-Jye
AU - Tseng, Sung-Nien
AU - Shih, Shin-Ru
AU - Chiang, Liao-Chun
AU - Mao, Yan-Chiao
AU - Tsai, Bor-Yu
AU - Dlamini, Nhlanhla Benedict
AU - Nguyen, Tien-Cuong
AU - Tsai, Chen-Hsin
AU - Yang, Yi-Yuan
PY - 2022/10
Y1 - 2022/10
N2 - Enterovirus 71 (EV71) is the major etiological agent contributing to the development of hand-foot-mouth disease (HFMD). There are not any global available vaccines or antibody drugs against EV71 released yet. In this study, we perform the virus immunization in a cost-effective and convenient approach by preparing virus particles from size exclusion and immunization of chicken. Polyclonal yolk-immunoglobulin (IgY) was simply purified from egg yolk and monoclonal single-chain variable fragments (scFv) were selected via phage display technology with two scFv libraries containing 6.0 × 106 and 1.3 × 107 transformants. Specific clones were enriched after 5 rounds of bio-panning and four identical genes were classified after the sequence analysis. Moreover, the higher mutation rates were revealed in the CDR regions, especially in the CDR3. IgY showed specific binding activities to both EV71-infected and Coxsackievirus 16-infected cell lysates and high infectivity inhibitory activity of EV71. However, while IgY detected a 37 kDa protein, the selected scFv seemingly detected higher size proteins which could be cell protein instead of EV71 proteins. Despite the highly effective chicken antibody generation, the purity of virus particles prepared by size exclusion is the limitation of this study, and further characterization should be carried out rigorously.
AB - Enterovirus 71 (EV71) is the major etiological agent contributing to the development of hand-foot-mouth disease (HFMD). There are not any global available vaccines or antibody drugs against EV71 released yet. In this study, we perform the virus immunization in a cost-effective and convenient approach by preparing virus particles from size exclusion and immunization of chicken. Polyclonal yolk-immunoglobulin (IgY) was simply purified from egg yolk and monoclonal single-chain variable fragments (scFv) were selected via phage display technology with two scFv libraries containing 6.0 × 106 and 1.3 × 107 transformants. Specific clones were enriched after 5 rounds of bio-panning and four identical genes were classified after the sequence analysis. Moreover, the higher mutation rates were revealed in the CDR regions, especially in the CDR3. IgY showed specific binding activities to both EV71-infected and Coxsackievirus 16-infected cell lysates and high infectivity inhibitory activity of EV71. However, while IgY detected a 37 kDa protein, the selected scFv seemingly detected higher size proteins which could be cell protein instead of EV71 proteins. Despite the highly effective chicken antibody generation, the purity of virus particles prepared by size exclusion is the limitation of this study, and further characterization should be carried out rigorously.
KW - Animals
KW - Hand, Foot and Mouth Disease
KW - Enterovirus
KW - Virion/genetics
KW - Single-Chain Antibodies/genetics
KW - Egg Yolk
KW - Chickens
KW - Enterovirus A, Human
U2 - 10.3390/ijms232112967
DO - 10.3390/ijms232112967
M3 - Article
C2 - 36361757
VL - 23
JO - International Journal of Molecular Sciences
JF - International Journal of Molecular Sciences
SN - 1661-6596
IS - 21
ER -