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A high throughput neutralization test based on GFP expression by recombinant rabies virus.

Jillybeth Burgado1, Lauren Greenberg1, Mike Niezgoda1

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|December 15, 2018
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Summary

A new automated high throughput neutralization test (HTNT) accurately detects rabies virus neutralizing antibodies (VNAs). This method offers a faster, more efficient alternative to the traditional Rapid Fluorescent Focus Inhibition Test (RFFIT).

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Area of Science:

  • Immunology
  • Virology
  • Biotechnology

Background:

  • Rabies virus neutralizing antibodies (VNAs) are crucial for vaccine effectiveness in humans and animals.
  • The traditional Rapid Fluorescent Focus Inhibition Test (RFFIT) for VNA detection is labor-intensive, low-throughput, and semi-quantitative.
  • RFFIT lacks image storage for future analysis, limiting its utility.

Purpose of the Study:

  • To develop and validate an automated, high-throughput neutralization test (HTNT) for rabies VNAs.
  • To compare the performance of HTNT with the established RFFIT method.
  • To assess the potential of HTNT for rabies VNA determination.

Main Methods:

  • Development of an HTNT utilizing a recombinant rabies virus (RABV) expressing green fluorescent protein (GFP) with a nuclear localization signal (NLS).
  • Comparison of HTNT with RFFIT using a panel of human and animal rabies serum samples (positive and negative).
  • Quantitative assessment of RABV-infected cells and image storage for future analysis in HTNT.

Main Results:

  • The HTNT demonstrated 100% correlation with RFFIT for all tested human and animal serum samples.
  • HTNT is a quantitative method that stores images for subsequent analysis.
  • While generally agreeable, HTNT titers were typically lower than RFFIT titers, likely due to quantification method differences.

Conclusions:

  • The developed HTNT is a viable, automated, and high-throughput alternative for rabies VNA determination.
  • HTNT offers advantages over RFFIT in terms of efficiency and data archiving.
  • The study highlights the potential of HTNT assays for routine rabies VNA titer determination.