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PubMed1980

Hemagglutination by canine parvovirus: serologic studies and diagnostic applications.

Conditions for canine parvoviral hemagglutination (HA) and hemagglutination-inhibition (HI) reactions were defined. The HA phenomena were used to differentiate canine parvovirus (CPV) from feline panleukopenia virus (FPV), mink enteritis virus (MEV), and minute virus of canines. Serologic comparisons of the CPV, FPV, and MEV by HA-HI and serum-neutralization tests indicated that CPV, FPV, and MEV were antigenically similar but were different from minute virus of canines. Diagnostic application of HA tests to fecal samples from acute cases of enteritis was discussed. Combinating HA tests with HI tests on fecal samples provided a rapid and specific diagnostic method for CPV infection. Secular seroprevalence studies indicated the emergence of CPV infeciton in the United States dog population-at-large in 1978.

Animals↗
PubMed1983

[Isolation of adenoviruses from calves with bronchopneumonia].

The virologic investigation of clinical and pathological material from calves affected with a respiratory disease at the age of 30 days to 10 months on 16 infected stock-breeding farms led to the isolation of two virus strains (D-117 and Ch-216) on primary cell cultures of fetal calf kidney from the nasal discharge of two 8-month-old calves. By their cytopathic changes, physico-chemical properties and the presence of a group-specific antigen they were said to belong to the adenovirus family. The results of the cross virus-neutralization test pointed to the assumption that they had to be referred to the bovine adenovirus type 1. Discussed is the problem concerning the participation and the importance of individual types of adenoviruses in the acute respiratory enzootics of grown-up calves.

Adenoviridae↗
PubMed1986

Antigenic comparison of envelope protein E between Japanese encephalitis virus and some other flaviviruses using monoclonal antibodies.

The antigenic relationships between Japanese encephalitis (JE) virus and several other flaviviruses have been investigated using anti-JE virus monoclonal antibodies (MAbs). Seventeen MAbs directed against envelope protein E of JE virus were characterized and divided into eight MAb groups based on reactivity patterns in haemagglutination inhibition test, neutralization (N) test, ELISA and competitive binding assay with JE virus. The results suggest the existence of at least eight epitopes on the E protein of JE viruses. Analysis of cross-reactivity of the antibodies with several other flaviviruses indicated these findings JE virus, belonging to West Nile (WN) subgroup, is antigenically closely related to viruses in the same subgroup, i.e. Murray Valley encephalitis (MVE), WN and St. Louis encephalitis (SLE) viruses. Of these three viruses, JE virus has the closest relationship with MVE virus. WN virus is relatively close to JE virus, whereas SLE virus is the least closely related. Dengue viruses types 1 and 2, which belong to another subgroup of flaviviruses, show markedly less antigenic homology to JE virus. One of the critical N sites on the E protein showed JE virus specificity. Some cross-reactive antibodies which did not neutralize JE virus showed low but significant N activity against several other flaviviruses. Mixtures of several MAbs, which showed different reactivity patterns, potentiated the N activity against not only JE virus but also other members of the WN subgroup of flaviviruses, namely MVE, WN and SLE viruses.

Antibodies, Monoclonal↗
PubMed2004

Allele excess at neutrally evolving microsatellites and the implications for tests of neutrality.

Skews in the observed allele-frequency spectrum are frequently viewed as an indication of non-neutral evolution. Recent surveys of microsatellite variability have used an excess of alleles as a statistical approach to infer positive selection. Using neutral coalescent simulations we demonstrate that the mean numbers of alleles expected under the stepwise-mutation model and infinite-allele model deviate from the observed numbers of alleles. The magnitude of this difference is dependent on the sample size, mutation rates (theta-values) and observed gene diversities. Moreover, we show that the number of observed alleles differs among loci with the same observed gene diversity but different mutation rates (theta-values). We propose that a reliable test statistic based on allele excess must determine the confidence interval by computer simulations conditional on the observed gene diversity and theta-values. As the latter are notoriously difficult to obtain for experimental data, we suggest that other statistics, such as lnRV, may be better suited to the identification of microsatellite loci subject to selection.

