TY - JOUR
T1 - Second generation of pseudotype-based serum neutralization assay for Nipah virus antibodies
T2 - Sensitive and high-throughput analysis utilizing secreted alkaline phosphatase
AU - Kaku, Yoshihiro
AU - Noguchi, Akira
AU - Marsh, Glenn A.
AU - Barr, Jennifer A.
AU - Okutani, Akiko
AU - Hotta, Kozue
AU - Bazartseren, Boldbaatar
AU - Fukushi, Shuetsu
AU - Broder, Christopher C.
AU - Yamada, Akio
AU - Inoue, Satoshi
AU - Wang, Lin Fa
PY - 2012/1
Y1 - 2012/1
N2 - Nipah virus (NiV), Paramyxoviridae, Henipavirus, is classified as a biosafety level (BSL) 4 pathogen, along with the closely related Hendra virus (HeV). A novel serum neutralization test was developed for measuring NiV neutralizing antibodies under BSL2 conditions using a recombinant vesicular stomatitis virus (VSV) expressing secreted alkaline phosphatase (SEAP) and pseudotyped with NiV F/G proteins (VSV-NiV-SEAP). A unique characteristic of this novel assay is the ability to obtain neutralization titers by measuring SEAP activity in supernatant using a common ELISA plate reader. This confers a remarkable advantage over the first generation of NiV-pseudotypes expressing green fluorescent protein or luciferase, which require expensive and specific measuring equipment. Using panels of NiV- and HeV-specific sera from various species, the VSV-NiV-SEAP assay demonstrated neutralizing antibody status (positive/negative) consistent with that obtained by conventional live NiV test, and gave higher antibody titers than the latter. Additionally, when screening sixty-six fruit bat sera at one dilution, the VSV-NiV-SEAP assay produced identical results to the live NiV test and only required a very small amount (2μl) of sera. The results suggest that this novel VSV-NiV-SEAP assay is safe, useful for high-throughput screening of sera using an ELISA plate reader, and has high sensitivity and specificity.
AB - Nipah virus (NiV), Paramyxoviridae, Henipavirus, is classified as a biosafety level (BSL) 4 pathogen, along with the closely related Hendra virus (HeV). A novel serum neutralization test was developed for measuring NiV neutralizing antibodies under BSL2 conditions using a recombinant vesicular stomatitis virus (VSV) expressing secreted alkaline phosphatase (SEAP) and pseudotyped with NiV F/G proteins (VSV-NiV-SEAP). A unique characteristic of this novel assay is the ability to obtain neutralization titers by measuring SEAP activity in supernatant using a common ELISA plate reader. This confers a remarkable advantage over the first generation of NiV-pseudotypes expressing green fluorescent protein or luciferase, which require expensive and specific measuring equipment. Using panels of NiV- and HeV-specific sera from various species, the VSV-NiV-SEAP assay demonstrated neutralizing antibody status (positive/negative) consistent with that obtained by conventional live NiV test, and gave higher antibody titers than the latter. Additionally, when screening sixty-six fruit bat sera at one dilution, the VSV-NiV-SEAP assay produced identical results to the live NiV test and only required a very small amount (2μl) of sera. The results suggest that this novel VSV-NiV-SEAP assay is safe, useful for high-throughput screening of sera using an ELISA plate reader, and has high sensitivity and specificity.
KW - Biosafety level 4
KW - Henipavirus
KW - Neutralization
KW - Pseudotyped virus
KW - Secreted alkaline phosphatase
UR - http://www.scopus.com/inward/record.url?scp=84855226612&partnerID=8YFLogxK
U2 - 10.1016/j.jviromet.2011.11.003
DO - 10.1016/j.jviromet.2011.11.003
M3 - Article
C2 - 22115786
AN - SCOPUS:84855226612
SN - 0166-0934
VL - 179
SP - 226
EP - 232
JO - Journal of Virological Methods
JF - Journal of Virological Methods
IS - 1
ER -