Expression and purification of polioviral proteins in E. coli, and production of antisera as reagents for immunological assays
Poliomyelitis, caused by poliovirus, is on the verge of eradication, and the world is preparing to shift from live to inactivated polio vaccine. In view of the requirement of non-infectious reagents, especially protein antigens, for surveillance during the final phase of poliovirus eradication, we h...
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| Published in | Protein expression and purification Vol. 128; pp. 115 - 122 |
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| Main Authors | , , , |
| Format | Journal Article |
| Language | English |
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01.12.2016
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| Abstract | Poliomyelitis, caused by poliovirus, is on the verge of eradication, and the world is preparing to shift from live to inactivated polio vaccine. In view of the requirement of non-infectious reagents, especially protein antigens, for surveillance during the final phase of poliovirus eradication, we have attempted to generate reagents that may be of use for the development of diagnostic tests. Polioviral proteins VP0, VP3, VP1, and 3AB were expressed in Escherichia coli using the autoinduction system, purified, and the proteins were used to raise antisera in rabbits. All antisera detected all three serotypes of PV from infected cell lysates in enzyme-linked immunosorbent assay, immunofluorescence and western blotting.
•Poliovirus structural and non-structural proteins were expressed in E. coli using autoinduction.•Purified proteins were used for raising antibodies.•Antisera raised could detect antigens in virus-infected cells by different immunoassays.•Antisera could detect both cleaved and uncleaved as well as native and denatured viral proteins.•These reagents may be useful in vaccine manufacturing and seromonitoring in the future. |
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| AbstractList | Poliomyelitis, caused by poliovirus, is on the verge of eradication, and the world is preparing to shift from live to inactivated polio vaccine. In view of the requirement of non-infectious reagents, especially protein antigens, for surveillance during the final phase of poliovirus eradication, we have attempted to generate reagents that may be of use for the development of diagnostic tests. Polioviral proteins VP0, VP3, VP1, and 3AB were expressed in Escherichia coli using the autoinduction system, purified, and the proteins were used to raise antisera in rabbits. All antisera detected all three serotypes of PV from infected cell lysates in enzyme-linked immunosorbent assay, immunofluorescence and western blotting. Poliomyelitis, caused by poliovirus, is on the verge of eradication, and the world is preparing to shift from live to inactivated polio vaccine. In view of the requirement of non-infectious reagents, especially protein antigens, for surveillance during the final phase of poliovirus eradication, we have attempted to generate reagents that may be of use for the development of diagnostic tests. Polioviral proteins VP0, VP3, VP1, and 3AB were expressed in Escherichia coli using the autoinduction system, purified, and the proteins were used to raise antisera in rabbits. All antisera detected all three serotypes of PV from infected cell lysates in enzyme-linked immunosorbent assay, immunofluorescence and western blotting. •Poliovirus structural and non-structural proteins were expressed in E. coli using autoinduction.•Purified proteins were used for raising antibodies.•Antisera raised could detect antigens in virus-infected cells by different immunoassays.•Antisera could detect both cleaved and uncleaved as well as native and denatured viral proteins.•These reagents may be useful in vaccine manufacturing and seromonitoring in the future. Poliomyelitis, caused by poliovirus, is on the verge of eradication, and the world is preparing to shift from live to inactivated polio vaccine. In view of the requirement of non-infectious reagents, especially protein antigens, for surveillance during the final phase of poliovirus eradication, we have attempted to generate reagents that may be of use for the development of diagnostic tests. Polioviral proteins VP0, VP3, VP1, and 3AB were expressed in Escherichia coli using the autoinduction system, purified, and the proteins were used to raise antisera in rabbits. All antisera detected all three serotypes of PV from infected cell lysates in enzyme-linked immunosorbent assay, immunofluorescence and western blotting. • Poliovirus structural and non-structural proteins were expressed in E. coli using autoinduction. • Purified proteins were used for raising antibodies. • Antisera raised could detect antigens in virus-infected cells by different immunoassays. • Antisera could detect both cleaved and uncleaved as well as native and denatured viral proteins. • These reagents may be useful in vaccine manufacturing and seromonitoring in the future. |
| Author | Hegde, N.R. Uma, Madala Nagalekshmi, K. Rao, P.P. |
| AuthorAffiliation | Ella Foundation, Genome Valley, Turkapally, Shameerpet Mandal, Hyderabad, 500078, India |
| AuthorAffiliation_xml | – name: Ella Foundation, Genome Valley, Turkapally, Shameerpet Mandal, Hyderabad, 500078, India |
| Author_xml | – sequence: 1 givenname: Madala surname: Uma fullname: Uma, Madala – sequence: 2 givenname: P.P. surname: Rao fullname: Rao, P.P. – sequence: 3 givenname: K. surname: Nagalekshmi fullname: Nagalekshmi, K. – sequence: 4 givenname: N.R. surname: Hegde fullname: Hegde, N.R. email: hegden@ellafoundation.org |
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| Keywords | Protein expression Autoinduction Poliovirus Immunoassays |
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| Title | Expression and purification of polioviral proteins in E. coli, and production of antisera as reagents for immunological assays |
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