Biological Evolution↗
PubMed1996

[Titration of rabies antibodies with the rapid fluorescence focus inhibition test].

Antirabies virus neutralization antibodies in sera and/or transudates modified RFFIT method by Smith et al. (1973). Sera were titrated on Lab-Tek 8 chamber TC slides. Sera and/or transudates (content of pleural cavity) as well as the challenge virus strain (vaccination strains of the rabies virus Vnukovo-32/107th passage and/or CVS 11/Paris) were incubated at 37 degrees C during 90 minutes subsequently BHK-21/C13 cell culture was added. The cultures were fixed after 24 to 48 hours and stained with antirabic fluorescent conjugate (Bioveta a.s., Ivanovice n. H., Czech Republic). The highest dilution of the virus was used as the challenge dose where 50 percent of the cells in the examined range of view were infected (fluorescent inclusions can be observed). The antirabic reference serum was used as a control in RFFIT in each examined serum. To ensure a good control, the serum was diluted to contain 0.5 IU/ml of antirabic virus neutralization antibodies. Sera and/or transudates which were sent to our Laboratory were examined in this way. We examined 40 sera or pleural transudates of orally vaccinated foxes by those methods. These sera were sent to National reference laboratories for rabies (NRPB) in Kosice. Samples were examined for the monitoring of efficiency of oral antirabic vaccination. The parallel quantification of antirabic antibodies by virus neutralization test (VNT) in vivo was applied to mice and indirect haemagglutination test (NHT). The results of these three tests are comparable or in correlation. RFFIT has many advantages. When using highly attenuated strain Vnukovo-32/107th passage as the challenge virus in RFFIT method the potential risk of laboratory exposition is absent.

Animals↗
PubMed1982

Collagen-anti-collagen complexes in rheumatoid arthritis sera.

Immune complexes (IC) were detected in rheumatoid sera by means of anti-antibody (AA) neutralization tests. The sera with rheumatoid factor (RF) that did not agglutinate human Rh+ erythrocytes sensitized by incomplete Rh antibodies, Ripley, could be tested in an AA neutralization test without any further treatment. On the other hand, 'Ripley-positive' sera had to be treated with 2-mercaptoethanol or dithiothreitol in order to destroy RF. IC-containing sera were further studied for dispersion of IC by collagen. Surprisingly, roughly 50% of IC-containing sera were affected by collagen in that they lost AA-neutralizing activity completely or partially. Significantly, very similar results were obtained with collagen type I, collagen type II and denatured collagen type II. It was concluded that these sera contained IC formed by denatured collagen and its antibodies. Further experiments were devoted to the study of the effect of collagen on reactions of RF with Ripley-sensitized erythrocytes. In several sera, the addition of collagen eliminated the prozone. This was interpreted as deblocking of RF due to the dispersion of the blocking IC by an excess of collagen. In other sera, the addition of collagen seemed to remove or block RF. This blocking effect was interpreted as a neutralization of multispecific RF by collagen. An alternative explanation proposed that collagen formed IC with free anti-collagen antibodies and that these IC blocked the RF.

Antibodies, Anti-Idiotypic↗
PubMed1970

Serological survey for arboviruses in Uganda, 1967-69.

In a serological survey for arboviruses carried out in 8 areas of Uganda between 1967 and 1969, altogether 1869 human sera were tested with 8 antigens by the haemagglutination-inhibition (HI) test. Mouse neutralization tests (NT) with Group-B viruses were also performed on selected sera.The prevalence of arbovirus antibodies was found to vary considerably between the different areas. Very few people in the highland area of Kigezi had antibody to any of the antigens used, whereas more than half of the sera from the Madi area near the Nile had antibody to several antigens. The chikungunya antibody prevalent in the West Nile, Madi and Bwamba areas is thought probably to represent previous infection with the cross-reacting o'nyong-nyong virus. Bunyamwera antibody was also highly prevalent in Madi (66.7% of children's sera and 81.5% of adult sera).The combined HI and NT results indicate that yellow fever continues to be infrequent in Uganda (only 31 sera had yellow fever neutralizing antibody); there is no evidence of recent yellow fever infection in Bwamba, an area in which this disease was previously considered endemic.

