Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “ELISA”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 145 records · Page 8Linked to original sources
PubMed1999

Comparison of commercial ELISA test kits from Australia and the USA with the serum neutralization test in cell cultures for the detection of antibodies to the infectious laryngotracheitis virus of chickens.

The objective of this work was the evaluation under our conditions of two commercial ELISA kits for the detection of antibodies to avian infectious laryngotracheitis (ILT) virus, one from Australia (Trop-ELISA, TropBio) and one from the USA (ProFLOK-ELISA, KPL), and to compare their performance with the conventional serum neutralization test (SNT) in chick embryo liver cells. Repeatability varied considerably, particularly when using the Trop-ELISA. Therefore, if individual results are important, at least two parallel measurements per serum sample are recommended. In 89.3% of the sera tested by SNT, results of two parallel measurements did not vary by more than one 2-fold dilution step. There was good linear correlation between both ELISAs and the SNT, the correlation coefficient for the Trop-ELISA being r = 0.758 and for the ProFLOK-ELISA r = 0.867. The negative/positive cut-off was redefined to suit our conditions. Sera with a SN titre of > or = 1:4 were considered positive. Sera with < or = 15% absorption in the ProFLOK-ELISA were considered clearly negative. For the Trop-ELISA, extinctions of > or = 0.477 were considered positive, < or = 0.168 clearly negative. Values in between were regarded as doubtful for young chickens and as possibly due to non-specific reactions in older chickens. The sensitivity and specificity of the ELISAs relative to the SNT were 87 and 77% for the Trop-ELISA, and 95 and 60%, respectively, for the ProFLOK-ELISA. However, the results indicate that the sensitivity of the ELISA is higher than that of the SNT, because most sera showed similar deviations from SNT results with both ELISAs. Generally, both ELISAs were a suitable alternative to the SNT under our conditions, as long as only negative/positive results are required.

Journal Article↗
PubMed1989

A blocking ELISA for the detection of specific antibodies to bovine respiratory syncytial virus.

A blocking enzyme-linked immunosorbent assay (ELISA) has been adapted to detect specific antibodies in bovine sera to respiratory syncytial virus using a horseradish peroxidase-labeled monoclonal antibody to the fusion protein of the virus. This assay plus an indirect blocking ELISA and indirect ELISA were used to detect antibodies to the bovine respiratory syncytial virus (BRSV) in 159 field-origin bovine sera. Results of these assays were compared with serum antibody titers measured by the serum neutralization (SN) test. Over a 56-day period, the mean neutralization titers and the mean delta absorbance values for the blocking ELISA, on the same sera, showed similar declines. However, the calculated correlation coefficients between mean SN titer and mean absorbance value for the blocking ELISA of the individual sera ranged from -0.2 to -0.5 depending on the source of sera. Similar values were obtained whether using crude or purified viral antigen in the assays. Corresponding calculated correlation coefficients were generally higher for the indirect blocking ELISA or indirect ELISA than for the blocking ELISA. The blocking ELISA was between 70 and 64% as sensitive as the serum neutralization test with a specificity of 100 or 90% using the crude and purified viral antigen, respectively. The indirect blocking ELISA and indirect ELISA had similar calculated sensitivities and specificities. The blocking ELISA was faster to run than either of the other ELISA's or the neutralization test. Further, nonspecific background absorbance was obviated because the blocking ELISA detects antibodies to 1 specific viral protein, the fusion protein. These studies suggest that the blocking ELISA should be useful as a serological test for BRSV antibodies.

Animals↗
PubMed2001

High sensitivity of a novel ELISA for anti-M2 in primary biliary cirrhosis.

