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An assay method for herpes simplex virus type 1 seroprevalence survey--detection of antibody in saliva by avidin-biotin complex enzyme-linked immunosorbent assay (ABC-ELISA).

To determine whether the avidin-biotin complex enzyme-linked immunosorbent assay (ABC-E) is a potentially useful method for detection of herpes simplex virus type 1 (HSV-1) antibody in saliva, paired serum and saliva samples from 129 healthy individuals aged 18 to 25 years were collected simultaneously and subjected to a neutralization test (NT) for neutralizing antibody and also to an indirect ELISA (IE) and ABC-E for HSV-1 specific IgG detection. Compared with the results of NT, the sensitivities of the IE and ABC-E for serum were both 100% (45/45), and for saliva 82.2% (37/45) and 93.3% (42/45), respectively. The specificity of all these methods was 100% (84/84). With the same ABC-E method, a significant correlation (r = 0.66, P < 0.001) between the OD-difference (d-OD) values of positive serum and saliva samples was observed. Furthermore, the consistency of ABC-E for salivary antibody detection was confirmed with the paired serum and saliva samples which were collected from four individuals followed up for eight months. It was clear that the ABC-E method for saliva can be used in place of the NT and ABC-E method for serum for seroprevalence studying of HSV-1 infection.

Adolescent↗

Construction of mutant genes for a non-toxic verotoxin 2 variant (VT2vp1) of Escherichia coli and characterization of purified mutant toxins.

The gene encoding a Verotoxin 2 variant, VTvp1, was mutated by oligonucleotide-directed site-specific mutagenesis. Among 6 mutant toxins encoded by the mutated genes, E167Q-R170L (glutamic acid at position 167 and arginine at position 170 from N-terminus of the A subunit were replaced by glutamine and leucine, respectively) was found to have markedly decreased activities; inhibition of protein synthesis, Vero cell cytotoxicity and mouse lethality of the purified E167Q-R170L were 1/1,900, 1/125,000 and 1/2,000, respectively, of those of the purified wild-type VT2vp1. Since the antigenic property of the E167Q-R170L was demonstrated to be similar to that of the wild-type VT2vp1 by Ouchterlony double gel diffusion test and by neutralization test of Vero cell cytotoxicity of the VT2vp1, a possibility to use the mutant VT2vp1, E167Q-R170L, as a toxoid is discussed.

Animals↗

Evaluation of a semiautomated latex agglutination test for the detection of pseudorabies antibody in swine sera.

Rapid screening of large numbers of swine sera for antibody is an essential element in the current eradication program for pseudorabies in the United States. We evaluated a recently introduced commercial semiautomated latex agglutination test (LAT) kit for pseudorabies antibody. A total of 1,191 swine sera were tested using the new procedure and 3 established tests: the manual LAT, the serum neutralization test (SNT), and an enzyme-linked immunosorbent assay (ELISA). There was close agreement among results of semiautomated LAT, the manual LAT, and ELISA but less agreement between semiautomated LAT and SNT. Overall, the sensitivities of the 4 tests were as follows: semiautomated LAT = manual LAT > ELISA > SNT. For 74 samples of known pseudorabies antibody status, the overall specificities were as follows: semi-automated LAT = manual LAT = SNT > ELISA. Because of its relative insensitivity, the SNT should no longer be considered the official "gold" standard for pseudorabies testing in the on-going eradication program. However, because no single test was perfect for all samples, a scheme including 3 tests in the following sequence is recommended: 1) screening using semiautomated LAT or ELISA and 2) confirmation testing of positives by manual LAT and SNT, with any sample that tests positive by any 2 tests being regarded as true positive.

Animals↗

Prevalence of Clostridium botulinum type C in substrates of phosphate-mine settling ponds and implications for epizootics of avian botulism.

Prevalence and conditions for occurrence of Clostridium botulinum type C were examined on phosphate-mine settling ponds and a natural wetland in northern Florida between April 1981 and March 1982. Substrate samples were collected monthly (winter) and semi-monthly (summer) from 16 locations on seven ponds. Selected environmental parameters were measured at each location at the time of sampling. Mouse inoculation tests and toxin neutralization tests using enrichment culture filtrates were conducted to identify C. botulinum type C in the samples. The bacteria were identified in 26 (5.6%) of 467 sediment samples. Occurrences were distributed over four of the seven ponds and included nine of the 16 sample locations, but were restricted to the months April through October. The organism occurred over a wide range of ecological conditions found on the ponds during these months. If the presence of C. botulinum type C in the substrate is a prerequisite for botulism to occur, the prevalence and fairly wide distribution of this organism on settling ponds makes it difficult to predict where future outbreaks may occur.

Animals↗

[Role of antineuraminidase antibodies in protection against influenza].

