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Immunodiagnosis of feline immunodeficiency virus infection using recombinant viral p17 and p24
G Reid1, M A Rigby, M McDonald
1MRC Retrovirus Research Laboratory, University of Glasgow, UK.
Abstract:
The coding sequences of p17 and p24 of the Glasgow-8 strain of feline immunodeficiency virus (FIV) were amplified using the polymerase chain reaction and cloned into plasmid vectors. The predicted amino-acid sequences of FIV/Glasgow-8 p17 and p24 were compared with those of the Petaluma and PPR isolates of FIV. As seen with other retroviruses, these gag gene products are highly conserved, indicating that the protein products would be suitable antigens to detect anti-FIV antibodies in an immunoassay. Both p17 and p24 were stably expressed in Escherichia coli as fusion proteins with glutathione S transferase. A pure preparation of each fusion protein was obtained from induced bacterial lysates by affinity chromatography using glutathione-agarose beads. These recombinant proteins were used in an enzyme-linked immunosorbent assay to detect antibodies directed against FIV p17 and p24 in cat sera. This assay allows the identification of seropositive cats following infection with FIV and has greater sensitivity and specificity than a currently available immunodiagnostic test.
Insights
Researchers developed a new feline immunodeficiency virus (FIV) immunoassay using recombinant FIV proteins. This test accurately identifies FIV-infected cats with improved sensitivity and specificity.
Area of Science:
- Veterinary Virology
- Immunology
- Molecular Biology
Background:
- Feline immunodeficiency virus (FIV) is a significant pathogen in domestic cats.
- Accurate diagnostic tools are crucial for managing FIV outbreaks.
- Existing immunodiagnostic tests for FIV may have limitations in sensitivity or specificity.
Purpose of the Study:
- To develop and evaluate a novel immunoassay for detecting FIV antibodies.
- To utilize conserved FIV gag gene products (p17 and p24) as antigens.
- To improve the sensitivity and specificity of FIV diagnostics.
Main Methods:
- Amplification and cloning of FIV Glasgow-8 p17 and p24 coding sequences.
- Expression of FIV p17 and p24 as glutathione S-transferase fusion proteins in Escherichia coli.
- Purification of recombinant proteins using affinity chromatography.
- Development of an enzyme-linked immunosorbent assay (ELISA) using recombinant antigens.
Main Results:
- FIV p17 and p24 amino acid sequences are highly conserved among different FIV isolates.
- Recombinant FIV p17 and p24 fusion proteins were successfully expressed and purified.
- The developed ELISA demonstrated high sensitivity and specificity in detecting anti-FIV antibodies in cat sera.
Conclusions:
- Recombinant FIV p17 and p24 proteins are suitable antigens for FIV detection.
- The novel ELISA offers a more sensitive and specific method for identifying seropositive cats.
- This assay can aid in the diagnosis and management of feline immunodeficiency virus infections.
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