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PubMed1993

Use of the cytomegalovirus antigenemia (CMV-Ag) assay for the detection of CMV in the blood of AIDS patients.

Direct specimen testing was performed on 186 peripheral blood specimens to identify the presence of antigen to cytomegalovirus (viz., the cytomegalovirus antigenemia (CMV-Ag) assay). Confirmatory testing was performed using the shell vial indirect immunofluorescence assay (SVA-IFA), the indirect immunoperoxidase assay (TC-IPA), and conventional tube culture isolation (TC-CPE). The primary reagent for the CMV-Ag assay consisted of anti-CMV monoclonal antibody directed against the internal matrix structural phosphoprotein (1C3; Clonatec-Biosoft, France). The 72-kDa early nuclear antigen (Dupont) was utilized in the SVA-IFA and the TC-IPA. All test systems received an equal number of polymorphonuclear leukocytes in the inoculum. CMV was detected and isolated from 30% (55/186) of the specimens evaluated by either one or a combination of the tests. Detection and (or) isolation of CMV from blood by the CMV-Ag assay, SV-IFA, TC-IPA, and TC-CPE occurred at a rate of 17 (31/186), 12 (22/186), 16 (29/186), and 26% (49/186). Three of 55 positive specimens were identified only by the CMV-Ag assay; each patient in question, however, had at least one previous CMV isolate. No significant differences in sensitivity occurred between the CMV-Ag assay, the SVA-IFA, or the TC-IPA. However, TC-CPE including the blind passage of all negative tube cultures yielded a significantly larger number of positive blood specimens than either of the rapid detection methodologies. The CMV-Ag assay encompasses the benefits of a nonculture system, is simple to perform and easy to read, permits a same-day diagnosis, and requires less reagents than the routinely used SVA-IFA or TC-IPA.(ABSTRACT TRUNCATED AT 250 WORDS)

Acquired Immunodeficiency Syndrome↗
PubMed2002

Rapid detection of avian pneumovirus in tissue culture by microindirect immunofluorescence test.

An indirect immunofluorescence (IFA) test with a 96-well, flat-bottomed microplate was developed to detect avian pneumovirus (APV) antigen in Vero cell cultures. Samples of nasal turbinates and swabs from infraorbital sinuses and trachea were collected from 4-week-old poults experimentally inoculated with APV. The APV titers by tissue culture IFA staining were compared with that of visual reading of cytopathic effect (CPE). The ability of IFA staining to detect APV antigen correlated well with visualizing CPE. The use of IFA staining of Vero cell cultures allowed detection of APV in substantially less time than the use of visualizing CPE. In addition, the use of IFA allowed specific identification of the virus in cell culture.

Animals↗
PubMed2003

The immunologic response of dogs to Bartonella vinsonii subspecies berkhoffii antigens: as assessed by Western immunoblot analysis.

Bartonella vinsonii subspecies berkhoffii is a recently recognized zoonotic pathogen that causes endocarditis, granulomatous rhinitis, and granulomatous lymphadenitis in dogs. Isolation of B. vinsonii (berkhoffii) from blood or tissue samples is frequently unsuccessful; therefore, diagnosis is primarily dependent on serologic or molecular testing modalities. Because previous canine serologic studies have used an indirect immunofluorescence assay (IFA), without Western immunoblot (WI) confirmation, the overall objective of this study was to examine the diagnostic use of WI for confirmation of B. vinsonii (berkhoffii) infection in dogs. To confirm that agar-grown and cell culture-grown organisms yielded similar patterns of WI antigenic protein recognition, the 2 preparations were compared using IFA-reactive sera obtained from dogs experimentally infected with B. vinsonii (berkhoffii). Temporal changes in the pattern of antigenic protein recognition were characterized using sera obtained from dogs at various time points after experimental B. vinsonii (berkhoffii) infection. The specificity of B. vinsonii (berkhoffii) WI was examined by testing canine sera that were reactive to B. henselae, B. clarridgeiae, Ehrlichia canis, Rickettsia rickettsii, Babesia canis, Anaplasma phagocytophilum (previously E. equi), or Brucella canis antigens. Clinical accessions including serum samples obtained from B. vinsonii (berkhoffii) culture-positive dogs and B. vinsonii (berkhoffii) culture-negative dogs that were IFA seroreactive to B. vinsonii (berkhoffii) antigens were examined by WI. The results of this study indicate that WI using agar-grown or cell culture-grown B. vinsonii (berkhoffii) antigens produce identical patterns of antigenic protein recognition. After experimental infection, there is a progressive increase in the number of antigenic proteins that are recognized by WI, with the 33-kD antigen representing the first and the most persistent antigen recognized by B. vinsonii (berkhoffii)-infected dogs. Regarding specificity, sera from dogs that were reactive to various heterologous antigens did not recognize B. vinsonii (berkhoffii) antigens by IFA or WI, and sera from dogs experimentally infected with B. henselae did not recognize B. vinsonii (berkhoffii) antigens by WI. Regarding clinical accessions, there was good agreement between B. vinsonii (berkhoffii) IFA test results and WI analysis. Western immunoblot analysis can be used to detect or confirm exposure to B. vinsonii (berkhoffii) in dogs.

