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Evaluation of solubilized herpes simplex virus membrane antigens in diffusion in gel-enzyme-linked immunosorbent assay (DIG-ELISA).

Cell membranes were prepared from herpes simplex virus (HSV) type 1-infected cells and solubilized with a low concentration of sodium deoxycholate. The supernatant after ultracentrifugation was used as antigen in a newly developed solid phase assay, diffusion in gel-enzyme-linked immunosorbent assay (DIG-ELISA). Antigen solubilization was almost complete, all HSV glycoproteins were represented, the yield of antigen in the solubilization process was high, and the presence of a detergent, sodium deoxycholate, did not interfere with the adsorption of antigen to the solid phase. DIG-ELISA was compared with the neutralization test for the determination of HSC antibodies and zone diameters showed a good correlation with the titre obtained by the neutralization test.

Animals↗

[Virus neutralization microreaction to determine antibodies against the infectious bovine rhinotracheitis virus].

Comparative studies were carried out with a total of 170 serum samples from cattle with the use of the virus-neutralization test and the virus-neutralization microreaction to demonstrate the presence of antibodies against the IBR virus. It was found that the two reactions were equally effective in terms of specificity. The coincidence of the results obtained with regard to the positive and negative tests was 97 per cent with variant alfa and 98 per cent with variant beta. Comparatively higher sensitivity was demonstrated with variant beta. It was established that the virus-neutralization microreaction was not influenced by the presence of antibodies against the virus in the initial cell cultures, it producing a well marked border line when reading the results. It was concluded that the virus-neutralization microreaction could in all cases be used instead of the virus-neutralization test as it is economical and is readily performed.

Animals↗

Immunoradiometric assay of inhibitors of antihaemophilic factor A.

An immunoradiometric assay (IRMA) for determination of antibodies against f. VIII:C in haemophilia A was developed. The assay was based on competitive binding of radiolabelled anti-VIII:C and antibodies in the test material to immobilized VIII:C. Fifteen haemophiliacs with known inhibitors were investigated with the new method and with a conventional neutralization test. In 3 cases the inhibitors were detected only with the IRMA and in the other 12 there was good agreement between the inhibitor levels found with the two methods. It was also possible to demonstrate the antibodies in three non-haemophilic patients with acquired inhibitors. The IRMA, which can detect the antibodies down to a concentration of 0.02 inhibitor units per ml, is more sensitive than conventional neutralization tests and is thus of practical importance in the investigation of patients with low inhibitor titres.

Antibodies↗

Development and evaluation of an indirect enzyme immunoassay for detection of porcine antibodies to pseudorabies virus.

An indirect enzyme immunoassay is described for detection of porcine serum antibody to pseudorabies virus. The analytical sensitivity of the enzyme immunoassay was found to be approximately 4.5 log 4 X 10 (5120 times) greater than the serum neutralization test, based on parallel end point titrations. The diagnostic sensitivity of the enzyme immunoassay was comparable or superior to that of the serum neutralization, based on the earliest detectable antibody after infection of swine with pseudorabies virus by intranasal or intrauterine routes or by contact with infected pigs. The enzyme immunoassay, at a screening dilution of 1:20, gave 100% agreement with ELISA results provided with a U.S. Department of Agriculture-Animal and Plant Health Inspection Service proficiency panel of 40 sera. One serum having demonstrable antibody by the enzyme immunoassay was seronegative by the serum neutralization test.

Animals↗

Three different serotypes of human rotavirus determined using an interference test with coxsackievirus B 1.

MA104 cells were used to titrate human rotavirus, and its capacity to render the cells resistant to coxsackievirus B 1 infection enabled us to determine easily the infectious titer of three prototype human rotaviruses. When rabbit antisera prepared against these prototype viruses were used for the neutralization test, the viruses could be discriminated from each other by a specific neutralization test.

Enterovirus B, Human↗

An enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies to bovine viral diarrhoea virus (BVDV) in cattle sera.

A microtitre ELISA has been established for the quantitation of antibodies to bovine viral diarrhoea virus (BVDV). Single dilutions of sera were assayed and units of antibody were calculated from a standard curve. In order to detect the maximum number of responding animals both IgG1 and IgG2 antibody should be assayed, although detection of IgG1 alone was nearly as effective. The ELISA was as sensitive as the virus neutralization test for detection of antibody; comparison of an ELISA that detected IgG1 plus IgG2 antibody to BVDV with the virus neutralization test gave a correlation coefficient (r) of 0.89 (P less than 0.001 for 95 compared sera). Although similar amounts of IgG1 and IgG2 antibodies were present in sera from both experimentally- and naturally-infected cattle, antibody to BVDV in colostrum and in the sera from young calves was predominantly IgG1. The number of adult cows with antibody was 40 out of 41 while 36 of 44 calves reared in a beef unit were found to have produced antibody by the time they were 31.5 weeks old, an indication of the high prevalence of BVDV in the cattle population.

Animals↗

Enzyme immunoassay for measurement of antibodies to herpes simplex virus infection: comparison with complement fixation, immunofluorescent-antibody, and neutralization techniques.