For improving the anti-influenza vaccination efficacy, the choice of strains carrying up dated neuraminidase antigen (NA) and the introduction of the optimal amount of NA antigen in the vaccine are critical. Monoclonal antibodies prepared against the neuraminidase N2 of A/Beijing/32/92 showed NA inhibition (NI) and neutralized (Nt) the cells infection by influenza virus either at an early stage (group 2 antibodies inhibit virus binding to cells) or at a late stage of infection (group 1 antibodies inhibit virus release). The specificity of the neutralization test is restricted to the homologous variant whereas the NI specificity is much broader. When both group 1 and group 2 antibodies are tested together, their neutralizing activity is significantly increased. The emergence in 1997 of an avian strain H5N1 in humans influenza infections at Hong Kong (Strain A/Hong-Kong/156/97) rose the threat of pandemic. The H5N1 strain carried H5 HA which is not recognized by the human immune system, but N1 might be related to other N1 antigens belonging to avian, swine and human strains. So we 1) characterized the N1 antigen from H5N1 in comparison with other known antigens, 2) we looked for anti N1 (H5N1) antibodies in humans according to the age and the vaccination status, 3) we checked the neutralizing activity of anti N1 antibodies. The N1 antigen (H5N1) appeared closely related to N1 from swine strains: Sw/31 correlated itself to the pandemic spanish virus (1918-19), and more recent swine isolates from 1982 and 1989. The anti N1 (H5N1) antibodies were present in sera collected from 75+ years old persons and these N1 antibodies were neutralizing H5N1 cells infection. Consequently, 75+ years old persons do not represent a priority group for vaccination in the case of H5N1 pandemic conditions.

Adolescent↗

Sero-response to measles-mumps-rubella vaccine campaign in Saudi Arabia.

OBJECTIVE: To evaluate the measles-mumps-rubella vaccine campaign targeting primary school children in the Kingdom of Saudi Arabia during the months of January and February 2000. METHODS: Pre and post measles-mumps-rubella vaccine blood samples were collected from 54 children from the first grade and 96 children from the 6th grade. Antibodies against measles, mumps and rubella were assayed using enzyme immunoassay and a backup test of plaque neutralization test. This assay was carried out at the Reference Center, Koch Institute, Berlin, Germany. RESULTS: When the backup test results were added, 96%-98% of children had protection level against measles before the campaign. It is interesting to notice that pre-vaccination measles mean titer and positivity rate with enzyme linked immunoassay in the first grade were significantly lower than the 6th grade. One hundred percent and 96.8% were positive for rubella antibody before the campaign, in the first and 6th grades. Seventy-seven percent and 67% were positive for mumps before the campaign, in the first and 6th grade. After the measles-mumps-rubella vaccine campaign, protection rate reached 100% for the 3 antigens with a statistically significant boosting effect and increase in geometric mean of the titre. CONCLUSION: Measles-mumps-rubella vaccine campaign was effective in increasing protective levels and boosting antibodies against the 3 targeted diseases which can prevent epidemics in the primary schools.

Antibodies, Viral↗

Serologic evidence for West Nile virus transmission in Puerto Rico and Cuba.

During the spring of 2004, approximately 1,950 blood specimens were collected from resident and Nearctic-Neotropical migratory birds on the Caribbean islands of Puerto Rico and Cuba prior to northerly spring migrations. Eleven birds and seven birds, collected in Puerto Rico and Cuba, respectively, showed evidence of antibody in a flavivirus enzyme-linked immunosorbent assay. Confirmatory plaque-reduction neutralization test results indicated neutralizing antibodies to West Nile virus in non-migratory resident birds from Puerto Rico and Cuba, which indicated local transmission.

Animal Migration↗

[Experimental imutiol stimulation of the primary immunological response to tetanus anatoxin].

The data on the influence of imuthiol on humoral and cell-mediated immunity developing after immunization against tetanus are presented. Experiments were made on 166 guinea pigs with the use of imuthiol in doses of 25 and 125 mg/kg and tetanus toxoid in doses of 1.5 and 6.0 BU/kg. The dynamic observation of the animals lasted 30 days. The data of the passive hemagglutination test and the neutralization test indicated that imuthiol had considerable influence on the formation of humoral immunity. The results of skin reactions with tetanin, in vitro cell reactions, as well as resistance tests, showed that imuthiol exerted influence on delayed hypersensitivity, the mechanisms of cell-mediated resistance and survival after challenge. The data on the influence of imuthiol could be registered even at early stages after immunization, the immunostimulating action of the preparation reaching its maximum with tetanus toxoid used in a dose of 6.0 BU/kg and imuthiol, in a dose of 125 mg/kg.

Animals↗

[A comparative study on the resistance of bacteriophage T4, phiXM174D, and f2 to glutaraldehyde in laboratory].

