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1.
An indirect microELISA test was performed for detection of maedi-visna antibodies in ovine and caprine species. The antigen consisted in viral particles, highly purified by successive ultracentrifugations.By comparative testing of 934 sera in ELISA and gel immunodiffusion, we found a good correlation between these two teṡts, and moreover, ELISA revealed another 11.3% of positive samples. The precocity of this ELISA was shown by experimental infection of sheep with different strains of maedi-visna: positive sera were detected 7 weeks post-infection, instead 4–5 months with gel immunodiffusion. The complement fixation test was compared with gel immunodiffusion and was found the less sensitive.This ELISA test appeared to be satisfactory, and may be used for early diagnosis of maedi-visna infection.  相似文献   

2.
The aim of this work was to investigate whether an enzyme-linked immunosorbent assay (ELISA) was useful for early detection of maedi-visna virus (MVV) infection in sheep under field conditions. An ELISA based on p25 recombinant protein and a gp46 synthetic peptide was used. Sequentially obtained serum samples (n = 1,941) were studied for 4 years. ELISA results were compared with those of the agar gel immunodiffusion (AGID) test, and results of both tests were compared with a reference result established using consensus scores for at least 2 of 3 serologic techniques (AGID, ELISA, and western blotting, which was used to resolve result discrepancies between the other 2 techniques). A total of 247 discrepancies were observed between ELISA and AGID. Of these, 131 were due to an earlier detection of 120 sera by the ELISA and 11 sera by AGID. The remaining discrepancies (116) were due to the presence of false reactions in both tests. Fewer false-negative results were found by ELISA than with AGID (6 vs. 69 sera, respectively), whereas the number of false-positive results was virtually the same for ELISA and AGID (21 vs. 20, respectively). In relation to the reference result, ELISA sensitivity and specificity were 97.8% and 98.2%, respectively, whereas values for AGID were 76.3% and 98.3%, respectively. The agreement between ELISA and the reference result was higher than that between AGID and the reference result (K value: 0.96 and 0.77, respectively). A variation in the ELISA signal (based on optical density) was observed during the study period, suggesting different antibody levels throughout the animal's life. The ELISA was useful for detecting MVV-infected sheep in field conditions and has potential for use in control and eradication programs.  相似文献   

3.
A study was conducted to compare the indirect enzyme-linked immunosorbent-assay (i-ELISA) test using antigen prepared by a simple technique using sodium dodecyl sulfate (SDS) treatment to the conventional agar gel immunodiffusion test (AGID). Ten specific-pathogen-free (SPF) sheep were inoculated with maedi-visna virus (MVV) and serum antibody titers compared over a period of 14 weeks. All the sheep seroconverted by the i-ELISA compared to 90% by the AGID. The i-ELISA detected antibody at a mean of 2.6 weeks prior to the AGID. In both tests, fluctuations were observed in the serum antibody response of two sheep. The i-ELISA had a specificity of at least 98.8% and an increased relative sensitivity of 15.5% compared to the AGID, based on the analysis of sera from experimental sheep with MVV free status and sera from sheep from various sources. Of the sera from a seronegative flock which had been monitored with the AGID after a "test and remove" eradication program, 10.2% were positive by the i-ELISA. It was concluded that the AGID test may not be adequate to monitor samples for an eradication scheme.  相似文献   

4.
An indirect enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies to maedi-visna virus (mvv) in sheep is described, in which microtitre plates are with a partly purified preparation of mvv. The antibodies bound are detected by a horseradish peroxidase conjugate.The results obtained with ELISA on a total of 493 serum samples from several commercial flocks were compared to those of a routine agar gel precipitation test (AGPT) and a complement fixation test (CFT).All samples which scored positive in AGPT, CFT or both (20.8%) were also found positive by ELISA. In addition, with ELISA a further 11.5% of the samples were positive. Serum samples from maedi-free flocks, from sheep suffering from sheep pulmonary adenomatosis and from lambs immunized against other viruses were all negative by ELISA. The assay has been used routinely for some years and proved to be specific, sensitive and suited for screening of large numbers of serum samples.  相似文献   