An enzyme immunoassay (EIA) was developed for detecting antibody to herpes simplex virus, and the results were compared with those of complement fixation, indirect immunofluorescent-antibody, and plaque reduction neutralization tests. Test sera showed very little nonspecific reactivity even at a starting dilution as low as 1:10. EIA results showed excellent correlation with results obtained by the neutralization test, with an average gain in sensitivity of 1.65. EIA proved very useful in detecting current herpes simplex virus infection, and antibody appeared in all cases soon after clinical onset. EIA appears to be a rapid, sensitive, and specific method for routine demonstration of herpes simplex virus antibody in a clinical setting.

Antibodies, Viral↗

Capsid mosaics of intermediate strains of human adenoviruses.

Antisera against isolated capsid components of intermediate adenovirus strains, types 3-16 (the San Carlos agent) and 15-9 and of "parental" prototype strains were compared in neutralization tests, hemagglutination-inhibition (HI) tests employing soluble and virion-associated hemagglutinin as antigens, and by electron microscopy. Hexons of the intermediate strains were found to be similar to, but not identical to, those of the prototype strains with which a cross-reaction occurred in neutralization tests (types 3 and 15). In contrast, fibers of intermediate strains displayed characteristics relating them to the corresponding components of prototype strains to which a relationship has been found in HI tests. Fibers (and possibly even pentons) of types 9 and 15-9 appeared to be identical, whereas fibers of types 3-16 and 16 displayed antigen specificities of both common and unique nature.

Adenoviridae↗

[Comparison of 4 technics for serologic titration of antibodies against rabies virus in dogs].

Four different serological techniques were used for the determination of antibody levels against rabies virus in 55 vaccinated street dogs: mouse neutralization test (reference), rapid fluorescent focus inhibition test, plaque reduction test and immunoenzymatic test (protein A). The results obtained with each of the last three methods were compared with those obtained with the reference test: correlation coefficients were, respectively, 0.810, 0.812 and 0.682, each being significantly correlated with the others. Therefore the three techniques may each be recommended as an alternative to the mouse neutralization test for routine titrations; the immunoenzymatic method, which titrates not only neutralizing antibodies being, nevertheless, better used as a screening test.

Animals↗

Detection of antibody against transmissible gastroenteritis virus of pigs by indirect immunoperoxidase antibody test.

Immunoperoxidase intibody (IPA) test was developed for detecting antibody against transmissible gastroenteritis (TGE) virus of pigs. The IPA antibody titers in sera collected in the field from 82 pigs were approximately seven times higher than those obtained in a serum-neutralization test. The correlation between the TGE antibody concentrations in the IPA and serum neutralization tests was positive (r = +0.74). The IPA tests appears to have the potential for routine laboratory use for serologic diagnosis of TGE.

Animals↗

The distribution and prevalence of group A arbovirus neutralizing antibodies among human populations in Southeast Asia and the Pacific islands.

Plaque reduction neutralization tests, using five group A arboviruses (chikungunya, Ross River, Getah, Bebaru and Sindbis), were done on sera from human populations in 44 Southeast Asia and Pacific island localities. Specificity of the plaque neutralization test was determined by examining convalescent sera from patients with known alphavirus infections. Chikungunya-specific neutralizing antibodies were demonstrated in sera of persons living in South Vietnam, Northern Malaysia, Indonesia (Kalimantan and Sulawesi), as well as Luzon, Marinduque, Cebu and Mindanao islands in the Philippines. Evidence of Ross River virus infection was found among populations living in West New Guinea and Papua New Guinea mainland, the Bismark Archipelago, Rossel Island and the Solomon Islands. There appeared to be no geographic overlap in the distribution of chikungunya and Ross River viruses, with the separation in their distribution corresponding with Weber's line in the Pacific. Sindbis neutralizing antibodies were found in 7 of 21 populations sampled, but in general the prevalence of infection was low. Four sera, from Vietnam, Malaysia and Mindanao gave monospecific reactions with Getah virus. No evidence of specific Bebaru virus infection was detected. The epidemiology of these five alphaviruses in Southeast Asia and the Pacific islands is discussed.

Adolescent↗

Studies concerning etiology of sheep-associated malignant catarrhal fever in Europe.

The etiological agent of sheep-associated malignant catarrhal fever (MCF) in Europe has not been isolated directly from sheep. The occurrence of antibodies against the African bovine herpesvirus (BHV-3, WC 11) in cattle and sheep was examined using recently modified indirect immunofluorescence test (IIFT) and neutralization test (NT) methods. Studies revealed that sheep and cattle sera in Europe were free from neutralizing antibodies of BHV-3. The IIFT could not establish the presence of antibodies to African BHV-3 in cattle but revealed that about 22.4% of sheep sera reacted to it. Apart from the well known ovine herpesvirus (BHV-5), occurrence of another herpesvirus in Europe had been expected. This virus is not identical with the WC 11 strain, but it is in antigenic relationship to it. We had for the first time substantial serological evidence to the effect that sheep-associated MCF in Europe is a herpesvirus related to the African strain.

