Rapid focus reduction neutralization test of Japanese encephalitis virus in microtiter system

Okuno, Yoshinobu; Fukunaga, T.; Tadano, Masayuki; Okamoto, Yutaka; Ohnishi, Tsukasa; Takagi, M.

doi:10.1007/bf01314119

Cited by 79 publications

(71 citation statements)

References 5 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The haemagglutination inhibition (HI) tests were performed by a standard microtitre assay method [15] in 96 round-bottomed well microtitre plates (Nunc, Roskilde, Denmark). SAP was diluted in TBS-Ca 2þ to a final concentration of 3.2 mg/ml and 50 ml was transferred to the first well.…”

Section: Methodsmentioning

confidence: 99%

Serum Amyloid P Component Binds to Influenza A Virus Haemagglutinin and Inhibits the Virus Infection In Vitro

Andersen

Ravn²,

Sørensen³

et al. 1997

Scand J Immunol

View full text Add to dashboard Cite

Serum amyloid P component (SAP) is a member of the phylogenetically conserved and structurally related group of proteins called pentraxins. SAP exhibits multispecific calcium-dependent binding to oligosaccharides with terminal N-acetyl-galactosamine, mannose and glucuronic acid. The authors report that SAP can bind to influenza A virus and inhibit agglutination of erythrocytes mediated by the virus subtypes H1N1, H2N2 and H3N2. SAP also inhibits the production of haemagglutinin (HA) and the cytopathogenic effect of influenza A virus in MDCK cells. The binding of SAP to the virus requires physiological calcium concentrations and is blocked by specific SAP antibodies. Denaturated and renaturated SAP retained inhibition of HA. Electron microscopy shows Ca 2þ -dependent binding of SAP to spikes on the viral envelope and immunoblotting indicates that SAP binds to a 50-55 kDa peptide corresponding to the mass of the HA1 peptide. Of several monosaccharides tested only D-mannose interfered with SAP's inhibition of both HA and infectivity. The glycosaminoglycans heparan sulfate and heparin, which bind SAP, reduced SAPs binding to the virus. The results indicate that the inhibition by SAP is due to steric effects when SAP binds to terminal mannose on oligosaccharides localized close to the sialic acid-binding site of the HA trimer.

show abstract

Section: Methodsmentioning

confidence: 99%

Serum Amyloid P Component Binds to Influenza A Virus Haemagglutinin and Inhibits the Virus Infection In Vitro

Andersen

Ravn²,

Sørensen³

et al. 1997

Scand J Immunol

View full text Add to dashboard Cite

show abstract

“…N test. Fifty percent focus reduction N test on BHK-21 cells using peroxidase-anti-peroxidase staining method was used (14). For the test, BHK-21 cells cultured on a 96-well microplate were used (9).…”

