Comparison of the detection and quantification of rabies antibodies in canine sera

Abstract

218Detection and quantification of rabies antibodies is intended in the first place for checking the immunity to rabies or effectiveness of rabies vaccines. Detection and quantification of virus neutralisation rabies antibodies in the serum is based on inhibition of rabies infection in vivo in animals or in vitro in cell cultures (Atanasiu, 1973;Bourhy and Sureau, 1991). Several suitable procedures have been recommended for determination of titres of virus neutralisation antibodies. e methods most frequently used for quantification of immune response in vaccinated animals after rabies vaccination challenge are serum neutralisation methods carried out on mice and in cell cultures (Smith et al., 1996). WHO recommends in vivo virus neutralisation test on mice (VNT) and in vitro rapid fluorescence focus inhibition test (RFFIT). e VNT on mice is time demanding and too expensive for routine use in virological laboratories. Recently it has been replaced by sensitive, less expensive and more rapid in vitro tests. e RFFIT is highly sensitive and advantageous because of its low time demand. e application of the RFFIT for detection and quantification of rabies antibodies also requires an OIE standard (WHO, 1992). e FAVN (fluorescent antibody virus neutralisation) test was first described by Zalan et al. (1979), and later reworked and modified in the AFSSA laboratory, Malzéville, Nancy, France. It is simple, rapid, safe and economically sustainable, it allows conduct of many serological examinations needed to check animals, particularly dogs, exported to other countries but also checking the immune status of vaccinated animals (Cliquet et al., 1998(Cliquet et al., , 2000. MATERIAL AND METHODS Examined seraEighty-three sera were chosen for comparison of individual methods for rabies antibodies detection from both non-vaccinated (12 sera) and vaccinated (71 sera) pet dogs. Before examination, they were inactivated by exposure to 56°C for 30 min and stored at -20°C. For comparison of reliability, sensitivity and reproducibility of rabies antibodies detection methods, if the sera were taken from vaccinated dogs we selected those that exhibited titres of rabies antibodies lower than 1.0 IU/cm 3 (International Units/cm 3 ), or near the level 0.5 IU per cm 3 , respectively, as detected by the RFFIT. e tests also included titration of reference serum (Copenhagen, Denmark, 30 IU in an ampoule, the dilution of serum containing 0.5 IU/cm 3 antibodies was used for the titration), and the control titration . e results of rabies antibodies levels in non-vaccinated dogs obtained by all three methods were in correlation. e comparison of rabies antibody titres determined in vaccinated dogs using VNT and FAVN methods showed 86.6% correspondence, while those obtained by RFFIT and FAVN methods corresponded in almost 95% of cases.

Achieving Population-Level Immunity to Rabies in Free-Roaming Dogs in Africa and Asia

“…Following seroconversion, protection from rabies virus challenge correlates with peak VNA titre and final titre prior to challenge for inactivated, DNA and modified-live vaccines, with increased susceptibility to challenge once titres drop to near negligible levels (VNA titres <0.1 IU/ml or mouse serum neutralizing antibody titres <1∶2 dilution) [31] – [35] , [37] – [41] . These studies used comparable antibody assays [42] , [43] and virus challenge doses. Titres measured repeatedly over 3–4 years initially peaked and then declined rapidly, followed by a more gradual decline [31] , [33] , [34] , [44] .…”

Generation of Recombinant Rabies Virus CVS-11 Expressing eGFP Applied to the Rapid Virus Neutralization Test

“…The FAVN test is a modified method or an adjusted RFFIT, which belongs among the standard WHO methods for the determination of antibody titers. RFFIT requires the manual counting of the number of fluorescent foci in virus-infected cells [ 11 ], whereas FAVN is objective and suitable for large-scale samples, thereby enabling automated high-throughput detection. The conventional FAVN and RFFIT methods require a FITC-conjugated monoclonal antibody to identify positive or negative cells by the green fluorescence of FITC using fluorescence microscopy.…”

“…However, quantification of the VNA levels against RABV is essential for the timely monitoring of immunization coverage in dog and cat populations and to evaluate the immunity effectiveness of RABV vaccines. According to the WHO, vaccinated animals are sufficiently protected from RABV when the levels of rabies VNA equal or exceed 0.5 IU/mL [ 11 ]. The principle of the detection and quantification of VNA titers is that the cells or animals are infected with viruses not inhibited by antibodies in vitro or in vivo .…”

Isatis indigotica root polysaccharides as adjuvants for an inactivated rabies virus vaccine

“…However, at least 3 injections are required to provide a sufficient virus neutralizing antibody (VNA) titer (at least 0.5 IU/ml), which is a reliable indicator that immune protection against rabies virus infection has been achieved [9,10]. In fact, the rabies neutralizing antibody titer is the most reliable indicator of immune protection [11]. Previous studies have shown that nearly one-third of dogs that received only one injection of a commercial rabies vaccine failed to produce a VNA titer of 0.5 IU/ml [12][13][14][15].…”