Adolescent↗
PubMed1985

Antiviral neutralizing antibody to Hantaan virus as determined by plaque reduction technique.

The 76--118 strain of Hantaan virus was titrated in E6 (Vero) cells by the plaque method using agarose overlay medium. Visible plaques, formed 10 days post-infection, were uniformly 2--3 mm in diameter. Dose-response experiments showed that a single infectious particle initiated the formation of a plaque. Infectivity titers by the plaque method were equivalent to those obtained by the endpoint method (TCID50) using the immunofluorescence antibody technique (IFA) for antigen detection. The single-cycle growth pattern of the virus showed an eclipse phase of 7 to 9 hours, with production of cell-free infectious virus 18 hours post-infection. Plaque reduction neutralization tests suggested that complement enhanced the neutralizing activity of sera; rat sera were particularly complement-dependent. The plaque reduction neutralization test was about 10 times more sensitive than the TCID50 neutralization test. Convalescent phase sera from patients with hemorrhagic fever with renal syndrome (HFRS) having higher IF antibody titers to Hantaan virus than to nephropathia epidemica (NE) virus were capable of neutralizing Hantaan virus, while sera from patients with higher IF antibody titers to NE virus than Hantaan virus did not contain neutralizing antibody to Hantaan virus.

Animals↗
PubMed1993

Serological response to Japanese encephalitis vaccine in a group of school children in South Arcot district of Tamil Nadu.

A trial with Biken Japanese encephalitis (JE) vaccine made in Japan was carried out in South Arcot district of Tamil Nadu state, India. A total of 113 school children were included in the trial. The efficacy (as determined by serological response) and safety of the vaccine were evaluated. Side effects, though minor, were noted in 54.9 per cent of the children after each dose. The serum antibody titres were determined by mouse neutralization test, plaque reduction neutralization test and haemagglutination inhibition test. An antibody response to two-dose primary vaccination schedule was observed in 72.7 per cent, whereas 87.8 per cent of the vaccines responded positively after the booster dose administered one year after. Only about 20 per cent of the children had persisting antibodies one year after the primary vaccination. The results indicated a probable need of the third dose in the primary vaccination schedule.

Adolescent↗
PubMed1975

A comparison of some immunological methods for the differentiation of strains of foot-and-mouth disease virus.

Two FMDV strains which had been previously differentiated by complement-fixation were compared by guinea-pig protection test, kinetic neutralization and micro-neutralization tests. It was found that these tests, which have not been previously applied by the methods described, were all capable of FMDV strain differentiation. Similar differences were found by all methods, which suggests that comparisons made by cross-CF, cross-neutralization or cross-protection involve measurement of the same antigen/antibody interactions.

Animals↗
PubMed1983

Persistence of antibodies and anamnestic response in calves vaccinated with inactivated infectious bovine rhinotracheitis virus and parainfluenza-3 virus vaccines.

Persistence of antibodies in calves vaccinated with 2 types of inactivated infectious bovine rhinotracheitis (IBR) virus and parainfluenza-3 (PI-3) virus vaccines were determined. Calves seronegative for IBR and PI-3 viruses were inoculated with 2 doses of inactivated IBR virus-PI-3 virus vaccines administered 2 weeks apart. Blood samples were obtained from the calves for serum at 2 weeks, 6 months, and 1 year after vaccination. The serums were tested by serum-neutralization tests. Antibody response to the vaccines persisted on a declining scale for 1 year. The anamnestic responses to the vaccines were determined by inoculating the same calves with a booster dose of vaccine 1 year after the original 2 doses were given. Blood samples were obtained from the calves for serum 2 weeks later. The serums were tested by serum-neutralization tests. The single booster dose of vaccine elicited an anamnestic response to both IBR and PI-3 viruses.