mentioning

confidence: 99%

Studies on Serological Cross‐Reaction in Sequential Flavivirus Infections

Makino

Tadano

Saito

et al. 1994

Microbiology and Immunology

View full text Add to dashboard Cite

Acute-and convalescent-phase sera from patients with dengue (DEN) hemorrhagic fever (DHF) and Japanese encephalitis (JE) that contained pre-existing flavivirus antibodies were tested for cross-reacting antibodies to DEN, JE and yellow fever (YF) viruses by a neutralization (N) test . A fourfold or greater rise in N antibody titer in the convalescent-phase was considered significant . Of 39 DHF cases, obtained at Chiang Mai University Hospital, Thailand, 15 (38.5%) showed a rise in DEN antibody titer, while another 15 (38.5%) showed a significant rise in both DEN and JE N antibody titers. On the other hand, eight (61.5%) of 13 JE cases obtained at the same Hospital , showed a significant rise in JE antibody titer, while two (15.4%) showed a significant rise in both DEN and JE antibody titers. Sucrose gradient centrifugation and fractionation of these two crossreactive JE sera revealed that IgM class antibody was specific for JE , while IgG class antibody was cross-reactive. Of three JE cases with pre-existing YF antibody obtained in Okinawa , Japan, two showed a significant rise in YF and JE antibodies. Both IgM and IgG class antibodies to YF virus were elevated. These results indicate that the cross-reactivity among flaviviruses in different subgroups (complexes), was observed quite often, even by the N test , in sequential flavivirus infection.Key words: Flaviviruses, Sequential infections, Cross-neutralization testThere are more than 60 flaviviruses , including Japanese encephalitis (JE), dengue (DEN) and yellow fever (YF) viruses, and contain crossreactive antigens which make serodiagnosis difficult (11 for review). This is especially true in the regions where two or more flaviviruses are prevalent . Among the serological tests, the neutralization (N) test is considered to be more specific than complement-fixation (CF), hemagglutination-inhibition (HI) and enzyme-linked immunosorbent assay (ELISA) (16 for review). On the basis of close relationship in cross-neutralization tests using hyperimmune polyclonal antisera, flaviviruses are divided into at least eight antigenic subgroups (complexes), where JE and DEN viruses are classified into different subgroups, while YF virus remains unassigned (I).In Japan, only one flavivirus , i.e. JE virus, has been prevailing and HI test or ELISA has been employed for serodiagnosis without problems. While in the Southeast Asian countries such as Thailand and Vietnam, JE and DEN viruses have been coexisting and the outbreaks caused by these viruses have taken place simultaneously in the rainy season (6). It has been reported that in the secondary DEN virus infection, the serotype of the causative virus of the recent infection could not be identified by N test because of the cross-reactions . It was possible only by virus isolation or detection of specific viral genomic sequence (13). Secondary DEN infection in previously immune individuals with another serotype of DEN virus elicits broadly

show abstract

“…Neutralizing activity of Fab preparations or a purified Fab molecule was determined by focus reduction neutralization test using JEV strain Nakayama on baby hamster kidney (BHK)-21 cells (24,29). Briefly, 70 µl of viral stock containing approximately 40 to 50 focus forming units (ffu) of JEV strain Nakayama was incubated with an equal volume of Fab preparation or a serially diluted purified Fab molecule or PBS (virus control) at room temperature for 90 min, then added to BHK-21 cells cultured in 96-well flatbottom plates.…”

Section: Methodsmentioning

confidence: 99%

Construction of Human Fab (Y1/K) Library and Identification of Human Monoclonal Fab Possessing Neutralizing Potency against Japanese Encephalitis Virus

Arakawa

Yamashiro

Uechi

et al. 2007

Microbiology and Immunology

View full text Add to dashboard Cite

A combinatorial human Fab library was constructed using RNAs from peripheral blood lymphocytes obtained from Japanese encephalitis virus hyper‐immune volunteers on pComb3H phagemid vector. The size of the constructed Fab library was 3.3 × 108 Escherichia coli transformants. The library was panned 3 times on the purified Japanese encephalitis virus (JEV) virion, and phage clones displaying JEV antigen‐specific Fab were enriched. The enriched phage pool was then screened for clones producing Fab molecule with JEV neutralizing activity by the focus reduction‐neutralizing test. Among 188 randomly selected clones, 9 Fab preparations revealed neutralizing activities against JEV strain Nakayama. An E. coli transformed with TJE12B02 clone, which produced human monoclonal Fab with the highest neutralizing activity was cultured in a large scale, and the Fab molecule was purified using affinity chromatography. The purified FabTJE12B02 showed the 50% focus reduction endpoint at the concentration of 50.2 μg/ml (ca. 1,000 nM) when JEV strain Nakayama was used. The FabTJE12B02 recognized E protein of JEV strain Nakayama, and the dissociation equilibrium constant (Kd) of the FabTJE12B02 against purified JEV antigen was calculated as 1.21 × 10–8 M. Sequence analysis demonstrated that TJE12B02 used a VH sequence homologous to the VH3 family showing 88.8% homology to germline VH3–23, and used a VK sequence homologous to the VκII subgroup showing 92.8% homology to germline A17.

show abstract

Rapid focus reduction neutralization test of Japanese encephalitis virus in microtiter system

Cited by 79 publications

References 5 publications

Serum Amyloid P Component Binds to Influenza A Virus Haemagglutinin and Inhibits the Virus Infection In Vitro

Serum Amyloid P Component Binds to Influenza A Virus Haemagglutinin and Inhibits the Virus Infection In Vitro

Studies on Serological Cross‐Reaction in Sequential Flavivirus Infections

Construction of Human Fab (Y1/K) Library and Identification of Human Monoclonal Fab Possessing Neutralizing Potency against Japanese Encephalitis Virus

Contact Info

Product

Resources

About