5.
An enzyme linked immunosorbent assay (ELISA) and the agar gel immunodiffusion test with bovine leukosis virus glycoprotein as antigen (AGIDT-BLV gp) were further used to test 633 bovine sera for antibodies to BLV. Both tests detected the same number of sera positive (149) or negative (464) for antibodies. Nine sera were negative in the ELISA but found to be weakly positive (2 sera) or bending the control line (7) in the AGDT-BLV gp. On the other hand 11 sera were scored negative in the AGIDT-BLV gp but were weakly positive (9 sera), positive (1), and strongly positive (1) in the ELISA. Both tests are used routinely in this Institute as they complement each other, specially if sera with low antibody titers are under investigation. It is concluded that ELISA can fully replace radioimmunoassays in the serodiagnosis of enzootic bovine leukosis.  相似文献   

6.
An enzyme-linked immunosorbent assay (ELISA) test was developed for the detection of specific antibodies against the unique infectious anemia (EIA) virus in equine sera. The ELISA test was faster and more sensitive when compared with the classic test of agar gel immunodiffusion (AGID). A total of 200 sera were tested: 100 from negative horses and 100 from positive horses by AGID. The ELISA test showed 92 horse sera negative and 100 horse sera positive by AGID with values of optical density (OD) less than 0.139 and higher than 0.139, respectively. Eight horse sera were negative by AGID and higher than 0.139 by ELISA. Six of these became AGID positive also when re-tested 30 days later, and two were of the horses that showed clinical signs of EIA and died before re-testing.  相似文献   

7.
Methods used to prepare antigens from caprine syncytial retrovirus (CSR) for use in the agarose gel immunodiffusion test (AGID) or an indirect enzyme-linked immunosorbent assay (ELISA) are described. Caprine and ovine sera were tested for antibody to CSR using the AGID test and ELISA incorporating a caprine system (CSR antigen and rabbit anti-goat IgG) or an ovine system (maedi-visna virus antigen and rabbit anti-sheep IgG). Good correlation was achieved in the results of the 3 tests when sera were devoid of antibody or were strongly positive. Variations in the results on weakly positive sera were considered to be more a matter of interpretation than due to basic differences in the reagents employed.  相似文献   

8.
Highly purified recombinant gag and env proteins derived from Icelandic strain 1514 of maedi-visna virus were used in an indirect enzyme immunoassay (ELISA) to detect antibodies to small ruminant lentiviruses in sheep and goat sera. The recombinant protein-based ELISA performed very well relative to whole maedi-visna virus and whole caprine arthritis-encephalitis-virus-based ELISAs in its ability to detect anti-maedi visna virus and anti-caprine arthritis-encephalitis virus antibodies, despite the antigenic and genomic variability that is known to exist within and between these two small ruminant lentiviruses. The data suggest that these recombinant maedi-visna virus proteins can be reliably used in an ELISA for the routine serodiagnosis of lentiviral infections in sheep and goats.  相似文献   

