Optimization and evaluation of an influenza A (H5) pseudotyped lentiviral particle-based serological assay

Abstract 

Background

Novel serological methods provide alternative options for sero-diagnosis, sero-epidemiology and for determining evidence of naturally acquired or vaccine induced immunity. Micro-neutralization tests are currently the gold standard for serological studies of highly pathogenic avian influenza in mammalian species but require handling live virus in a biosafety level (BSL) 3 environment. We previously reported the use of H5 pseudotyped lentiviral particles (H5pp) as an alternative to micro-neutralization tests in a BSL-2 setting (Nefkens et al., 2007).

Objective

To optimize and evaluate this newly developed H5pp assay on relevant clinical specimens.

Study design

We optimise and evaluate the performance of the H5pp assay using well-characterized sera from humans with confirmed H5N1 disease or controls.

Results

The H5pp assay is a reliable serological method for the detection and quantification of neutralizing antibody to H5-viruses.

Conclusion

H5pp provide a reliable and safe alternative for sero-diagnosis and sero-epidemiology of H5N1 infections in a BSL-2 setting.

Abbreviations: HPAI, highly pathogenic avian influenza, MN, micro-neutralization, BSL, biosafety level, H5, haemagglutinin of H5N1 HPAI virus, H5pp, H5 pseudotyped viral particles, HI, haemagglutination inhibition assay, RT-PCR, reverse transcription polymerase chain reaction, HA, haemagglutinin, MDCK, Madin–Darby canine kidney, DMEM, Dulbecco modified Eagle medium, RLU, relative luminescence unit, ROC curve, receiver operating characteristic curve, CI, confidence interval, AUC, area under curve, SE, standard error, “c”, closest-to-(0,1) criterion, “J”, Youden index, PPV, positive predictive value, NPV, negative predictive value, NA, neuraminidase

Keywords: Influenza, Pseudotyped lentiviral particles, H5N1, Diagnosis, Serology, Pseudoparticle