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Calculating rabies virus neutralizing antibodies titres by flow cytometry
Juliano Bordignon1, Fabiano Comin, Sílvia Córdoba P Ferreira
1Departament of Microbiology, Immunology and Parasitology, Federal University of Santa Catarina, Florianópolis, SC, 88040-900, Brazil.
Revista Do Instituto De Medicina Tropical De Sao Paulo
|August 7, 2002
Summary
Flow cytometry offers a reliable method for assessing rabies neutralizing antibody (VNA) levels. This technique accurately measures VNA titers in immunized animal sera, correlating well with standard methods.
Area of Science:
- Virology
- Immunology
- Analytical Chemistry
Background:
- Rabies neutralizing antibody (VNA) levels are crucial for evaluating vaccine efficacy and treatment success.
- Existing methods for VNA assessment, such as mouse inoculation and cell-culture fluorescence inhibition tests, have limitations in accuracy and ease of use.
- There is a need for improved, reliable, and accessible methods for VNA determination.
Purpose of the Study:
- To describe and validate a novel flow cytometry (FC)-based neutralization assay for quantifying VNA.
- To compare the performance of the FC assay with established methods for VNA titration.
- To assess the potential of FC as a suitable technique for VNA evaluation in various serum samples.
Main Methods:
- A cell-culture-based neutralization test was developed using flow cytometry (FC).
- Serum samples were incubated with rabies virus and BHK-21 cells.
- Cells were stained with a rabies virus nucleocapsid protein-specific antibody conjugate, and infection inhibition was measured by FC.
- A correlation curve was established using a reference serum to extrapolate VNA titers (IU/ml).
Main Results:
- The FC-based assay demonstrated a significant correlation (r = 0.7, p < 0.01) with standard VNA titration methods.
- The results indicate effective pairing between titers obtained by FC and standard tests.
- The study successfully built a correlation curve for extrapolating VNA titers from percentage of infective inhibition.
Conclusions:
- Flow cytometry presents a suitable and accurate technique for evaluating VNA in sera from immunized animals.
- The FC method shows promise for application in human serum samples.
- Further studies are required to determine the sensitivity and specificity of FC compared to gold-standard techniques.