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1.
OBJECTIVE: To determine the value of virus isolation (VI), immunofluorescent antibody (IFA) assay, serum neutralization (SN), and ELISA for the diagnosis of clinical feline herpesvirus-1 (FHV-1) infection in cats. ANIMALS: 46 clinically normal cats, 17 cats with signs of acute respiratory tract disease, and 38 cats with signs of chronic ocular disease. PROCEDURE: Conjunctival swabs for VI, conjunctival scrapings for IFA testing, and venous blood samples for SN or ELISA testing were obtained from all cats. RESULTS: FHV-1 was detected in 10.9 and 28.3% of clinically normal cats and in 18.2 and 33.3% of cats with FHV-1-associated disease by VI and the IFA assay, respectively. There were no significant differences in the viral detection rate between cats with acute respiratory tract disease and cats with chronic ocular disease or between diseased cats and clinically normal cats; however, FHV-1 was never detected by both methods in clinically normal cats. Overall FHV-1 seroprevalence was 97% when tested by ELISA and 66% when tested by SN. Seroprevalence did not vary significantly among the 3 groups for either serologic test. Magnitude of SN and ELISA titers varied greatly but independently of presence or absence of clinical signs of FHV-1-associated disease. Sensitivity, specificity, and positive and negative predictive values were assessed for VI and the IFA assay--jointly and individually--and for each SN and ELISA titer magnitude. Values never all exceeded 50%. CLINICAL IMPLICATIONS: Because FHV-1 can be detected commonly in clinically normal cats by the IFA assay or VI, neither test appears to aid in the clinical diagnosis of FHV-1 infection. Seroprevalence does not appear to vary between affected and clinically normal cats. SN, ELISA, VI, and the IFA assay appear to be of limited value in the diagnosis of FHV-1-associated disease in cats. Concurrent assessment of the IFA assay and VI results may permit exclusion of FHV-1 as an etiologic agent if results of both tests are negative.  相似文献   

2.
A specific and sensitive enzyme-linked immunosorbent assay (ELISA) was established for the detection of antibodies to bovine viral diarrhea virus (BVDV) in bovine sera. Polyethylene-glycol concentrated, equilibrium density gradient purified BVDV was used as test antigen at an optimal amount of 1 microgram/well, whereas the optimal concentration of conjugate was at 1/2000 dilution. The standardized test encountered no non-specific reaction with test sera at a starting dilution of 1/10. A total of 50 bovine serum samples was assayed for the presence of antibodies against BVDV by ELISA and serum neutralization test (SNT). A positive correlation between the 2 tests was found. However, ELISA could be as much as 500-fold more sensitive than SNT in detecting low levels of BVDV antibodies.  相似文献   

3.
OBJECTIVE: To determine whether detection of virus-specific serum antibodies correlates with resistance to challenge with virulent feline herpesvirus 1 (FHV-1), feline calicivirus (FCV), and feline parvovirus (FPV) in cats and to determine percentages of client-owned cats with serum antibodies to FHV-1, FCV, and FPV. DESIGN: Prospective experimental study. ANIMALS: 72 laboratory-reared cats and 276 client-owned cats. PROCEDURES: Laboratory-reared cats were vaccinated against FHV-1, FCV, and FPV, using 1 of 3 commercial vaccines, or maintained as unvaccinated controls. Between 9 and 36 months after vaccination, cats were challenged with virulent virus. Recombinant-antigen ELISA for detection of FHV-1-, FCV-, and FPV-specific antibodies were developed, and results were compared with results of hemagglutination inhibition (FPV) and virus neutralization (FHV-1 and FCV) assays and with resistance to viral challenge. RESULTS: For vaccinated laboratory-reared cats, predictive values of positive results were 100% for the FPV and FCV ELISA and 90% for the FHV-1 ELISA. Results of the FHV-1, FCV, and FPV ELISA were positive for 195 (70.7%), 255 (92.4%), and 189 (68.5%), respectively, of the 276 client-owned cats. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that for cats that have been vaccinated, detection of FHV-1-, FCV-, and FPV-specific antibodies is predictive of whether cats are susceptible to disease, regardless of vaccine type or vaccination interval. Because most client-owned cats had detectable serum antibodies suggestive of resistance to infection, use of arbitrary booster vaccination intervals is likely to lead to unnecessary vaccination of some cats.  相似文献   