Animals↗
PubMed2005

A highly immunogenic trivalent T cell receptor peptide vaccine for multiple sclerosis.

BACKGROUND: T cell receptor (TCR) peptide vaccination is a novel approach to treating multiple sclerosis (MS). The low immunogenicity of previous vaccines has hindered the development of TCR peptide vaccination for MS. OBJECTIVE: To compare the immunogenicity of intramuscular injections of TCR BV5S2, BV6S5 and BV13S1 CDR2 peptides in incomplete Freunds adjuvant (IFA) with intradermal injections of the same peptides without IFA. METHODS: MS subjects were randomized to receive TCR peptides/IFA, TCR peptides/saline or IFA alone. Subjects were on study for 24 weeks. RESULTS: The TCR peptides/IFA vaccine induced vigorous T cell responses in 100% of subjects completing the 24-week study (9/9) compared with only 20% (2/10) of those receiving the TCR peptides/saline vaccine (P =0.001). IFA alone induced a weak response in only one of five subjects. Aside from injection site reactions, there were no significant adverse events attributable to the treatment. CONCLUSIONS: The trivalent TCR peptide in IFA vaccine represents a significant improvement in immunogenicity over previous TCR peptide vaccines and warrants investigation of its ability to treat MS.

Adult↗
PubMed2006

Surveys on seroprevalence of canine monocytic ehrlichiosis among dogs living in the Ivory Coast and Gabon and evaluation of a quick commercial test kit dot-ELISA.

Canine monocytic ehlichiosis (CME), an enzootic disease in Africa, has been studied in canine blood samples (serum). These dogs, without any clinical sign of disease, were living in Abidjan (Ivory Coast) and in several small villages located in northeasst Gabon (Ogooué Ivindo). The results obtained by indirect fluorescent antibody (IFA) test, used as a point of reference, and by a quick test dot-Elisa were compared. Blood samples taken from 390 asymptomatic dogs in 2003 (137 in Ivory Coast and 253 in Gabon) were screened by IFA (antigen from Symbiotics Europe, Lyon) with a positive threshold set at 1/80. Afterwards, CME was detected by the commercial test kit Dot-Elisa in solid phase Snap 3Dx (Idexx, Westbrook, Maine, USA), using recombinant proteins which belong to Ehrlichia canis, p30 and p30-1. Using the IFA test, CME seroprevalence in the Ivory Coast is found to be 67.8%. Among 93 Ivorian seropositive blood samples, 76 samples show an antibody titer >1/2560. In Gabon, IFA showed that seroprevalence is only 3.1%. Among 8 seropositive Gabonese dogs, only one sample shows an antibody test titer> 1/2560. Results from the Snap 3Dx test used on 390 blood samples are 100 positive samples and 290 negative ones. Comparison between IFA and Snap test 3DX revealed that the Snap test shows 97.9% specificity, 93.1% sensitivity, a 94% positive predictive value, a 97.6%, negative predictive value, and 96.6% reliability. In conclusion, CME seroprevalence in Abidjan is very high. Dogs studied for CME were watchdogs, living in kennels, where infection transmitted by Rhipicephalus sanguineus seems to be higher in the Gabonese area called Ogooué Ivindo, where semi-stray dogs were subjected to the test. These dogs where carried ticks identified as Haemaphysalis leachi, but this kind of tick is not considered as bearing Ehrlichia canis. Finally, results of the Snap 3Dx show that it is a simple and reliablemeans for quickly detecting dogs suspected of asymptomatic canine ehrlichiosis.