Animals↗

Immunologic comparison of the proteins of pseudorabies (Aujeszky's disease) virus and bovine herpesvirus-1.

The immunologic relationship between bovine herpesvirus-1 and pseudorabies virus was examined by 80% serum cross-neutralization test, enzyme-linked immunosorbent assays, and Western immunoblotting procedures. Immunogenic cross reactivity between the 2 viruses was observed with both the serum-neutralization test and the enzyme-linked immunosorbent assay. A probing of viral Western immunoblots with rabbit hyperimmune antiserum showed that there were a number of viral-specific cross-reactive proteins between bovine herpesvirus-1 and pseudorabies virus.

Animals↗

[Immunological correlations between 2 strains of mucosal disease virus diarrhea (short report].

Cross-neutralization tests of the C24V strain and the "Kotschmar" strain provided evidence that the two strains are serological variants. Independenthy of quantitative serological differences, these strains had a better immunizing action in calves against a virulent strain of MD/VD virus. This virulent strain evidently formed a third serological variant, because when calves were infected with it, there was a decrease in the serological differences between antibodies. A practical conclusion is that diagnostic serological testing using only one strain is unlikely to detect all the infected animals, particularly those with low titres of antibody. In the presence of complement the neutralizing capacity of immune serum increased and, in the case of low antibody titres, serological differences between strains decreased. Therefore the complement-dependent neutralization test with at least two strains of virus is recommended for use in diagnostics.

Animals↗

Aujeszky's disease: blocking ELISA for the detection of serum antibodies.

A blocking ELISA for the detection of serum antibodies to Aujeszky's disease virus has been developed. The test was designed in particular with a view to examining large numbers of blood samples. It has been found to be sensitive, specific and precise. In comparison with the serum neutralization test it was superior in detecting low levels of antibodies in pig sera. The blocking ELISA gave higher titre values in serum samples taken early after experimental infection of pigs with Aujeszky's disease virus, than did the serum neutralization test.

Animals↗

[Application of single radial hemolysis technique for diagnosis of influenza A (H5N1)].

BACKGROUND: To understand the optimal condition of single radial hemolysis (SRH) for diagnosis of avian influenza A (H5N1) virus in order that SRH could be performed in general laboratories. METHODS: The effect of different concentration of virus and species of red blood cells, as well as kind and concentration of agarose on testing sensitivity of SRH was determined. Meanwhile the sensitivity and specificity of this method were compared with those of micro-neutralization test. RESULTS: The optimal condition of SRH included the viral concentration of 1000 HA units per 0.1 ml packed chicken red blood cells, the agarose concentration of 1.0%, the compliment added into agarose-virus-rbc slides after diffusion of sera. The sensitivity and specificity of SRH were very similar to those of micro-neutralization test. Meanwhile, no cross reaction between antibodies, especially antibodies against N1 antigens, H5N1 and H1N1 viruses was detected. CONCLUSION: The sensitivity and specificity of SRH were very similar to those of micro-neutralization assay. SRH could be performed in normal laboratories and be used for testing large scale serum samples.

Animals↗

Standardization of poliovirus neutralizing antibody tests.

Recently the Forum for Advancement of Immunization Research sponsored a Collaborative Study on Poliovirus Antibody Titration. Twenty laboratories from 12 countries participated. There were considerable differences in detail of test performance and test results among laboratories. The sensitivity of the tests varied over a 10-fold range (geometric mean titer from 21 to 288). The average coefficient of variation ranged from 4.5% to 13.5%. Tests performed at the Food and Drug Administration indicated that Hep-2 cells were more suitable than Vero cells for poliovirus titration by cytopathic effect end point or plaque assay. However, the cell lines were equally suitable for polio antibody titration by neutralization of cytopathic effect. A high degree of sensitivity and optimal reproducibility of antibody assay were observed in tests utilizing a medium to low dose of virus and serum-virus incubation overnight at 36 C. On the basis of current experience, a protocol is proposed for a test that provides high sensitivity and reproducibility and is practical for large-scale determinations of poliovirus antibody.

Animals↗

[Selective photoimmunoinactivation of influenza virus].

Antibodies to influenza virus were conjugated with hematoporphyrin. The conjugates were verified by the peak point identity in protein absorption (lambda-280 nm) of elution profiles and by the porphyrin-specific fluorescence. The antibody conjugates have been shown to preserve the specific antigenic activity tested by enzyme immunoassay. The conjugated hematoporphyrin nuclei have also preserved the photochemical activity since the antibody conjugates lose their native properties being exposed to visible light. In the neutralization test in the dark the conjugates of specific antibodies with hematoporphyrin decrease the viral cultures infectivity to the same extent as nonconjugated antibodies. This fact verifies antibodies stability in the conjugates. The infectivity of viral cultures exposed to visible light after neutralization test was further decreased by several orders. The data suggest undoubtedly perspective use of this method in viral infections control.

Antibodies, Monoclonal↗