4.
试验旨在利用胶体金免疫层析技术建立快速检测犬血清中犬细小病毒（canine parvo virus, CPV）血凝抑制（haemagglutination inhibition, HI）抗体效价的方法,用于CPV疫苗免疫效果评价。采用双抗体夹心法,以抗CPV血凝相关抗原的单克隆抗体制备CPV抗原检测试纸条;将犬血清进行不同比例系列稀释后,分别与定量CPV抗原充分反应,滴入CPV胶体金试纸条,根据试纸条检测线（test line,T线）消失时的血清最高稀释倍数判断血清中CPV抗体的HI效价;用此方法检测86份犬血清样品,并与传统血凝抑制试验方法进行分析比较。结果显示,成功制备CPV抗原检测试纸条,确定了试纸条检测犬血清CPV-HI效价的反应条件和结果判定标准。结果表明,在检测不同稀释倍数犬血清反应后的CPV抗原时,能使试纸条T线消失时的血清最高稀释倍数与HI效价具有正相关性,犬血清最高稀释倍数乘以4即为HI效价;两种方法的符合率达90.7%。本试验初步建立了胶体金试纸条检测CPV血凝抑制效价的方法,为检测CPV-HI效价提供了一种操作简单、快速的试验方法,可用于CPV疫苗免疫效果评价。  相似文献   

5.
An enzyme-linked immunosorbent assay (ELISA) was developed to measure specific antibody activity in sera of chickens exposed to Newcastle disease virus (NDV). A near-linear relationship existed between the log of the corrected absorbance of antisera at a single working dilution and the corresponding observed serum titers as determined by a standard serial-dilution method. Regression analysis was used to construct a standard curve and extract an equation from this relationship. The equation was used to convert corrected absorbance readings of the single working dilution directly into predicted ELISA antibody activity titers. In a comparative study, a correlation (P less than 0.01) was found between ELISA and hemagglutination-inhibition (HI) antibody titers to NDV. ELISA titers were as much as 160 times greater than the HI titers. ELISA was also able to detect much lower levels of antibody activity than the HI test.  相似文献   

6.
The study was aimed to use colloidal gold immune chromatography technology to establish a rapid method for detection of canine serum canine parvovirus (CPV) hemagglutination inhibition (HI) titer and CPV vaccine immunization effect assessment.Double antibody sandwich method and monoclonal antibodies of anti-CPV hemagglutination antigen were used to prepare CPV antigen test strip.Canine serum with different proportion respectively was mixed with quantitative CPV antigen for full reaction,then dropped the mixture into the CPV colloidal gold test strip,so according to the highest serum dilution ratios when the test strip line T (line T) vanishes,it was to judge CPV antibodies in serum of the HI titer.This method had been used to detect 86 canine serum samples,at the same time,analyzing and comparing it with traditional hemagglutination inhibition test method.The results showed that the CPV antigen detection test strip was successfully prepared,and the reaction conditions and results of the test strip for detecting the titer of CPV-HI in canine serum were determined.The results indicated that when detecting CPV antigen after the dilution of different ratios of canine serum,the highest serum dilution ratios when the strip line T vanished and the HI titer had positive correlation.The highest dilution ratios of canine serum multiplied by 4 was the HI titer.The results of two methods had 90.7% consistency.This experiment established the colloidal gold immune chromatography test strip for the detection of CPV-HI titers method initially.This CPV-HI detection provided a simple and fast test method for the effect evaluation of CPV vaccine immune.  相似文献   