The use of an ELISA for the detection of anti-M2, a specific autoantibody in primary biliary cirrhosis (PBC), has been common in Japan. However, there are some problems in the sensitivity of this ELISA, especially in PBC patients showing antimitochondrial antibody (AMA)-negative sera or low AMA titers by immunofluorescence. Recently, a new ELISA for anti-M2 was developed, using porcine heart mitochondrial protein as the antigen. We report here comparative studies of the new and the former anti-M2 ELISAs. Porcine heart mitochondrial protein was prepared and used as the antigen for the new ELISA for anti-M2. Sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of this protein showed three major M2 antigen proteins. As the second antibody, peroxidase-conjugated anti-human mouse monoclonal IgM, in addition to monoclonal IgG, was included. The sera of 171 PBC patients were examined. As controls, we examined the sera of 167 non-PBC patients and the sera of 115 normal controls. The cut-off index was set at 10 U/ml, based on the results for the normal controls. No sera from the non-PBC patients or the normal controls were positive for anti-M2 by either the new or the former ELISA. However, the positivity rate for anti-M2 in PBC patients with the new ELISA was 78%; in contrast, that with the former ELISA was only 54%; this difference was significant (P = 0.00001). In particular, in 65 patients showing AMA titers of 1:20 or less, the positivity rate with the new ELISA was 51%; in contrast, that with the former ELISA was only 17%. As the sensitivity of the new ELISA is significantly higher than that of the former ELISA, especially for sera from patients showing AMA-negativity or low titers of AMA, the new ELISA is considered to be more effective than the former ELISA for use in anti-M2 screening assays in patients with PBC.

Animals↗
PubMed2005

Development of monoclonal antibody-linked ELISA for sero-diagnosis of vesicular stomatitis virus (VSV-IN) using baculovirus expressed glycoprotein.

The gene encoding the envelope glycoprotein (GP) of vesicular stomatitis virus serotype, Indiana (VSV-IN), was expressed under the polyhedron promoter of baculovirus. The recombinant GP was applied as a diagnostic antigen for the detection of cattle and horse antibodies to VSV. In addition, the neutralizing monoclonal antibody (Mab) to GP of VSV-IN was used as trapping antibody in a Mab-linked indirect ELISA (MLI-ELISA) or detecting antibody in a Mab-linked competitive ELISA (MLC-ELISA). The diagnostic efficiencies of MLI-ELISA and MLC-ELISA were evaluated with currently available C-ELISA from OIE reference laboratory for vesicular stomatitis as a gold standard by using VSV-positive equine sera and negative bovine sera vaccinated against foot-and-mouth disease (FMD) in the field. When naturally infected equine sera and FMDV vaccinated bovine sera were tested, MLI-ELISA and MLC-ELISA showed relative sensitivities of 80% and 95% with relative specificity of 97% and 99%, respectively. However, both ELISAs cross-reacted with equine sera against New Jersey (VSV-NJ) serotype. The comparison of the two ELISAs revealed that MLC-ELISA was relatively more sensitive and specific than MLI-ELISA, indicating that MLC-ELISA can be applied to sero-diagnosis for VSV-IN infection.

Animals↗
PubMed1999

Comparison of a maedi-visna virus CA-TM fusion protein ELISA with other assays for detecting sheep infected with North American ovine lentivirus strains.

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.

Animals↗
PubMed2000

Comparative evaluation of a field-based dot-ELISA kit with three other serological tests for the detection of Brucella antibodies in goats.

A dot-ELISA (d-ELISA) test was evaluated and compared with the serum agglutination test (SAT), micro-complement fixation test (CFT) and a plate-ELISA (p-ELISA) for field use in screening herds of goats against brucellosis. During the standardization of the dot-ELISA kit on 1732 caprine serum samples, 1571 samples out of 1666 were found to be negative in d-ELISA, SAT and micro-CFT, while 59 were positive in different combinations. Of a further 66 serum samples, 34 were negative and 31 were positive in different combinations in d-ELISA, SAT, micro-CFT and p-ELISA. A total of 1584 goats belonging to different herds were then screened for brucellosis. Of the 694 serum samples screened in the first batch using d-ELISA, a positive reaction was observed in 26 cases. Further screening of these cases revealed 13 and 21 goats as positive reactors in SAT and CFT, respectively. In a second batch of 890 goats there were 109 positive reactors in d-ELISA. Among these 109 goats, 34, 40 and 80 goats were positive reactors in SAT, CFT and p-ELISA, respectively. The results of d-ELISA correlated well with those of p-ELISA. Dot-ELISA was found to be a more suitable and rapid test for screening large numbers of goats in the field.