Animals↗
PubMed1992

Involvement of the consensus sequence motif at coil 2b in the assembly and stability of vimentin filaments.

Nearly all intermediate filament (IF) proteins share two sequence motifs located at the N- and the C-terminal ends of their helical rod domain ('coil 1a' and 'coil 2b', respectively). To examine the structural role of the coil 2b motif, we have performed in vitro assembly studies and in vivo microinjection experiments employing two site-specific reagents: (a) a 20-residue synthetic peptide (C-2) representing the conserved motif itself and (b) a monoclonal antibody (anti-IFA) that recognises an epitope within the conserved coil 2b sequence. We demonstrate here that vimentin protofilaments, when induced to assemble in the presence of C-2 or anti-IFA, show a lower propensity to polymerise and yield various abberant structures. The few filaments that are formed under these conditions appear much shorter than normal IFs and are unravelled or aggregated. Furthermore, when preformed vimentin filaments are exposed to C-2 or anti-IFA, most of the normal IFs are converted into shorter filamentous forms that possess an abberant morphology. None of these effects is seen when vimentin subunits are coincubated with control peptides. Microinjection of anti-IFA into the cytoplasm of interphasic 3T3 cells provokes collapse of vimentin IFs into a juxtanuclear mass and formation of numerous amorphous aggregates distributed throughout the cytoplasm. These two effects are not seen when the anti-IFA is microinjected into the cell nucleus. Our results provide experimental evidence supporting previous suggestions for a role for the conserved coil 2b sequence in filament assembly. We propose that this region is interacting with other sites along the vimentin molecule and that these interactions are essential for proper protofilament-protofilament alignment and filament stability.

3T3 Cells↗
PubMed1987

Serodiagnosis of cilia-associated respiratory bacillus infection by the indirect immunofluorescence assay technique.

Antibody to cilia-associated respiratory (CAR) bacillus was detected by the indirect immunofluorescence assay (IFA) technique using tracheal sections of infected mice as antigen in serum samples collected from rats infected naturally and experimentally. Nine of 23 cases of natural infection were positive in IFA antibody, with titres ranging from 1:10 to 1:80, and all these antibody-positive cases were also histologically positive. The remaining 14 cases were negative in both IFA antibody and histological diagnosis, even though some of them were infected with Sendai virus and Mycoplasma pulmonis. In the experimental infection, serum samples collected from 18 rats on days 4, 7, 14, 21, 28 and 56 post-inoculation (PI) (three rats for each point) and examined for IFA antibody revealed that seroconversion occurred in one rat on day 14 PI and in three rats on day 21 PI. Antibody titres of 1:80 to 1:160 remained to the termination of the experiment. The IFA technique was useful for the diagnosis of CAR bacillus infection except in the early stage of the infection.

Animals↗
PubMed2004

Detection of antinuclear antibodies by solid-phase immunoassays and immunofluorescence analysis.

BACKGROUND: Antinuclear antibodies (ANAs) are associated with several inflammatory rheumatic diseases. The aim of the present work was to evaluate enzyme immunoassays (EIAs) and compare them with classic immunofluorescent analysis (IFA) for the detection of ANA. METHODS: Seven enzyme immunoassays were used in this study. All assays were applied as described by the manufacturers. Three populations were included in the study: (a) a population of patients with well-established autoimmune inflammatory disease (n = 102); (b) a population in which a rheumatic disease was diagnosed up to 5 years after an IFA was performed (n = 164); and (c) a population of consecutive outpatients suspected to have a rheumatic disease (n = 101). The current clinical diagnoses of the patients served as the standard against which performance of the assays was evaluated. RESULTS: In patients with well-established rheumatic disorders, the newly developed EIA in which HEp-2 extracts were included had sensitivities and specificities comparable to or in some instances better than the IFA. The assays without HEp-2 extracts included had significantly lower sensitivities and specificities. In the outpatient population, up to 51% of patients had positive ANA tests that did not correspond to classic ANA-associated disease. However, in the assays in which the HEp-2 extracts were not included, the false-positive rate was <10%. The false-negative rate judged against IFA differed from assay to assay and disease to disease and was mostly <10%. CONCLUSIONS: In this study, the sensitivities of EIAs and IFA were largely comparable. However, EIAs without HEp-2 extracts included had a low sensitivity but a high specificity, particularly in nonselected populations. The choice of test is highly dependent on the clinical setting in which the ANA test is to be used and on laboratory policy.