7.
An indirect enzyme-linked immunosorbent assay (ELISA) was developed which was specific and sensitive in detecting antibodies to Ehrlichia risticii in Potomac horse fever (PHF). The ELISA antibody titers were correlated with the indirect fluorescent antibody (IFA) titers. E. risticii propagated in human histiocyte culture was purified on renografin gradient and the band of the organisms at a density of 1.182 g/ml was used as antigen. ELISA antibody titers were determined through computer assisted analysis, the observed antibody titers were derived by serial serum dilutions and using a resultant standard curve the predicted antibody titers were obtained from a single serum dilution. The standard curve had a correlation coefficient of 0.8975. The observed and predicted antibody titers were in good agreement, as the respective titers fell within a two-fold range. There was a good correlation between ELISA and IFA test results, but the ELISA titers were several times higher. In experimental infections of horses produced with the infected equine whole blood and the Ehrlichia infected macrophage culture, the antibodies were first detected in two weeks and one week postinoculation (PI), respectively. In both cases the titers reached a peak in about 4 weeks PI with a mean titer of 1:16558 and 1:4030, respectively. The antibody titers of the convalescent sera of field cases of PHF were comparatively lower than the experimentally infected horses.  相似文献   

8.
An enzyme linked immunosorbent assay (ELISA) was developed to detect antibody to bovine viral diarrhoea virus (BVDV) in bovine serum. The ELISA results were compared with those of the serum neutralisation test (SNT) using serums from 6 experimentally infected calves bled at intervals from 0 to 154 days postinfection and 886 field samples. The optical density (OD) produced by a single dilution of test serum was compared with a standard curve and the result expressed in ELISA units. Despite wide variation between absolute ELISA and SNT results, an agreement of 97% was obtained when reciprocal SNT titres greater than or equal to 8 and ELISA units greater than or equal to 10 were taken as indicative of a specific reaction. The ELISA was shown to be an efficient method of measuring antibody in bovine serum samples and would assist in any large scale screening of cattle herds for BVDV antibody.  相似文献   

9.
This study was designed to investigate the application of indirect enzyme-linked immunoassay (ELISA) in detecting IgG against Japanese encephalitis virus in swine sera and the qualitative nature of this test. The attenuated strain SA14-14-2 of Japanese encephalitis virus (JEV) was inoculated into 9-day-old chicken embryos and virus was harvested, purified and suspended in 0.9% saline as JEV antigen. The control antigen was prepared by the same method as for the antigen. In the ELISA, the optimal concentrations of antigen coated and dilution factor were selected using chi2 test. Ninety-two swine sera negative to haemagglutination inhibition (HI) were tested by this assay and the positive threshold was determined. The results of this study indicate that indirect ELISA has high specificity, sensitivity and reproducability. Simultaneous testing of 74 serum samples from nine pig farms was carried out to compare the existing HI test and the indirect ELISA. The coincidence rate of the two assays was 85.1% (63/74) and no significant difference was observed between them (p > 0.05). This ELISA test can detect 46 swine serum samples qualitatively and the titre of eight swine serum samples through endpoint dilution quantitatively within one 96-well plate.  相似文献   

10.
Egg yolk was evaluated in the enzyme-linked immunosorbent assay (ELISA) as an alternative source of antibodies for detection of Mycoplasma gallisepticum (MG) and M. synoviae (MS) infections in chickens. There was no statistically significant difference (P greater than 0.05) between the ELISA geometric mean titers (GMTs) of saline-diluted egg yolk and chloroform-extracted egg yolk, and both preparations had a high correlation coefficient (0.87 for MG; 0.97 for MS). The saline-diluted and chloroform-extracted yolk had a relative sensitivity of 90% and specificity of 98% in the MG ELISA; in MS ELISA they were 100% and 96%, respectively. Hemagglutination-inhibition (HI) results with chloroform-extracted samples were satisfactory, but those with saline-diluted samples were not. Neither preparation was satisfactory for use in the rapid plate agglutination (RPA) test. A 1-ml sample of yolk was compared with the whole-yolk method. The chloroform-extracted whole yolk yielded a significantly higher (P less than 0.05) GMT in the MG ELISA; however, there was no statistically significant difference (P greater than 0.05) between GMTs yielded by the two procedures in the MS ELISA. The correlation coefficients for the two sampling methods were 0.73 for MG ELISA and 0.63 for MS ELISA. ELISA detected no statistically significant difference (P greater than 0.05) between GMTs of serum and chloroform-extracted yolk from individual birds. Results with the HI test were comparable to those with ELISA on the same samples. The RPA test yielded comparable results on the serum samples. No statistically significant differences (P greater than 0.05) were observed in HI or ELISA antibody levels between egg-yolk samples and sera on random samples collected from nine flocks that were MG- and MS-free or were infected with MG, MS, or both; however, egg-yolk samples tended to have slightly higher titers than sera in both tests. The optimum screening dilution of chloroform-extracted yolk for detecting MG and MS antibodies by ELISA was 1:800.  相似文献   