Agglutination Tests↗
PubMed1999

Indirect ELISAs based on recombinant and affinity-purified glycoprotein E of Aujeszky's disease virus to differentiate between vaccinated and infected animals.

Two indirect ELISAs for the detection of antibodies against glycoprotein E (gE) of Aujeszky's disease virus (ADV) in sera have been developed. The rec-gE-ELISA is based on the E. coli-expressed recombinant protein containing the N-terminal sequences of gE (aa 1-125) fused with the glutathione S-transferase from Schistosoma japonicum. The affi-gE-ELISA is based on native gE, which was purified from virions by affinity chromatography. The tests were optimised and compared with each other, as well as with the recently developed blocking gE-ELISA (Morenkov et al., 1997b), with respect to specificity and sensitivity. The rec-gE-ELISA was less sensitive in detecting ADV-infected animals than the affi-gE-ELISA (sensitivity 80% and 97%, respectively), which is probably due to the lack of conformation-dependent immunodominant epitopes on the recombinant protein expressed in E. coli. The specificity of the rec-gE-ELISA and affi-gE-ELISA was rather moderate (90% and 94%, respectively) because it was necessary to set such cut-off values in the tests that provided a maximum level of sensitivity, which obviously increased the incidence of false positive reactions. Though the indirect ELISAs detect antibodies against many epitopes of gE, the blocking gE-ELISA, which detects antibodies against only one immunodominant epitope of gE, showed a better test performance (specificity 99% and sensitivity 98%). This is most probably due to rather high dilutions of the sera used in the indirect gE-ELISAs (1:30) as compared to the serum dilution in the blocking gE-ELISA (1:2). We conclude that the indirect gE-ELISAs are sufficiently specific and sensitive to distinguish ADV-infected swine from those vaccinated with gE-negative vaccine and can be useful, in particularly affi-gE-ELISA, as additional tests for the detection of antibodies to gE.

Animals↗
PubMed2006

Analysis of the seroprevalence of bovine paratuberculosis and the application of modified absorbed ELISA to field sample testing in Korea.

Paratuberculosis (PTB) is a major disease problem worldwide, and causes major economic losses in the dairy industry. Although PTB has been reported in Korea, no studies have been conducted to determine its prevalence and no program has been developed to control the disease. In this study, the sera of beef (n = 1,056) and dairy cattle (n = 1,105) from all provinces in Korea were tested to determine the prevalence of PTB using two different ELISA: an 'in house' modified absorbed ELISA (P-ELISA) based on sonicated antigen from Mycobacterium avium subsp. paratuberculosis ATCC 19698, and a commercial ELISA (C-ELISA). Receiver operating characteristic analysis was used to determine the cutoff point for P-ELISA. Based on C-ELISA results, the area under the curve for P-ELISA was 0.913 (95% CI, 0.883 to 0.943). Using a cutoff point of 0.100, P-ELISA showed a sensitivity of 62.0% and a specificity of 93.7%. The kappa value and the percent agreement between the two ELISAs were 0.322 and 92.5%, respectively. Both ELISAs showed a significant correlation between age and seropositivity (p < 0.01). According to C-ELISA, 71 of 2,161 sera (3.3%, 95 CI, 2.6% to 4.1%) were test-positive. The national true prevalence of PTB was estimated to be 7.1%. The findings suggest that a control program should be implemented to limit the spread of this disease, and that P-ELISA could be used as a screening test that produces results similar to C-ELISA.

Animals↗
PubMed2006

Comparison of sensitivity and specificity in three commercial foot-and-mouth disease virus non-structural protein ELISA kits with swine sera in Taiwan.