Adolescent↗
PubMed2003

Clinical and cerebrospinal fluid (CSF) profile and CSF criteria for the diagnosis of spinal cord schistosomiasis.

OBJECTIVES: To describe the clinical and CSF findings among patients with presumptive neuroschistosomiasis (NS) and to suggest a classification for the CSF diagnosis of presumptive NS. METHOD: The charts of all patients whose CSF exam was performed at the CSF Lab, Jos Silveira Foundation, Salvador, Brazil, from 1988 to 2002 were reviewed. Those with clinically suspected NS whose indirect fluorescent antibody test (IFA) and or hemagglutination-inhibiting antibodies test (HAI) were positive to S. mansoni were identified. RESULTS: Of 377 patients, 67.9% were males; the median age was 36 years (mean 37 + 16 yrs, range 3-82 yrs). The most frequent complaints were paraparesis (59.9%), urinary retention (36.2%), lower limb pain (22.8%). WBC of CSF (count/mm ) was > 4 in 66.0% (mean 83 + 124, median 40, range 4.3-1,100), protein (mg/dl) was > 40 in 84.6% (mean 185 + 519, median 81, range 41-6,800) and eosinophils were present in 46.9%. IFA and HAI were positive in 75.3%. WBC > 4 and presence of eosinophils were associated with IFA and HAI positive (67.3% versus 51.4%, p 0.014; 49.1% versus 23.0%, p 0.0001, respectively) and protein > 40 was not (85.4% versus 77.0%, p 0.09). Presence of WBC > 4, protein > 40 and eosinophils was associated with IFA and HAI positive (71.6% versus 38.2%, p 0.0003) but presence of eosinophils and any other combination of WBC and protein were not. CONCLUSION: NS should be considered as a possible diagnosis in patients who had had contact with schistosome-infected water and present with spinal cord compromising. Presence of IFA and HAI positive to S. mansoni, WBC > 4, protein > 40 and presence of eosinophils in the CSF may be considered as a criterium of highly probable presumptive diagnosis.

Adolescent↗
PubMed2004

Comparison of results for serologic testing and a polymerase chain reaction assay to determine the prevalence of stray dogs in eastern Tennessee seropositive to Ehrlichia canis.

OBJECTIVE: To determine the prevalence of stray dogs in eastern Tennessee seropositive to Ehrlichia canis and examine the correlation between results for an ELISA, indirect immunofluorescent antibody (IFA) test, and polymerase chain reaction (PCR) assay. SAMPLE POPULATION: Blood samples obtained from 90 adult dogs admitted to an animal shelter in eastern Tennessee. PROCEDURE: Serum samples were analyzed for antibodies against E. canis by use of a commercially available ELISA kit, 2 IFA tests, and a PCR assay; testing was performed at the University of Tennessee (TN) and North Carolina State University (NCSU). The PCR amplification was performed by use of DNA extracted from EDTA-anticoagulated blood and primers designed to amplify DNA of Ehrlichia spp. RESULTS: Antibodies against E. canis were detected in only 1 dog by use of the ELISA. By IFA testing at TN, 10 of 90 (11%) dogs were seroreactive against E. canis antigens, all of which had medium to high titers to E. canis. Only 5 of the 10 TN seroreactors were also reactive against E. canis antigens in IFA tests conducted at NCSU, and all 5 had low to medium titers. The DNA of Ehrlichia spp was not amplified in any blood samples by use of PCR assays conducted at the TN or NCSU. CONCLUSIONS AND CLINICAL RELEVANCE: The discordant ELISA, IFA, and PCR results obtained in this study were unexpected and may have been related to exposure of dogs to an Ehrlichia species other than E. canis, such as E. ewingii.