11.
H3N8亚型马流感病毒间接ELISA抗体检测方法建立及应用   总被引：5，自引：0，他引：5  
为建立马流感血清学ELISA诊断方法,本研究以马流感病毒中国分离株A/马/新疆/07(H3N8)通过SPF鸡胚培养和增殖,收取含病毒尿囊液经蔗糖密度梯度离心纯化后作为ELISA包被抗原,首次在我国建立了检测H3N8亚型马流感抗体的间接ELISA诊断方法。试验的最佳反应条件为:最佳抗原稀释度7μg/mL,封闭液5%脱脂乳,血清稀释度1∶100,二抗稀释度1∶10000,稀释液PBS(pH7.4),血清反应时间1.5h,二抗反应时间1h。通过本方法对555份临床样品进行检测并与血凝抑制(HI)试验检测结果比较,证明本方法特异、敏感,具有良好的稳定性和可重复性,适于马流感的流行病学调查和监测工作。  相似文献   

12.
OBJECTIVE: To determine whether uveitis in cats was associated with intraocular production of feline herpesvirus type 1 (FHV-1)-specific antibodies or with detection of FHV-1 DNA in aqueous humor (AH). ANIMALS: 44 cats with idiopathic uveitis, 29 cats with uveitis attributed to Toxoplasma gondii infection, 13 FHV-1 seropositive cats without uveitis, and 9 FHV-1 seronegative cats without uveitis. PROCEDURE: ELISA were used to detect FHV-1-specific antibodies and total IgG antibodies in serum and AH, and the Goldmann-Witmer coefficient (C-value) for intraocular antibody production was calculated. A polymerase chain reaction assay was used to detect FHV-1 DNA in AH. RESULTS: FHV-1 seroprevalence among cats with uveitis was not significantly different from seroprevalence among cats without uveitis. Intraocular FHV-1 antibodies were never detected in cats without uveitis. Significantly more cats with idiopathic uveitis (22/44) or with toxoplasmic uveitis (11/29) had evidence of intraocular antibody production (C-value > 1) than did cats without uveitis. Only cats with idiopathic uveitis had FHV-1 C-values > 8. Among cats with evidence of intraocular antibody production, cats with idiopathic uveitis had a significantly higher median FHV-1 C-value (9.61) than did cats with toxoplasmic uveitis (2.56). Overall, FHV-1 DNA was detected in AH from 12 cats, 11 of which had uveitis. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that FHV-1 can infect intraocular tissues of cats and that intraocular FHV-1 infection may be associated with uveal inflammation in some cats.  相似文献   

13.
Three serological assays were compared for detection of antibodies to bovine herpes-virus type 1. These were virus neutralization (VN), enhanced complement fixation (CF) and enzyme-linked immunosorbent assay (ELISA). The ELISA was developed using an infected cell lysate antigen and purified virus and was optimized in relation to antigen and antisera dilutions. The CF assay was enhanced by the addition of bovine complement. These 3 assays were compared for detection of: specific virus antibody titers; sero-conversions; early antibody response in experimentally-infected cattle. Both ELISA end-point titers and single dilution values were found to be more sensitive than the CF or VN assays for specific antibody level quantitation. With a single dilution ELISA test procedure a correlation was obtained between ELISA values and VN titers. Using the single dilution ELISA test the assay also detected antibodies in experimentally-infected cattle before either the VN or CF assays, and agreed with the VN test in 35/38 seroconversions found by 4-fold or more VN changes between acute and convalescent paired sera from naturally-infected animals. The single dilution ELISA was a rapid and sensitive test for routine antibody detection in bovine sera.  相似文献   

