Development and evaluation of an anti-rabies virus phosphoprotein-specific monoclonal antibody for detection of rabies neutralizing antibodies using RFFIT

PLoS Negl Trop Dis. 2017 Dec 21;11(12):e0006084. doi: 10.1371/journal.pntd.0006084. eCollection 2017 Dec.

Abstract

Background: Rabies is a major public health problem with a fatality rate close to 100%; however, complete prevention can be achieved through pre- or post-exposure prophylaxis. The rapid fluorescent focus inhibition test (RFFIT) is one of the recommended testing methods to determine the production of neutralizing antibodies after vaccination. Here, we report the development of a new monoclonal antibody (mAb) designed to react specifically with Rabies virus (RABV) phosphoprotein (P protein), and the evaluation of its applicability to the RFFIT and its effectiveness as a diagnostic reagent for human rabies.

Methodology/principal findings: The mAb KGH P 16B8 was produced to target the P protein of the Korean KGH RABV strain. An indirect immunofluorescence assay (IFA) was conducted to detect various strains of RABV in various cell lines. Alexa-conjugated KGH P 16B8 (16B8-Alexa) was developed for the RFFIT. The IFA test could detect RABV up to a 1:2,500 dilution, with a detection limit comparable to that of a commercial diagnostic reagent. The sensitivity, specificity, positive predictive value, and negative predictive value of the RFFIT using 16B8-Alexa in 414 clinical specimens were 98.67%, 99.47%, 99.55%, and 98.42%, respectively. The results of the RFFIT with 16B8-Alexa were strongly correlated with those obtained using an existing commercial diagnostic reagent (r = 0.995, p<0.001).

Conclusions/significance: The mAb developed in this study shows high sensitivity and specificity, confirming its clinical utility with the RFFIT to measure the rabies neutralizing antibody titer and establish a diagnosis in human. Thus, 16B8-Alexa is expected to serve as an alternative diagnostic reagent that is widely accessible, with potentially broad applications beyond those of the RFFIT in Korea. Further studies with 16B8-Alexa should provide insight into the immunological mechanism of the P protein of Korean RABV.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibodies, Monoclonal / immunology*
  • Antibodies, Neutralizing / blood*
  • Antibodies, Neutralizing / immunology
  • Antibodies, Viral / blood*
  • Antibodies, Viral / immunology
  • Fluorescent Antibody Technique, Indirect / methods*
  • Molecular Chaperones
  • Neutralization Tests / methods*
  • Phosphoproteins / immunology*
  • Post-Exposure Prophylaxis / methods
  • Rabies / prevention & control*
  • Rabies / virology
  • Rabies Vaccines / immunology
  • Rabies virus / immunology*
  • Vaccination
  • Viral Structural Proteins / immunology*

Substances

  • Antibodies, Monoclonal
  • Antibodies, Neutralizing
  • Antibodies, Viral
  • Molecular Chaperones
  • P phosphoprotein, Rabies virus
  • Phosphoproteins
  • Rabies Vaccines
  • Viral Structural Proteins

Grants and funding

This study supported by grant no. 4837-301-210-13 from the Korea Centers for Disease Control and Prevention(http://cdc.go.kr/CDC/main.jsp). The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.