9.
An enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies in bovine sera against Parafilaria bovicola nematodes was developed and its sensitivity was compared with the immunodiffusion (ID) method. An exoantigen of P. bovicola which was shown to contain four major polypeptides was used in these procedures. The serological reactivity of the antigen polypeptides was defined by using the enzyme-linked immunoelectrotransfer blot technique (EITB) and whole-worm extract proteins. It identified only four serologically reactive polypeptides with sera from one experimentally infected calf and a verified field case. These two positive sera reacted mainly with four major antigens which coincided in molecular weights of the polypeptides of the exoantigenic preparation, namely, 43, 39, 28 and 25 KDa. Calves experimentally infected with P. bovicola showed a positive reaction with ELISA at 4 months after inoculation, and after this period a rapid increase in serum antibody response occurred. In these cases the ID reaction was observed for the first time at 7 months after inoculation. The specificity of an ELISA method using crude exoantigen preparation of P. bovicola was tested for the diagnosis of bovine parafilariasis. No cross-reactivity was detected when the P. bovicola exoantigen preparation was tested against sera from calves experimentally infected with Onchocerca lienalis, as well as against the sera from cattle naturally infected with Dictyocaulus viviparus or from cattle chronically infected with Ostertagia ostertagi. In addition, testing of 740 field sera from cattle in areas non-endemic and endemic for P. bovicola indicated a specificity of the antigen preparation used. Forty sera from laboratory-confirmed field cases of P. bovicola infection were tested by ELISA and immunodiffusion. All of these sera were ELISA positive, whereas only 70% of these were positive in the ID test. Seven (2.1%) of 328 sera from 21 herds from non-endemic P. bovicola areas were ELISA positive, as opposed to none in the ID test. Of the 94 sera from six herds in areas endemic for P. bovicola infection, 51 (54%) were ELISA positive whereas only 24 (26%) were positive in the ID test. When 56 slaughtered cattle, with varying degrees of meat condemnations due to parafilariasis, were tested for P. bovicola specific antibody, 91% of the serum samples were positive by ELISA. These results suggest that the exoantigen of P. bovicola can be used in a sensitive and reliable serological detection of parafilariasis by ELISA.  相似文献   

10.
A blocking enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies against bovine leukemia virus (BLV) is described. The test is based on the biotin-streptavidin system using unlabelled polyclonal bovine IgG against BLV as catching antibody and biotinylated bovine anti-BLV IgG as detecting antibody. The sensitivity was found to be 50-100 times higher than the agar gel immunodiffusion test, with a specificity of practically 100%. The blocking ELISA proved to be suitable for detection of antibodies against BLV in serum and milk. In 34 paired milk/serum samples, the average ratio of BLV antibody titres was 1:26. So far, more than 700,000 sera have been screened by blocking ELISA for BLV antibodies in the course of the Danish surveillance programme for BLV infection.  相似文献   

11.
When six cattle persistently infected with bovine virus diarrhoea virus (BVDV) were inoculated with lymphocytes infected with bovine leukosis virus (BLV), a depressed antibody response to BLV was observed by ELISA which was due to a decrease in IgG1 synthesis. The ELISA was more sensitive and more reliable than the agar gel immunodiffusion (AGID) test in detecting BLV infection in cattle persistently infected with BVDV. Decreased antibody responses were manifested in the AGID test by negative, inconclusive or weakly positive reactions: only two of the six cattle developed antibodies that generated positive AGID reactions.  相似文献   

12.
A maedi-visna virus CA-TM fusion protein ELISA (MVV ELISA) was evaluated for the detection of antibody in sheep infected with North American ovine lentivirus (OvLV). The results of the MVV ELISA were compared with other assays for OvLV antibody and with viral infection in an intensively studied group of 38 sheep with a high prevalence of OvLV infection and disease. The sensitivity, specificity, and concordance of assays for OvLV antibody (MVV ELISA, indirect ELISA, Western blot, and AGID), virus (virus isolation, PCR, antigen ELISA), and OvLV-induced disease in each animal were compared with OvLV infection status as defined by a positive result in two or more of the assays. Five sheep met the criteria for absence of OvLV infection. The sensitivity of the MVV ELISA in detecting OvLV infected sheep was 64%, whereas the sensitivity of the other three tests for antibody ranged from 85 to 94%. All the antibody assays were 100% specific in this group of animals. Of the assays for virus, the PCR test had the highest sensitivity and the best concordance with OvLV infection, but it also had the lowest specificity of any of the virus or antibody assays. Among the antibody tests, the concordance of the MVV ELISA compared most favorably with the AGID test for detecting OvLV-infected sheep. Analysis of serum samples from 28 lambs experimentally-infected with one of three North American strains of OvLV suggested that there were no significant strain differences detectable by antibody assay. Twenty virus-inoculated lambs were positive by both the MVV ELISA and the AGID test, five lambs were MVV ELISA negative and AGID test positive, and three lambs were MVV ELISA positive and AGID test negative. No pre-inoculation samples were positive by either assay. In a longitudinal study involving seven lambs, antibodies to OvLV were detected by AGID 3-5 weeks post-inoculation, but were not detected by MVV ELISA until 5-10 weeks post-inoculation. Among 128 naturally and experimentally-infected sheep that were seropositive in the AGID test, the overall sensitivity of the MVV ELISA was higher in the naturally infected sheep (84%) than in the experimentally infected sheep (69%). The data indicated that the MVV ELISA represents a less sensitive, but specific alternative for the detection of OvLV antibodies.  相似文献   