Animals↗
PubMed1987

[Detection of IgG antibodies with immunofluorescent antibody technique in human trichomoniasis]

The indirect fluorescent antibody(IFA) test was used to detect serum IgG and IgM antibodies to Trichomonas vaginalis in 31 vaginal trichomoniasis, 7 candidiasis and in 20 non-infected healthy wonem with antigen prepared from axenic culture of Trichomonas vaginalis isolated from vulvovaginitis patient. The results were as follows: In 31 vaginal trichomoniasis the positive reactions of IgG antibody were 27 in the 1/8 dilution or higher and 4 in the 1/4 dilution whereas in healthy women the reaction showed signigicantly low as in the 1/4 dilution of below. The sensitivity and specificity of IFA test for IgG antibody to trichomonad antigen in this study were 87.1% and 100%, respectively. No significant difference of IgM antibody levels between vaginal trichomoniasis and healthy women was observed. No relation between the levels of IgG and IgM antibodies to trichomonad antigen by IFA test was observed. No relation between the time lapse and the level of serum IgG antibodies in IFA test of vaginal trichomoniasis was regarded. In conclusion the present study suggests that IFA test in trichomoniasis could be a useful tool for detection of anti-trichomonad IgG antibodies and applicable as an immundiagnostic method.

Journal Article↗
PubMed1997

Antibodies against equine herpesviruses in free-ranging mountain zebras from Namibia.

Twenty-one blood samples of free-ranging mountain zebras (Equus zebra) from Namibia were tested for equine herpesvirus (EHV-1, -2, -3, -4) specific antibodies by immunofluorescence assay (IFA) and neutralization test (NT). Additionally, type-specific nested polymerase chain reactions (nested PCR) were employed for detection of EHV-1, -2 and -4 DNA. Equine herpesvirus-1 antibodies were detected by IFA in all zebras, while only seven serum samples contained EHV-4 IFA antibodies. Sera with high IFA antibodies also were found to neutralize EHV-1 and -4. Furthermore, 20 zebras were EHV-2 seropositive by IFA, and one zebra had EHV-2 neutralizing antibodies. Equine herpesvirus-3 specific antibodies were not detected. We did not amplify EHV-1, -2 or -4 specific DNA sequences in peripheral blood leukocytes of the same zebras using type-specific nested PCR. EHV infections appear to be widespread in free-ranging zebras, as they are in domestic horses.

Animals↗
PubMed2000

Circulating anticentromere CENP-A and CENP-B antibodies in patients with diffuse and limited systemic sclerosis, systemic lupus erythematosus, and rheumatoid arthritis.

OBJECTIVE: To determine the disease sensitivity and specificity of testing for autoantibodies against 2 of the 3 main human centromere antigenic components, CENP-A and CENP-B (recombinant, expressed in baculovirus). METHODS: ELISA with CENP-A and CENP-B antigens were used to test 45 sera showing a centromere pattern by immunofluorescence (IFA) and sera from 96 patients with systemic sclerosis (SSc), subdivided into diffuse (dSSc) and limited (lSSc) forms. For controls, the same tests were performed on sera from 100 patients with rheumatoid arthritis (RA), 100 with systemic lupus erythematosus (SLE), and 50 random blood donors. Sera from all the patients with SSc were also tested for the presence of anti-Scl70 antibody by ELISA (bovine antigen), and for pattern and titer by IFA (HEp-2 cells). RESULTS: Of the 45 IFA positive sera, 93% were positive for anti-CENP-A and 91% for anti-CENP-B. There was a very good quantitative correlation between the antibody levels against these 2 centromere components (r = 0.597; p<0.001). Anti-CENP-A and B were found in 48% of patients with lSSc, and in 11% and 9%, respectively, of those with dSSc. The difference in the frequency of anti-CENP-A between the 2 patient groups was significant (chi-squared, p<0.001). Similar levels of anticentromere staining pattern by IFA were observed for these 2 groups. Anti-Scl70 was elevated in 8% of lSSc and 25% of dSSc patients; this difference was also significant (chi-squared, p = 0.02). Neither CENP-A nor CENP-B reacted with IgG from SSc patients containing anti-Scl70. The frequency of abnormal levels in patients with SLE and RA was, respectively, 11% and 3% for anti-CENP-A and 4% and 3% for anti-CENP-B. The reaction of IgG from SLE and RA patients with CENP-A was not inhibited by histone H3, i.e., it was not due to recognition of the histone-like domain in CENP-A. Thus, when 96 SSc patients were compared to 200 patients with RA and SLE, the disease specificity of anti-CENP-A and B was 93% and 96.5%, respectively. CONCLUSION: In addition to IFA, ELISA tests for CENP-A and CENP-B yield results with similar sensitivity and specificity for the diagnosis of SSc. CENP-A and CENP-B are primarily associated with lSSc. In SSc the autoantibody response is directed simultaneously and with similar amplitude against these 2 components of the centromere structure, whereas in other autoimmune diseases the response is directed mainly against one of the 2 components.