Three commercialized ELISA kits for the detection of antibodies to the non-structural proteins (NSPs) of FMD virus were compared, using sera from uninfected, vaccinated, challenged and naturally infected pigs. The kinetics of the antibody response to NSPs was compared on sequential serum samples in swine from challenge studies and outbreaks. The results showed that ELISA A (UBI) and ELISA B (CEDI) had better sensitivity than that of the 3ABC recombinant protein-based ELISA C (Chekit). The peak for detection of antibodies to NSPs in ELISA C was significantly delayed in sera from natural infection and challenged swine as compared to the ELISA A and B. The sensitivity of the three ELISAs gradually declined during the 6-month post-infection as antibodies to NSP decline. ELISA kits A and B detected NSP antibody in 50% of challenged pigs by the 9-10th-day and 7-8th-day post-challenge, respectively. ELISA B and C had better specificity than ELISA A on sequential serum samples obtained from swine immunized with a type O FMD vaccine commercially available in Taiwan. Antibody to NSPs before vaccination was not detected in swine not exposed to FMD virus, however, antibody to NSPs was found in sera of some pigs after vaccination. All assays had significantly lower specificity when testing sera from repeatedly vaccinated sows and finishers in 1997 that were tested after the 1997 FMD outbreak. However, when testing sera from repeatedly vaccinated sows or finishers in 2003-2004, the specificity for ELISAs A, B and C were significantly better than those in 1997. This effect was less marked for ELISA A. The ELISA B was the best test in terms of the highest sensitivity and specificity and the lowest reactivity with residual NSP in vaccinates.

Animals↗
PubMed1997

Glycoprotein gE blocking ELISAs to differentiate between Aujeszky's disease-vaccinated and infected animals.

Three blocking ELISAs for the detection of antibodies against glycoprotein E (gE) of Aujeszky's disease virus (ADV) in sera (an indirect blocking ELISA (gE-iELISA), a direct blocking ELISA (gE-dELISA) and a two-site 'sandwich' blocking ELISA (gE-sELISA)) have been developed. The gE-ELISAs are based on monoclonal antibodies (mAbs) directed to gE and detect gE-specific antibodies in sera by blocking the binding of mAbs to one (in the gE-iELISA and the gE-dELISA) or two (in the gE-sELISA) epitopes of gE. From a panel of thirteen gE-specific mAbs, mAbs, their conjugates and the combination coating mAb/conjugate that optimally detect anti-gE antibodies in the assays were selected. Sera from uninfected unvaccinated swine or swine vaccinated against different swine viral disorders were negative by three gE-ELISAs, the blocking gB-ELISA, and VNT. Swine vaccinated with gE-negative vaccine were seropositive in the gB-ELISA and VNT but were seronegative by three gE-ELISAs. Infected unvaccinated swine, infected swine vaccinated with gE-negative vaccine, and swine vaccinated with gE-positive vaccine were detected as seropositive by three gE-ELISAs as well as in the gB-ELISA. The gE-dELISA and the gE-sELISA proved to be specific and the most sensitive and reliable assays to distinguish ADV-infected swine from those vaccinated with gE-negative vaccine. These two gE-ELISAs were at least as sensitive as the gB-ELISA for detecting ADV-specific antibodies.

Animals↗
PubMed2002

[Evaluation of a new ELISA (Bartels) for detection of Legionella pneumophila antigen in urine].

BACKGROUND: Detection of Legionella pneumophila soluble antigens allows rapid diagnosis of pneumonia caused by these bacteria. A new ELISA (Bartels) for antigenuria detection has recently been commercialized. We compared the new ELISA with another well-established ELISA (Binax). METHODS: To evaluate ELISA-Bartels (Legionella Urinary Antigen, Intracel, Issaquah, Washington, United States), urine samples previously characterized by ELISA Binax (Legionella Urinary Antigen Enzyme Immunoassay Kit, Binax, Portland, Maine, United States) were used. Samples came from Legionella outbreaks (n = 48), from sporadic legionellosis (n = 38), and from children with viral pneumonia (n = 21). Samples from the External Quality Control of Legionella of the European Working Group on Legionella Infections (n = 102) were also tested. Of the samples analyzed, 109 were positive in ELISA-Binax, 2 were equivocal and 98 were negative. Samples showing equivocal results were excluded from the analysis. RESULTS: The sensitivity of ELISA-Bartels in comparison with that of ELISA-Binax was 98.2% (107/109) and specificity was 82.7% (81/98). In the 17 samples that were positive in ELISA-Bartels and negative in ELISA-Binax, 10 were positive in ELISA-Binax after concentration by selective ultrafiltration and 6 further cases showed serology indicating or compatible with recent Legionella infection and were thus classified as true positives. CONCLUSIONS: ELISA-Bartels showed good sensitivity and specificity. Sensitivity was even higher than that of ELISA-Binax. Thus, we consider it to be an appropriate method for diagnosis of Legionella pneumonia.