14.
Serial dilution and single dilution enzyme linked immunosorbent assays (ELISA) were standardised and their sensitivity and specificity were compared for serodiagnosis of Babesia equi infection. The antibody titres of 24 donkey sera of known identity were determined separately by serial dilution ELISA using three different B. equi antigens namely whole merozoite (WM), cell membrane (CM) and high speed supernatant (HSS). The ratios of the optical density (OD) of known positive and known negative sera at different serum dilutions were calculated and termed as the positive/negative (P/N) ratio. The coefficients of correlation (r) were calculated between the P/N ratios at different dilutions of sera and the log10 antibody titres of the same sera were ascertained by serial dilution ELISA. The highest value of 'r' was obtained at a serum dilution of 1:200. From log10 antibody titre of sera (y) and their P/N ratio at a dilution of 1:200 (x), regression equations (y = a + bx) were calculated separately for the three antigens. Test sera were diluted to 1:200, their OD were read in duplicate wells and were converted to the P/N ratio. Antibody titres were predicted from the P/N ratio using a regression equation separately for the three antigens. Titres obtained by both ELISAs were not significantly different from each other, thus confirming that single dilution ELISA could be successfully used to replace conventional serial dilution ELISA. The sensitivity, specificity and predictive value of single dilution ELISA was validated statistically using 42 B. equi disease-positive sera and 106 B. equi disease-negative sera. The WM antigen was found to be the most sensitive with a higher predictive value for negative test sera as compared to the CM or HSS antigens. Sera positive for other equine infections including Babesia caballi showed no cross-reaction with the three B. equi antigens in ELISA, thus the test was immunologically specific. Antibody titres of 109 unknown field donkey/horse sera obtained by serial and single dilution ELISA using the WM antigen did not show any significant difference. Since the single dilution ELISA was found to be more economical, convenient, sensitive, specific than the serial dilution ELISA and has a high predictive value, it is suitable for use in sero-epidemiological studies on B. equi infections in the field.  相似文献   

15.
Two groups of feline panleukopenia virus (FPV), feline calicivirus (FCV), and feline herpesvirus-1 (FHV-1) seronegative cats (five cats per group) were administered one of two modified live feline viral rhinotracheitis, calicivirus, and panleukopenia virus (FVRCP) vaccines and the serological responses to each agent were followed over 28 days. While all cats developed detectable FPV and FCV antibody titers; only two cats developed detectable FHV-1 antibody titers using the criteria described by the testing laboratory. For FPV and FHV-1, there were no differences in seroconversion rates between the cats that were administered the intranasal (IN) FVRCP vaccine and the cats that were administered the parenteral FVRCP vaccine on any day post-inoculation. For FCV, the cats that were administered the IN FVRCP vaccine were more likely to seroconvert on days 10 and 14 when compared to cats that were administered the parenteral FVRCP vaccine.  相似文献   

16.
A rapid screening assay for determining antibodies to canine parvovirus in dog serum using monoclonal antibodies and enzyme-linked immunosorbent assay (ELISA) technology was developed. The ELISA could be read visually, and the results correlated well with serum neutralization (SN) and hemagglutination inhibition (HI) titers. Sera with SN less than or equal to 1:4 or HI less than or equal to 1:10 had an 87.9% correlation with ELISA and sera with SN greater than or equal to 1:64 or HI greater than or equal to 1:80 had a 94.4% correlation. The assay took only 10 to 15 minutes to perform and did not require specialized equipment. The ELISA should be useful in monitoring dogs for the presence of maternal antibodies against parvovirus and for determining seroconversion after vaccination.  相似文献   

