An Innovative Pseudotypes-Based Enzyme-Linked Lectin Assay for the Measurement of Functional Anti-Neuraminidase Antibodies.
Antibodies (Ab) to neuraminidase (NA) play a role in limiting influenza infection and might help reduce the disease impact. The most widely used serological assay to measure functional anti-NA immune responses is the Enzyme-Linked Lectin Assay (ELLA) which relies on hemagglutinin (HA) mismatched virus reassortants, or detergent treated viruses as the NA source to overcome interference associated with steric hindrance of anti-HA Ab present in sera. The difficulty in producing and handling these reagents, which are not easily adapted for screening large numbers of samples, limits the routine analysis of functional anti-NA Ab in clinical trials. In this study, we produced influenza lentiviral pseudoparticles (PPs) containing only the NA antigen (NA-PPs) with a simple two-plasmid co-transfection system. NA-PPs were characterized and tested as an innovative source of NA in the NA inhibition (NI) assay. Both swine A/California/07/2009 (H1N1) and avian A/turkey/Turkey/01/2005 (H5N1) N1s within NA-PPs retained their sialidase activity and were specifically inhibited by homologous and N1 subtype-specific, heterologous sheep sera. Moreover, A/California/07/2009 N1-PPs were a better source of NA compared to whole live and detergent treated H1N1 viruses in ELLA, likely due to lack of interference by anti-HA Ab, and absence of possible structural modifications caused by treatment with detergent. This innovative assay is safer and applicable to all NAs. Taken together, these results highlight the potential of NA-PPs-based NI assays to be developed as sensitive, flexible, easy to handle and scalable serological tests for routine NA immune response analysis.
University of Siena, Department of Life Sciences, Via A. Moro, 53100, Siena, Italy.
Marua
M
Prevato
GSK, Research Center, Via Fiorentina 1, 53100, Siena, Italy.
Roberta
R
Cozzi
GSK, Research Center, Via Fiorentina 1, 53100, Siena, Italy.
Alfredo
A
Pezzicoli
Section of Electron Microscopy, Great Equipment Center, Tuscia University, 01100, Viterbo, Italy.
Anna Rita
AR
Taddei
GSK, Research Center, Via Fiorentina 1, 53100, Siena, Italy.
Ilaria
I
Ferlenghi
GSK, Vaccine Research, Holly Springs, North Carolina, 27540, United States of America; GSK, Vaccine Research, Cambridge, Massachusetts, 02139, United States of America.
Avishek
A
Nandi
University of Siena, Department of Molecular and Developmental Medicine, Via A. Moro, 53100, Siena, Italy.
Emanuele
E
Montomoli
GSK, Vaccine Research, Cambridge, Massachusetts, 02139, United States of America.
Ethan C
EC
Settembre
GSK, Research Center, Via Fiorentina 1, 53100, Siena, Italy.
Sylvie
S
Bertholet
GSK, Research Center, Via Fiorentina 1, 53100, Siena, Italy.
Alessandra
A
Bonci
GSK, Research Center, Via Fiorentina 1, 53100, Siena, Italy; GSK, Peter Merian Strasse, 4056, Basel, Switzerland.
Francois
F
Legay
D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology*; D057809:HEK293 Cells; D006801:Humans; D007124:Immunoenzyme Techniques / Q000379:methods*; D009980:Influenza A virus / Q000201:enzymology / Q000276:immunology*; D037102:Lectins / Q000737:chemistry; D051379:Mice; D008807:Mice, Inbred BALB C; D009439:Neuraminidase / Q000276:immunology*; D014764:Viral Proteins / Q000276:immunology*
D000914:Antibodies, Viral; D037102:Lectins; D014764:Viral Proteins; D009439:Neuraminidase