Adult↗
PubMed2001

An enzyme-linked immunosorbent assay using immonoaffinity-purified antigen in the diagnosis of caprine paratuberculosis and its comparison with conventional ELISAs.

An enzyme-linked immunosorbent assay (APA-ELISA) using an immunoaffinity-purified antigen was developed and compared with the unabsorbed and absorbed ELISA procedures, using a crude antigenic preparation, for its efficacy in detecting antibodies in goat sera against Mycobacterium ovium paratuberculosis. Serum samples from 89 goats belonging to three different flocks, two with a history and evidence of paratuberculosis and one without it, were subjected to each ELISA, which had been standardized on known positive sera from goats experimentally infected with paratuberculosis. Faecal culture, faecal examination and histopathology were used as indicators of infection. The diagnostic sensitivities of the unabsorbed, absorbed and APA-ELISA were 81.8%, 77.3% and 77.3% and the specificities were 90.6%, 93.7% and 96.8%, respectively. The positive predictive values of APA-ELISA (94.4%) was the highest, followed by absorbed ELISA (80.9%) and unabsorbed ELISA (72.0%). The negative predictive values for APA-ELISA, absorbed ELISA and unabsorbed ELISA were 93.0%, 92.7% and 93.8%, respectively. The results indicated the value of APA-ELISA in avoiding the need to absorb individual test sera with Mycobacterium phlei and giving more consistent results than the absorbed ELISA. The APA-ELISA was also better than the other two procedures in terms of specificity and positive predictive values.

Animals↗
PubMed2003

Detection of malaria sporozoites by standard ELISA and VecTestTM dipstick assay in field-collected anopheline mosquitoes from a malaria endemic site in Ghana.

We compared the VecTestTM dipstick assay for detection of Plasmodium sporozoites in Anopheles vectors of malaria with standard circumsporozoite (CS) microplate ELISA for detection of Plasmodium falciparum circumsporozoite protein (PfCSP) in Anopheles mosquitoes. Mosquitoes were collected from a malaria endemic site (Kassena Nankana district) in northern Ghana. Of 2620 randomly sampled mosquitoes tested, the standard CS-ELISA gave a sporozoite rate of 10.8% compared with 11.2% by VecTestTM, which was not statistically different (P = 0.66). Visual reading of the CS-ELISA results gave a sporozoite rate of 13.4%, which was higher than the other tests (P > 0.05). To allow a more objective evaluation of the sensitivity of the dipstick, an additional 136 known CS-ELISA-positive specimens were analysed. The prevalence of the test (including the additional samples) was 14.6% and 14.7% for CS-ELISA and dipstick, respectively (P > 0.05). The estimated prevalence by visual assessment of the CS-ELISA results was 17.5%. The relative specificity and sensitivity of the VecTestTM dipstick and visually read ELISA were estimated based on the CS-ELISA as a gold standard. The specificities of the dipstick and visual ELISA were high, 98.0% and 96.6%, respectively. However, the sensitivities of the two assays were 88.8% for VecTest and 100% for visual ELISA (P < 0.01). Concordance between VecTest and CS-ELISA was good (kappa = 0.86). Similarly, there was a good concordance between the dipstick and the visually read ELISA (kappa = 0.88). Extrapolating from PfCSP controls (titrated quantities of P. falciparum sporozoites), mean sporozoite loads of CS-ELISA-positive An. gambiae (286 +/- 28.05) and An. funestus (236 +/- 19.32) were determined (P = 0.146). The visual dipstick grades showed high correlation with sporozoite load. The more intense the dipstick colour, the higher the mean sporozoite load (+ = 108, ++ = 207, +++ = 290, r = 0.99, r2 = 1). The VecTest dipstick offers practical advantages for field workers needing rapid and accurate means of detection of sporozoites in mosquitoes.