Adult↗
PubMed2000

HEp2 ANA EIA: a new fully automated assay for the screening of antinuclear antibodies.

BACKGROUND: Screening for antinuclear antibodies is performed for various systemic autoimmune diseases with various different techniques. A more recent approach to ANA screening is the use of enzyme immunoassays. OBJECTIVES: To introduce the new COBAS CORE HEp2 ANA EIA, the first fully automated enzyme immunoassay for the detection of ANA in serum and plasma of patients with suspected systemic autoimmune disease. METHODS: A method comparison between the new HEp2 ANA EIA and immunofluorescence assays on HEp2 cells was performed in two groups--977 patients with systemic autoimmune disorders and 952 healthy subjects with no autoimmune disease-related diagnosis. RESULTS: The clinical sensitivity in the patient group was 79.8% for HEp2 ANA EIA and 89.5% for IFA, and the clinical specificity in the control group was 91.7% for HEp2 ANA EIA and 93.9% for IFA. The analysis of 830 routine samples showed an overall good concordance of 85% between both assays. Further examination of the discrepant results showed 8% discrepant negative results for the HEp2 ANA EIA assay compared to 21% for IFA. Discrepant positive results were found in 37.9% and 33% of samples for the HEp2 ANA EIA and the IFA, respectively. CONCLUSION: The HEp2 ANA EIA is the first fully automated assay for the detection of ANA in serum and plasma. The assay shows improved sensitivity in ANA determination and a similar performance, compared to the standard method of IFA.

Antibodies, Antinuclear↗
PubMed2003

Selective detection of autoimmune antibodies to single- and double-stranded DNA by enzyme immunoassay.

Presence in serum of anti-double-stranded deoxyribonucleic acid antibodies (anti-dsDNA) is one of the diagnostic criteria for systemic lupus erythematosus. Anti-single stranded DNA antibodies (anti-ssDNA) also occur, but their clinical significance is unclear. Use of enzyme immunoassay (EIA) kits that are specific for anti-dsDNA is desirable but problematic, since preparation of dsDNA of high integrity is difficult (ie, regions of ssDNA may persist). This study evaluated an EIA kit that uses low Mw plasmid/bacteriophage DNA as a nucleic acid source. Anti-dsDNA results were compared to results obtained by an anti-ssDNA EIA and an immunofluorescent assay (IFA) that uses Crithidia luciliae (specific for anti-dsDNA). Consecutive serum samples (n=139, 88% female) submitted to the clinical laboratory for anti-dsDNA analysis were evaluated. EIA precision was determined at three levels. Intra-assay precision [mean +/- SD (CV)] for anti-dsDNA: 36 +/- 3.5 IU/ml (9.8%); 98 +/- 4.4 (4.5%); and 245 +/- 5.8 (2.4%); and anti-ssDNA. 40 +/- 1.4 U/ml (3.5%); 190 +/- 4.8 (2.5%); and 283 +/- 10.3 (3.7%) (n=8). Inter-assay precision for anti-dsDNA: 36 +/- 6.3 IU/ml (17.5%); 90 +/- 5.9 (6.6%); and 207 +/- 20.9 (10.1%); and anti-ssDNA: 48 +/- 8.2 U/ml (17.0%); 193 +/- 12.7 (6.6%); and 263 +/- 21.5 (8.2%) (n=8). Linearity was assessed with (a) high dsDNA/high ssDNA and (b) low dsDNA/high ssDNA samples (no high dsDNA/low ssDNA samples were identified). Linearity (>200 IU/ml) was found for both sample types with a correlation coefficient (r) of 0.995-0.999. Anti-dsDNA immunoreactivity was not apparent with the low dsDNA/high ssDNA sample. More patients were positive for anti-ssDNA (54%), compared to anti-dsDNA (17%). IFA confirmation (Crithidia) indicated a relative sensitivity and specificity of 94.1% and 93.4% for the anti-dsDNA EIA. IFA positivity correlated with increased anti-dsDNA level: 20% (30-60 IU/ml); 70% (61-200 IU/ml); and 89% (>200 IU/ml). Of specimens that were anti-ssDNA positive and anti-dsDNA negative (n=51), only one was IFA positive. When IFA was compared to an anti-dsDNA EIA kit that used high Mw calf thymus DNA, lower relative specificity (88.2%) and sensitivity (72.6%) was obtained. Anti-ssDNA was found in many false positive specimens (87%).