17.
COMBSCORES determined using the ImmunoComb solid-phase immunoassay were compared with hemagglutination-inhibition (HI) titers specific for Newcastle disease virus (NDV) and infectious bronchitis virus (IBV) and with mean enzyme-linked immunosorbent assay (ELISA) titers determined using Agritech Systems, Inc., ELISA. COMBSCORES for NDV and IBV increased proportionately in a stepwise manner as HI titers increased. The ImmunoComb solid-phase immunoassay was ablt to produce endpoint titers on sera with NDV-HI titers of 0 through 320 and IBV-HI titers of 0 through 1024 without reaching the maximum S-value. The ImmunoComb showed good correlation with the HI assay and the Agritech ELISA and should prove to be a useful tool for serological profiling, either alone or in conjunction with the HI test or commercial ELISA.  相似文献   

18.
Chickens were experimentally infected with a duck adenovirus that has been shown to be serologically indistinguishable from Adenovirus 127. Sera and eggs were collected at intervals after exposure for antibody determination by the hemagglutination-inhibition (HI) test, the enzyme-linked immunosorbent assay (ELISA), and the immunodiffusion (ID) test. Egg yolks were processed for use in the serological tests by (a) dilution in phosphate-buffered saline (PBS), (b) extraction of the water-soluble fraction with chloroform, or (c) freezing and thawing PBS-diluted yolks and testing the supernatant fluid. HI antibody titers from serum and extracted yolk were similar except during the initial 2 weeks, when yolk antibody levels were low or absent. Chloroform-extracted yolks were suitable material for the HI, ELISA, and ID tests. Heat inactivation of the chloroform-extracted yolk had no effect on titers.  相似文献   

19.
Samples were collected from 36 cats with feline herpesvirus (FHV-1)-related ocular disease (conjunctivitis, epithelial or stromal keratitis, or corneal sequestration), and 17 cats without ocular changes. Corneoconjunctival swabs, scrapings and biopsies were tested in various combinations for presence of FHV-1 DNA using single round (sr) polymerase chain reaction (PCR) and nested PCR (nPCR). Additional swabs from the inferior conjunctival fornix were tested by enzyme-linked immunosorbent assay for Chlamydophila felis antigen. Cytologic evaluation was carried out on conjunctival (cats with conjunctivitis) and corneal (cats with keratitis) cytobrush preparations. FHV-1 DNA was detected by PCR in 14 (39%) cats with ocular disease and 1 (6%) of the control group. Agreement between srPCR and nPCR results was significant (P < 0.01). FHV-1 DNA was detected in 3/7 cats with conjunctivitis, 5/6 cats with epithelial keratitis, 3/11 cats with stromal keratitis, and 3/12 cats with corneal sequestration. There was a significant association (P = 0.0027) between viral presence and epithelial keratitis. However, no significant association was found between viral presence and conjunctivitis (P = 0.059), stromal keratitis (P = 0.15), or corneal sequestration (P = 0.18). With respect to FHV-1 DNA detection, intersample agreement was significant (P < 0.03). No sampling technique seemed more likely than another to harvest detectable viral DNA, except for cats with corneal sequestrum in which viral DNA was not detected using corneoconjunctival swabs. FHV-1 DNA was detected in 6/9 samples with intranuclear inclusion bodies and in 6/7 cats with eosinophils on cytologic examination. All samples tested negative for C. felis antigen.  相似文献   

20.
During an outbreak of bovine enzootic encephalomyelitis caused by the Akabane virus (AKAV) in 2010, 210 serum samples were collected from the affected cattle, and serological investigations for the AKAV were performed using a serum neutralization test (SNT) and an enzyme-linked immunosorbent assay (ELISA). The seropositive rates for SNT and ELISA were 90.0 and 85.2 %, respectively. The titers of SNT (log2) against the AKAV were higher than 4.0 in the highly affected cattle (80.0 %). This finding indicates that most affected cattle were infected with the AKAV and that strong immune responses against this virus were elicited in affected cattle. The strong immune response to the AKAV in cattle may provide insight into the occurrence of bovine encephalomyelitis caused by the AKAV.  相似文献   

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