Animals↗
PubMed1999

Screening tests for antinuclear antibodies (ANA): selective use of central nuclear antigens as a rational basis for screening by ELISA.

Rational, fully automated ELISA screening tests for ANA have become accessible for routine use. Full length, recombinant or purified naturally expressed antigens can be selected to deliver well defined screening tests, designed to detect ANAs of established clinical significance, while unknown specificities with no clear biological impact are omitted. By comparing results of ANA screening by immunofluorescence (IIF) on tissue section with HEp-2 cells, a correlation of 88% was revealed, while 86% of HEp-2-positive sera bound tissue sections. Eighty-nine percent of HEp-2 ANA-positive sera bound in ANA ELISA I. Seventy-five percent of ANAs detected in ELISA I also bound in ELISA II. These correlations were statistically significant. ANAs detected in ANA ELISA I were all detected upon retesting in both ANA screening ELISA systems, and also by antigen-specific ELISAs. Specific ANAs defined by ELISA, immunodiffusion or Crithidia tests were detected in both ANA ELISA tests. These data demonstrate that most ANAs detected by immunofluorescence tests contain a dominating repertoire of specificities covered by the ANA ELISA systems. The sensitivity of the different ELISA tests can easily be adjusted to internationally defined consensus levels, and screening for ANA using both ELISA systems detects expected ANA specificities in ongoing international quality assessment programs.

Antibodies, Antinuclear↗
PubMed2003

Evaluation of dot-ELISA for serological diagnosis of amebiasis.

We improved the dot enzyme-linked immunosorbent assay (dot-ELISA) reported by Itoh and Sato, and assessed the usefulness of this test for the diagnosis of amebiasis. The sensitivity of dot-ELISA was compared with that of plate ELISA, the indirect hemagglutination test (IHA), and the indirect fluorescent antibody test (IFA) for the diagnosis of amebiasis. Of 37 serum samples from patients with documented amebiasis, 36 (97.3%) were positive by dot-ELISA. There was consistency among the results of dot-ELISA, plate ELISA, and IFA, although the positive rate of IHA was lower than that of the others (78.4%; 29 of 37 cases were positive). The specificities of dot-ELISA and plate ELISA were assessed using a total of 68 sera, collected from 38 patients infected with seven different parasites other than Entamoeba histolytica, 10 patients showing diarrhea or liver abscess without parasitic infection, and 20 healthy individuals. The two assays showed no false-positive results. There were no differences in sensitivity and specificity between dot-ELISA and plate ELISA. However, the dot-ELISA technique seems to be more feasible for clinical application than plate ELISA techniques, because the assay does not require any specific equipment.

Animals↗
PubMed1994

Clinical evaluation of a modified ELISA, using photobiotinylated DNA, for the detection of anti-DNA antibodies.

The measurement of anti-dsDNA antibodies is important for the diagnosis and the follow-up of patients with systemic lupus erythematosus (SLE). For routine detection of anti-dsDNA, the Farr assay and the immunofluorescence technique (IFT) on Crithidia luciliae proved to be very useful. The anti-dsDNA ELISA is not used for routine purposes in our institute since it is flawed by false-positive results due to binding of negatively charged (immune) complexes to the employed precoat (protamine sulphate). Recently, a new anti-dsDNA ELISA has been described in which photobiotinylated dsDNA is coated to streptavidin coated plates. To investigate whether this modified ELISA is more specific than the classical anti-dsDNA ELISA, we tested sera of patients with SLE (n = 51), myasthenia gravis (MG, n = 25), rheumatoid arthritis (RA, n = 25) and Sjögren's syndrome (SS, n = 23) and sera of healthy blood bank donors (BBD, n = 25). In both assays the sera of the SLE patients gave significantly higher values than the sera of healthy blood bank donors. In the classical ELISA, 84% of the sera from patients with RA and 28% of sera of patients with MG were found positive. For the modified assay the figures were 8% and 24%, respectively. This modified ELISA was further studied and clinically evaluated by comparing it with the classical anti-DNA ELISA and two other anti-DNA assays (Farr assay and IFT), using 500 sera sent to our institute for routine anti-DNA determination and sera of an additional 75 healthy blood bank donors. Quantitatively, both ELISAs showed the same high degree of correlation with the IFT. The modified ELISA gave a better correlation with the Farr assay than the classical anti-DNA ELISA. From our data we conclude that the ELISA using photobiotinylated DNA is a more reliable assay than the classical anti-DNA ELISA.