Adult↗
PubMed2003

[Serum antibodies detection for serological diagnosis of severe acute respiratory syndrome].

OBJECTIVE: To evaluate the significance of serum specific antibodies against severe acute respiratory syndrome (SARS) virus in the diagnosis of SARS and the concordance of IFA and ELISA in the antibodies measurements. METHODS: Specific antibodies of IgM and IgG against SARS virus in 267 serum specimens from patients with SARS and 132 from non-SARS patients or healthy persons were assayed with IFA or ELISA. The sensitivity, specificity, positive predictive value (ppv), negative predictive value (NPV), and accuracy were used to evaluate the significance of the antibodies in diagnosis of SARS. The concordance of different measurements was tested with Kappa. RESULTS: The positive rate of specific IgM and IgG detected by IFA increased sharply on the eleventh day of the disease. The sensitivity, specificity, PPV, NPV, and accuracy of IgM in diagnosis of SARS on eleventh day or later were 65.6%, 100.0%, 100.0%, 71.0% and 81.3%, respectively, and IgG were 91.1%, 97.0% 97.3%, 90.1%, and 93.8%, respectively. Similar results could be seen when IgM and IgG were detected by ELISA. The Kappa of IFA and ELISA for IgM and IgG were 0.640 and 0.779 respectively. CONCLUSIONS: The antibodies against SARS virus can be detected in the mid-and late stage of SARS. The antibodies can be used in the serological diagnosis of SARS. There is high concordance of IFA and ELISA results in measurement of SARS antibodies.

Adult↗
PubMedDate not supplied

[Comparison of the methods used for the detection of antinuclear antibodies].

The production of one or more autoantibodies in the majority of rheumatic diseases is a common clinical finding. Indirect immunofluorescent antibody (IFA) test is the former and widely used method in detecting autoantibodies. Recently, commercial enzyme immunoassays (EIA) and immunoblot (IB) assays are available for the routine laboratories. The aim of this study was to compare the efficacies of these methods in 121 sera samples which were collected from 46 male and 75 female patients, of whom antinuclear antibody (ANA) testing were requested by clinicians. All the samples were simultaneously studied by IFA with HEp-2 and Crithidia lucillae cells, and by two different commercial kits of EIA and IB, and the results were evaluated. When IFA is accepted as reference method, the sensitivities and specificities of the other methods were found as follows, respectively; 82-100% and 84-100% for ANA-EIA, 48-54% and 98-100% for ANA-IB, 66-100% and 97-98% for anti-dsDNA-EIA, and 50-66% and 86-100% for anti-dsDNA-IB. The agreement rates of ANA-EIA, ANA-IB, anti-dsDNA-EIA and anti-dsDNA-IB methods with IFA were detected as 82-90%, 74%, 96-97%, and 84-98%, respectively. As a result, it may be concluded that IFA test should be used as a primarily test for the detection of ANA and anti-dsDNA autoantibodies, while EIA and IB methods are useful for antigen differentiation and confirmation of the results.

Antibodies, Antinuclear↗
PubMed2003

[Development and application of an indirect immunofluorescence assay for the detection of serum immunoglobulin G to human herpesvirus 6].

AIM: To establish an indirect immunofluorescence assay(IFA) for detection of serum IgG antibody against human herpesvirus 6(HHV-6). METHODS: Cord blood mononuclear cells infected by HHV-6 strain CN(5) were used to prepare cell antigen smear, so as to establish an IFA and make an epidemical investigation on serum specimens of child-bearing age, women. RESULTS: The specificity of the IFA for HHV-6 was confirmed by absorption assay(test). The IFA detection showed that in serum specimens from 116 cases of child-bearing age, women, the positive rate of anti-HHV-6 IgG was 72.4% and geometric mean titer(GMT) was 1:61. The positive rate and GMT of serum anti-HHV-6 IgG had no differences between pregnant women and unpregnant women, neither among pregnant women at different pregnant stages. CONCLUSION: A specific IFA has been developed successfully for epidemical investigation of HHV-6 infection rate in child-bearing women.

Antibodies, Viral↗