Animals↗
PubMed2005

Insufficient neutralization in testing a chlorhexidine-containing ethanol-based hand rub can result in a false positive efficacy assessment.

BACKGROUND: Effective neutralization in testing hand hygiene preparations is considered to be a crucial element to ensure validity of the test results, especially with the difficulty to neutralize chlorhexidine gluconate. Aim of the study was to measure the effect of chemical neutralization under practical test conditions according to EN 1500. METHODS: We have investigated two ethanol-based hand rubs (product A, based on 61% ethanol and 1% chlorhexidine gluconate; product B, based on 85% ethanol). The efficacy of products (application of 3 ml for 30 s) was compared to 2-propanol 60% (v/v) (two 3 ml rubs of 30 s each) on hands artificially contaminated with Escherichia coli using a cross-over design with 15 volunteers. Pre-values were obtained by rubbing fingertips for 1 minute in liquid broth. Post-values were determined by sampling immediately after disinfection in liquid broth with and without neutralizers (0.5% lecithin, 4% polysorbate 20). RESULTS: The neutralizers were found to be effective and non-toxic. Without neutralization in the sampling fluid, the reference disinfection reduced the test bacteria by 3.7 log10, product B by 3.3 log10 and product A by 4.8 log10 (P = 0.001; ANOVA). With neutralization the reference disinfection reduced the test bacteria by 3.5 log10, product B by 3.3 log10 and product A by 2.7 log10 (P = 0.011; ANOVA). In comparison to the reference treatment Product B lead to a lower mean reduction than the reference disinfection but the difference was not significant (P > 0.1; Wilcoxon-Wilcox test). Without neutralizing agents in the sampling fluid, product A yielded a significantly higher reduction of test bacteria (4.8; P = 0.02) as compared to the situation with neutralizing agents (2.7; P = 0.033). CONCLUSION: The crucial step of neutralization lies in the sampling fluid itself in order to stop any residual bacteriostatic or bactericidal activity immediately after the application of the preparation, especially with chlorhexidine gluconate-containing preparations. This is particularly important at short application times such as the 30 s.

Chlorhexidine↗
PubMed1973

Neutralization studies with Marek's disease virus and turkey herpesvirus.

Use of Marek's disease virus (MDV) in a neutralization test presents several problems, which are described, making this potentially useful test difficult. To obviate these difficulties, a plaque reduction test has been designed based on cross-neutralization of turkey herpesvirus (HVT) by serum-neutralizing MDV. The technique for such a neutralization test is outlined. Kinetics of development of neutralizing antibodies in chickens inoculated with HVT and MDV are described. The neutralization test can be used to evaluate viability of HVT vaccines and the possible role of neutralizing antibodies in the protection afforded by vaccination against MDV-induced tumors.

Animals↗
PubMed1986

Global survey of antibody to Hantaan-related viruses among peridomestic rodents.

A global serological survey of rodents was conducted to determine the distribution and prevalence of antibody to Hantaan-related viruses, which are the causative agents of haemorrhagic fever with renal syndrome (HFRS) in man. Over 1700 rodent sera from more than 20 sites worldwide were examined by immunofluorescent antibody assay. High-titred positive sera were further tested by plaque reduction neutralization tests with prototype Hantaan virus and urban rat-associated Hantaan-like virus. Antibody-positive rodents were found in most, but not all, sites sampled. The highest antibody prevalence rates were found in Baltimore, MD, USA and Bélem, Brazil, and Rattus norvegicus was the species most often found positive. Bandicota indica and B. bengalensis, species previously not recognized as hosts of hantaviruses, were also positive. Neutralization tests detected antibody in Rattus sera specific for urban rat-associated Hantaan-like virus, but failed to establish the specificity of antibody in Bandicota sera. These results indicate that Hantaan-related viruses exist beyond the currently recognized boundaries of HFRS in man and suggest that human HFRS-like disease might be occurring in other areas of the world where rodent-human contact is common.