13.
I evaluated an enzyme-linked immunosorbent assay (ELISA) and agar gel immunodiffusion (AGID) test simultaneously for the detection of bovine leukemia virus (BLV) antibodies. Total 1,293 serum samples were tested for ELISA and AGID test and the results were compared. The results of ELISA and AGID agreed by 1,156 out of 1,293 (89.4%). All of AGID-positive 356 sera were positive by ELISA. However, of 451 ELISA-positive sera, 95 sera were either negative or equivocal by AGID test. Eleven animals which showed ELISA-positive but AGID-negative or equivocal became AGID-positive in a year. It may be inferred that ELISA detects infected cattle earlier and with greater sensitivity than AGID.  相似文献   

14.
An indirect enzyme-linked immunosorbent assay (ELISA) for the diagnosis of bovine leukaemia virus (BLV) infection was developed and compared with the agar gel immunodiffusion test (AGIDT). Western blotting (WB) was used as confirmatory test. ELISA and AGIDT had specificities that were comparable with that of WB, however, ELISA showed a higher sensitivity than AGIDT. The ELISA was useful for screening a large number of samples, whereas WB was important for detecting the antibody response against the individual BLV-proteins. Different types of positive serological reactions were discerned in WB, that correlated with reactions of sera in AGIDT and ELISA. The most important antigen in WB and ELISA was the BLV protein p24, whereas the BLV glycoproteins gp51 and gp30 were of special importance in AGIDT. The relevance of repeatedly testing the antibody response in BLV-infected herds for control and eradication programmes using assays with higher sensitivity than AGIDT was demonstrated.  相似文献   

15.
A liquid-phase enzyme-linked immunosorbent assay (ELISA) was developed for the detection of vesicular stomatitis virus (VSV) types New Jersey (VSV-NJ) and Indiana subtype Indiana I (VSV-IND1) antibodies in the sera of naturally and experimentally infected cattle, horses and swine. Four different VSV preparations were compared for use as antigen in the ELISA: virus used in neutralization tests, complement-fixation antigen, immunodiffusion ager gel antigen and viral glycoprotein. Comparative antibody titration results of virus neutralization (VN) and ELISA showed no statistically significant difference between serum titers obtained with the four antigens to VSV-NJ (P=0.21) and VSV-IND1 (P=0.14). The viral glycoprotein antigen was incorporated in the ELISA system because it is non-infectious and induces neutralizing antibodies. The reliability and variability of the ELISA was determined by testing 516 bovine, equine and swine sera which originated from areas free of vesicular stomatitis, and by testing 186 sera from areas where outbreaks occur. ELISA and VN results were correlated (P < 0.001 for both serotypes), and no statiscafically difference was found between replications of the tests. The ELISA allows the testing of a larger number of sera in a shorter time than is possible with the VN test and it can be used in diagnostic laboratories in VSV-free areas for the support of epidemiological surveillance programs.  相似文献   

16.
Two control programs were evaluated for their efficiency in eradicating the maedi-visna (M-V) virus from a single sheep flock. In both programs, the agar gel immunodiffusion test was used for the detection of M-V infected animals at regular intervals. In program 1, the test and remove program, ewes that were serologically positive for M-V were immediately removed along with their offspring. The prevalence of infected sheep decreased gradually and a seronegative flock was obtained after 30 months of monitoring. Program 2 entailed the removal of replacement ewe lambs at birth prior to the ingestion of colostrum. Maedi-visna antibodies have not been detected in this flock. These results show that under conditions similar to the industry norms, M-V can be expelled. Although the approach of program 1 is more practical for sheep producers, program 2 is more effective because of the earlier development of a M-V seronegative flock. Because of the nature of the humoral response, a longer time period than four years is required to ensure that M-V has been completely eradicated from each flock.  相似文献   