Antibodies, Antinuclear↗
PubMed1994

An improved enzyme linked immunosorbent assay (ELISA) for the detection of porcine serum antibodies against Mycoplasma hyopneumoniae.

An ELISA for the detection of serum antibodies to Mycoplasma hypopneumoniae in pigs and based on a 43 kDa purified protein derived from the cytoplasmic membrane of M. hyopneumoniae strain J is described. This ELISA (MHPP ELISA) was compared with another recently described (Auspharm ELISA, Sheldrake and Romalis 1992) that is based on column-purified sonicated proteins of strain J. Using sample to negative ELISA ratios of 3 and 4 as cutoffs for inconclusive and positive reactors respectively (compared to 2 and 3 for the Auspharm ELISA), the two tests had high specificity (MHPP 99.6%; Auspharm 100%) in 280 SPF pigs. In 176 pigs from commercial herds with endemic M. hyopneumoniae, the MHPP ELISA showed a higher sensitivity than the Auspharm ELISA in both high lung score (LS > or = 5) (85.5% vs. 69.9%) and low lung score (0 < LS < 5) (57.9% vs. 49%) pigs when the positive cutoff for each test was selected. The sensitivity when the inconclusive cutoff was selected was similar in both tests (85%; 85.7%) when low and high lung score pigs were pooled. Altough the MHPP also gave more positive reactors in 36 pigs from M. hyopneumoniae-infected herds with no lung pathology at slaughter than the Auspharm ELISA (11 vs. 4), the total number of inconclusive and positive reactors in these pigs was similar for both tests (18 vs. 14). The MHPP ELISA gave significantly higher ELISA ratios in infected pigs (up to 17.9) than the Auspharm ELISA (up to 9), and earlier seroconversion in naturally-infected (6-8 weeks vs. 9-10 weeks) and experimentally-infected pigs (2-4 weeks vs. 4-6 weeks post infection).

Animals↗
PubMed2005

Evaluation of three ELISAs for Mycobacterium avium subsp. paratuberculosis using tissue and fecal culture as comparison standards.

Three serum ELISAs for detection of antibodies against Mycobacterium avium subsp. paratuberculosis (Mptb) were evaluated against culture of tissue and feces samples from 994 dairy cows collected at slaughter. Culture of ileum and associated lymph nodes for Mptb were positive for 160 (16.1%) of the 994 cows and 36 (3.6%) were fecal culture-positive for Mptb. Two of the ELISAs evaluated were absorbed indirect assays and the third was a non-absorbed indirect assay. Estimated sensitivities of the absorbed ELISAs when compared to tissue culture were 8.8% and 6.9%, while the unabsorbed ELISA had a sensitivity of 16.9%. Specificities were 97.6%, 96.0% and 90.8%, respectively. When compared to fecal culture, the sensitivities of the absorbed ELISAs were 16.6% and 13.9%, respectively, and the sensitivity of the unabsorbed ELISA was 27.8%. Specificities were 97.1%, 95.9% and 90.1%, respectively. Area under the curve (AUC) of receiver operator characteristic curves for the absorbed ELISAs when tissue culture was the standard were 0.553 and 0.547, while the unabsorbed ELISA had an AUC of 0.540. When fecal culture was the comparison standard, the AUC of the absorbed ELISAs was 0.575 and 0.574, while the unabsorbed ELISA was 0.529. Overall, the sensitivities of the ELISAs when compared to tissue culture were low. The apparent advantage of the unabsorbed ELISA with respect to sensitivity is at the cost of lowered specificity and test accuracy.

Adsorption↗