OBJECTIVE: To identify the best indicative bacteriophage in disinfection tests through comparing the resistance of bacteriophage T4, phiX174D, and f2 to glutaraldehyde. METHODS: The virucidal activities of glutaraldehyde against bacteriophage T4, phiX174D, and f2 were assessed with suspension tests along with neutralizer tests. The double-agar-layer plaque technique was used to detect the bacteriophage T4, phiX174D, and f2. RESULTS: (1)In a condition of 3000 mg/L of glutaraldehyde and 20 minutes of contact or 6000 mg/L of glutaraldehyde and 5 minutes of contact, "disinfection" level for bacteriophage T4 was achieved, with the log10 inactivation value (LIV) or log10o reduction value (LRV) (=log10No-log10Nt) > or = 4. 00 log10. (2) In a condition of 2500 mg/L of glutaraldehyde and 20 minutes of contact or 5000 mg/L of glutaraldehyde and 5 minutes of contact, the LIV for bacteriophage phiX174D reached "disinfection" level; (3) In a condition of 4000 mg/L of glutaraldehyde and 40 minutes of contact or 8000 mg/L of glutaraldehyde and 10 minutes of contact, the LIV for bacteriophage f2 reached "disinfection" level. CONCLUSION: Bacteriophage f2 and bacteriophage phiX174D has the strongest and weakest resistance to glutaraldehyde respectively.

Bacteriophage T4↗

[Demonstration of the tick-borne encephalitis virus by an immunoenzyme method during a study of its persistence and the chronicity of the process in experimental animals].

The studies carried out by the authors indicated the advantages of the enzyme-immunoassay (EIA) for the detection of the persisting tick-borne encephalitis (TBE) virus and in verification of the chronic course of TBE as compared with HA test, HI test and biologic neutralization test. This method detected TBE virus in the brain of most animals developing the disease within 30-300 days postinoculation. In mice developing no disease the persisting virus was detected both directly in the organs and after preliminary amplification in a sensitive biological model and by the tissue explant method. The EIA was particularly effective in the study of persistence at later intervals (100-300 days), when the hemagglutinating and infective activities of the virus declined. The method is more informative, specific and less labor-consuming.

Animals↗

[Reactogenicity and antigenic activity of a chromatographic cultured purified and concentrated inactivated dried vaccine against tick-borne encephalitis].

The study of the characteristics of a new dried tissue-culture purified concentrated inactivated vaccine against tick-borne encephalitis, manufactured in the USSR, has revealed that the preparation is moderately reactogenic and produces no definite side effects in the vaccinees. In the process of the controlled epidemiological trial the optimum vaccination schedule for the primary immunization of adults against tick-borne encephalitis with the new preparation has been determined by the study of serum samples from the vaccinees in the hemagglutination inhibition test and the neutralization test in tissue culture. In accordance with this vaccination schedule the course of primary immunization with the chromatographic variant of the concentrated vaccine consists of two injections in a dose of 0.5 ml, made at an interval of 6 months.

Adolescent↗

[Prevention of tetanus in the hospital first-aid service].

It has been well known that every wound can become infected by C. tetani. Up to now, the antitetanic prophylaxis and/or therapy were based on empirical criteria, not capable of guaranteeing protection for the wounded. In fact: a) the type of wound and the environment in which the trauma occurred cannot exclude the possibility of infection owing to the ubiquity of the tetanic spore; b) the anamnestic data are often not reliable; c) repeated doses of tetanus anatoxin administered according to different vaccination schedules could give unexpected results; d) the protective effects of a vaccination declines in time in a manner different from one individual to another; e) there are subjects who do not acquire protection even after a vaccination treatment given according to schedule (low responders); f) the excessive number of recall doses of anatoxin, in some subjects can exhibit hypersensitivity side effects. Therefore, for each wounded patient a rational intervention of antitetanus prophylaxis is necessary, based on the rapid determination of antitetanus antibody level. A correlation has been documented between titre of antibody determined by the passive haemoagglutination test (HA) and neutralization test. The HA assay can be a useful tool (because of its easy application and rapidity of the results) in discriminating in wounded patients what type of rational therapeutic approach must be performed.

Antibodies↗

[Serodiagnosis and epidemiology of a California encephalitis group of infections in the Ryazan' region].

