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Production of measles nucleoprotein in different expression systems and its use as a diagnostic reagent
A Warnes1, A R Fooks, J R Stephenson
1Biologics Division, PHLS, CAMR, Porton Down, Salisbury, UK.
Abstract:
Measles nucleoprotein has been successfully expressed in three different hosts, bacterial (Escherichia coli BL21 DE3), insect (Spodoptera frugiperda; Sf9) and mammalian (primary human fibroblasts) cells, each producing an antigenic protein of M(r) 60 kDa. The nucleoprotein produced in all three hosts was used in an ELISA for the detection of antibodies to measles virus in a cohort of haemagglutinin-positive or -negative human sera. Data produced from baculovirus and adenovirus-based antigens indicated that there was good correlation between the ELISA results and previous haemagglutination inhibition test data, and there was little background interference by cellular proteins or the development of false positive or negative results. The assay was rapid as it could be carried out in under 4 h, sensitive as the sera could be diluted by at least 1000-fold, and versatile as both IgG and IgM could be detected and differentiated.
Insights
Measles nucleoprotein was expressed in bacterial, insect, and mammalian cells for antibody detection. The developed ELISA assay is rapid, sensitive, and versatile for diagnosing measles infections.
Area of Science:
- Virology
- Immunology
- Biotechnology
Background:
- Measles virus diagnosis relies on detecting specific antibodies.
- Previous diagnostic methods may lack speed, sensitivity, or versatility.
Purpose of the Study:
- To express measles nucleoprotein in multiple hosts for diagnostic applications.
- To develop and evaluate an Enzyme-Linked Immunosorbent Assay (ELISA) for measles antibody detection.
Main Methods:
- Measles nucleoprotein was expressed in Escherichia coli, Spodoptera frugiperda (Sf9), and primary human fibroblast cells.
- An ELISA was developed using the expressed nucleoprotein to detect measles-specific IgG and IgM antibodies in human sera.
- Assay performance was compared with traditional haemagglutination inhibition tests.
Main Results:
- Antigenic nucleoprotein (60 kDa) was successfully produced in all three host systems.
- ELISA results showed good correlation with haemagglutination inhibition test data.
- The assay demonstrated high sensitivity (dilution up to 1000-fold), speed (under 4 hours), and specificity with minimal background interference.
Conclusions:
- Recombinant measles nucleoprotein expressed in diverse hosts is suitable for developing sensitive diagnostic assays.
- The developed ELISA offers a rapid, sensitive, and versatile method for detecting measles virus antibodies, differentiating between IgG and IgM.
- This approach holds promise for improved measles diagnosis and surveillance.