Africa↗
PubMed1973

Guinea pig herpes-like virus infection. I. Antibody response and virus persistence in guinea pigs after experimental infection.

Guinea pigs, experimentally infected with guinea pig herpes-like virus produced antibodies which were detectable by indirect hemagglutination (IHA), complement-fixation (CF), and neutralization tests. The IHA test appeared to be a more sensitive method than the CF or neutralization test for determining antibody response in guinea pigs immediately after infection, but the IHA method was not suitable for the detection of antibody in animals receiving small doses of virus. Antibody titers obtained by CF tests were generally higher than those obtained by the neutralization test, and they followed the same time course when individual animals were studied serially. Intracardiac inoculation produced the best antibody response in guinea pigs when compared with other routes of infection. Guinea pigs infected by the intraperitoneal, intranasal, or oral route showed rising antibody titers but the levels were low. Infectious virus was isolated from and persisted in all inoculated animals in the presence of antibody regardless of the route of inoculation. Recovery of infectious virus required cultivation or cocultivation of tissue cells containing virus. The administration of antilymphocyte sera delayed the appearance of IHA antibody but had no effect on antibodies determined by the CF- and neutralization tests.

Animals↗
PubMed2006

Development of passive haemagglutination (PHA) and haemagglutination inhibition (HAI) technique for potency estimation of Cobra Antisnake Venom Serum (ASVS).

Antibodies against snake venom or antivenom potency are assayed quantitatively by in-vivo neutralization test in mice, which requires large number of laboratory animals. In potency assays of biological substances such as antivenoms, it is highly desirable to avoid suffering and death of animals by substituting in-vivo method with in-vitro methods, provided such methods measure life-saving capability with precision similar to that of in-vivo method. The in-vitro tests determine the neutralizing power of antivenom by permitting the evaluation of a particular biological activity of the venom and its neutralization after mixing the venom with the antivenom [Theakston RDG, Reid HA. Development of simple standard assay procedures for the characterization of snake venom. Bull WHO 1983;61:949-956; Gutierrez JM, Rojas G, Lomonte B, Gene JA, Chaves F, Alvarado J, et al. Standardizing of assays for testing the neutralizing ability of antivenoms. Toxicon 1990;28:1127-1129; Theakston R.D.G. Comments on letter of Gutierrez et al. on standardization of assays for testing the neutralizing ability of antivenoms. Toxicon 1990;28:1131-1132; Harvey AL, Barfaraz A, Thomson E, Faiz A, Preston S, Harris JB. Screening of snake venom for neurotoxic and myotoxic effects using simple in-vitro preparation from rodents and chicks. Toxicon 1994;32:257-265; World Health Organization Progress in characterization of venom and standardization of anti-venoms. Geneva: WHO offset publication; 1981. p. 58.]. Hence, the ideal requirements for an assay in detecting venom and venom antibody include high level of sensitivity, specificity (ability to differentiate between venom and venom antibody produced by closely related species of snakes), reproducibility and simplicity. A new in-vitro procedure for quantitative analysis of potency of ASVS by passive haemagglutination (PHA) and haemagglutination inhibition (HAI) has been explored. The methods described are simple, rapid, economical, reproducible and useful in replacing the more expensive in-vivo neutralization assays. Moreover, it also eliminates the use of laboratory animals.

Animals↗
PubMed1982

Experimental maedi virus infection in sheep: early cellular and humoral immune response following parenteral inoculation.

The early immune response (19 weeks) in sheep inoculated parenterally with maedi virus was studied, using the lymphocyte transformation test, immunodiffusion test, complement-fixation test, and neutralization test. Cellular immune responses were detected between 3 and 7 weeks after the sheep were inoculated. Antibodies were detected by immunodiffusion test in 3 of 5 animals in weeks 7 to 11 and by complement-fixation test in weeks 15 to 19. Neutralizing antibodies were not detected. There was no sign of virus-induced immunosuppression as determined by lymphocyte responsiveness to mitogens or by in vivo and in vitro immune responses following BCG vaccination.

Animals↗