17.
In order to study pathogenicity of sheep lentiviruses, to obtain monospecific sera and to perfect ELISA, 3 experiments with different strains were carried out for 4 yr. In expt 1, one clone only of a French maedi-visna strain (564-79) elicits a clear seroconversion in inoculated sheep. In expt 2, K1514 is more immunogenic than K796 and PPV: intratracheal route seems more efficient than intracerebral route. Sheep infected by ts mutants (expt 3) are early positive as wild strain K796. Nevertheless, the level of positivity is less important than for the parental strain, suggesting that the defect of the ts mutants is not limiting their replication in vivo. An important result is the lack of clinical signs and anatomical and histopathological lesions, in spite of frequent isolations of virus from buffy coat cells. These results suggest that: different enhancing factors have to be taken in account in the apparition of clinical signs; all the clones are not infectious; viral infection might be effective with several types of virions.  相似文献   

18.
Sera from two sheep experimentally infected with ovine lentivirus (OLV) and from 186 sheep selected from flocks with known high or low prevalence of infection or on the basis of virological or histopathological examination were simultaneously tested by whole virus (WV) ELISA, recombinant transmembrane (r-TM) ELISA and AGID assay. Antigens for both the WV ELISA and AGID were prepared from an Italian field isolate; recombinant antigen was derived from the N-terminal region of the transmembrane envelope protein of strain K1514. The WV ELISA detected the highest number of seropositives, followed by the r-TM ELISA and AGID test. The sensitivity and specificity of the r-TM ELISA relative to the WV ELISA were 0.66 and 0.95, respectively. Immunoblot analysis of 14 WV ELISA-positive and r-TM ELISA-negative sera showed that the major core protein was immunodominant on WV antigen. It is concluded that the r-TM ELISA was more sensitive than the AGID test but less sensitive that the WV ELISA, particularly for detecting antibodies in the early stages of infection.Abbreviations AGID agar gel immunodiffusion - bp base pair - ELISA enzyme-linked immunosorbent assay - FBS fetal bovine serum - IB immunoblot - kD kilodalton - OLV ovine lentivirus - MEM minimal essential medium - p.i. post-infection - r recombinant - SDS-PAGE sodium dodecyl sulphate-polyacrylamide gel electrophoresis - TM transmembrane - WV whole virus  相似文献   

19.
An indirect ELISA test was developed for the diagnosis of Brucella canis infection in dogs. A bacterial whole cell extract was used as a solid phase antigen, using B. canis isolated from an infected animal. Sera from culture-positive and healthy negative animals were used as internal reference controls. The cut-off point was determined by a mathematical formula for a statistically valid value, which defined the upper prediction limit, based on the upper tail of the t-distribution of 21 negative control sera readings, for the confidence level of 99.5%. The sensitivity and specificity of the ELISA test were 95% and 91%, respectively. The ELISA test showed a significant concordance index (K=0.84) with the agar gel immunodiffusion test. The reliability of the ELISA for the detection of infected animals was established by a double blind study testing 280 sera provided by serum banks from different diagnostic and research institutions and analyzed by ROC Curve.  相似文献   

20.
An enzyme-linked immunosorbent assay (ELISA) for detecting antibodies to enzootic bovine leukosis (EBL) virus is described and its sensitivity compared with that of the agar gel immunodiffusion test (AGIDT) using 198 sera collected in Great Britain. There was 95 per cent agreement between the ELISA and AGIDT, when sera with positive/negative ratio (P/N) values of 1 . 5 or greater were considered positive. A total of 259 out of 264 sera (98 per cent) collected in Northern Ireland had P/N values of less than 1 . 5, the remaining sera having P/N values of 1 . 5 and 1 . 6. As Northern Ireland is clinically and serologically free of EBL infection it is proposed that sera with P/N values of 1 . 5 and 1 . 6, which account for approximately 3.5 per cent of the total sera tested, are considered doubtful and should be tested by another serological test.  相似文献   

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