Blood sera of 221 healthy residents of 4 districts of Ryazan region were tested in the neutralization test (NT) with Tahyna and Inkoo viruses. The sum of positive responses in the region was 40.7% on an average, this indicating numerous contacts of the local population with these agents. Blood sera of 520 patients with acute seasonal (May-September) fevers and neuroinfections were tested for antibodies to Tahyna and Inkoo viruses in serologic tests (NT and MAC-ELISA-IgM-antibody capture method). Etiologic relationship of the diseases with Tahyna and Inkoo viruses was detected in 51 (9.8%) cases. The patients developed two types of specific immune response, primary or secondary. The best diagnostic result was attained in testing paired sera of patients in NT and MAC-ELISA. MAC-ELISA helped etiologically diagnose the disease after a single testing of a blood serum collected in the acute period of the disease. Inkoo infection was more incident in subjects aged 21 to 40 in May and July-August. The majority of cases of Tahyna infection were diagnosed in subjects aged under 20 in June and July. The incubation period was from 3 to 7 days, the morbidity was sporadic.

Adolescent↗

Lupus anticoagulant in Thai systemic lupus erythematosus patients.

In the past decade, interest in the potential clinical significance of lupus anticoagulant (LA) has grown tremendously. Recent reviews from the Western countries have found an average frequency of 34% for LA in patients with systemic lupus erythematosus (SLE). By using various laboratory procedures, namely, standard and diluted activated partial thromboplastin time, kaolin clotting time, tissue thromboplastin inhibition test and platelet neutralization test, we found the frequency of LA in 91 consecutive Thai SLE patients to be 17.5%, compared with 0.8% in the age-matched normal control population. The presence of LA was significantly associated with disease activity (p = 0.01). A statistically significant association was also observed between the presence of LA and convulsive disorders (p = 0.04), thrombocytopenia (p = 0.001) and autoimmune hemolytic anemia (p = 0.02).

Adolescent↗

[Feasibility of constructing species-specific antigenic erythrocyte diagnosticum for the detection of antibodies to Rickettsia prowazekii].

Species-specific antigenic erythrocyte diagnosticum (SAED) for the detection of antibodies to R.prowazekii was developed. The cells used as the basis for the preparation of the antigenic erythrocyte diagnosticum were formulated and tanned sheep red blood cells (FT-SRBC). Original indication antigens, solubilized with Triton X-100 from common R.prowazekii antigen for the complement fixation test, the antigen neutralization test and the fluorescent analysis, were used as sensitin. The optimum parameters for sensitizing FT-SRBC were established. The SAED thus obtained was stabilized by the method of lyophilization. In a separate series of experiments the specificity and activity of SAED were studied in comparison with those of preparations obtained with the use of different sensitins. The newly developed SAED was shown to be highly specific.

Antibodies, Bacterial↗

A retrospective serological survey of humans in India for Wanowrie virus.

Wanowrie virus (WAN) is an ungrouped arbovirus having morphogenetic characters in common with Bunyavirus. It was isolated for the first time from ticks, Hyalomma marginatum isacci (6700 strain) and subsequently from a human case (68923 strain). In the absence of any reports on the prevalence of this virus amongst humans in India, 600 human sera were tested in the Complement Fixation (CF) test against both the strains of WAN virus. One hundred human sera were randomly selected from six different states, viz Jammu and Kashmir, Rajasthan, Orissa, Maharashtra, Karnataka and Tamil Nadu. CF antibodies were detected in 77 sera (12.8%) against the 6700 antigen while 91 sera (15.1%) possessed antibodies against the 68923 antigen; 60 sera were found to be reactive against both the antigens, with almost identical titres. There was no statistical difference in the distribution of CF reactive sera with respect to age, gender and geographic locations. All the CF reactive sera were found to be negative when tested in the neutralization test against the respective virus strains. The CF antibodies detected in these sera may have been produced during exposure to an antigenically closely related virus yet to be identified.

Adolescent↗

Antigenic variation among bovine viral diarrhea virus (BVDV) strains and the role of different cell fixation methods in immunoassays.

Antigenic variation among 13 Quebec isolates of bovine viral diarrhea virus (BVDV), 4 reference strains and 2 American isolates were studied by peroxidase-linked antibody assay (PLA assay) and neutralization test (NT). The Quebec strains consisted of 3 isolates before 1993 and 10 isolates from 1993. In the PLA assay, we compared 2 different fixatives, acetone and formalin. Acetone-fixation allowed us to identify 6 groups from amongst the viruses tested. All the Quebec isolates were different from the reference strains. In addition, antigenic variation was detected between Quebec isolates obtained before and during 1993. However, PLA assays performed after formalin fixation did not detect these antigenic variations. Neutralization tests were carried out with 2 polyclonal antibodies (PAb) and 6 monoclonal antibodies (MAb). They were used to classify BVDV strains and isolates into 4 groups and 7 subgroups respectively. In conclusion, we demonstrated that the BVDV isolates from the 1993 outbreak in Quebec are antigenically different from reference strains and from isolates existing in Quebec before 1993. In addition, we have shown that 2 internationally used fixation-methods in PLA assay give different results. The usefulness of each method is discussed.

Animals↗