Positive EIA Research Articles

DNA Amplification for Diagnosis of C. difficile Infection: The Solution to False Positive Cytotoxin EIA Testing

DNA Amplification for Diagnosis of C. difficile Infection: The Solution to False Positive Cytotoxin EIA Testing

P1-S6.09 An audit of management of prenatal syphilis serology in the STI Clinic, Calgary, AB, Canada

BackgroundOutbreaks of syphilis have been occurring across Canada since 2001, with the province of Alberta having the 2nd highest incidence in 2007. As a result, there has been a national...

S11.2 Which algorithm performs better, screening with a non-treponemal or treponemal test?

BackgroundA growing number of diagnostic laboratories have recently adopted treponemal EIA tests that permits automation for syphilis screening thus reducing time and labor. This leads to a reverse sequence approach...

The construction of new immunopotentiating MUC1-VNTR-C144 DNA vaccine for pancreatic cancer

Objective To construct new MUC1-VNTR-C144 DNA vaccine for pancreatic cancer and study both CTL responses and antibody responses specific to VNTR enhanced by the C144 gene. Methods DNA encoding the 144 amino acids of the N-terminus of HBV core gene was cloned and then inserted into the recombinant plasmid pcDNA3.1-VNTR/Myc-his( + ) A. C57BL/6 mice were randomly immunized intramusscularly into anterior tibialis muscle with 100 μg plasmid DNA encoding VNTR ( V group,n = 15) or VNTR/CI44 (VC group,n = 15). Mice injected with the plasmid pcDNA3.1-C144/ Myc-his( + ) A vector (C group, n = 15 ) were used as controls. Four weeks later, all mice were injected a-gain with the same solution as before. Two weeks after the last immunization, spleen cells were obtained and analyzed for cytotoxic activity 6 days after in vitro stimulation by the synthesized VNTR polypeptide (Calcein AM assay). Blood was collected for the ELISA assay of anti-VNTR antibodies. Results The specific CTL killing rate against VNTR polypeptide induced in the VC group (46.7±8.2)% was signifi-cantly higher than in the V group (34.8±3.1 ) % and the C group ( 12.9±1.2) % ( P < 0.01 ) ,indicating the C144 gene enhances the CTL response induced by the vaccine. However,CTL lysed much more VNTR positive EIA than VNTR negative EIA ( P < 0.01 ), while the anti-VNTR antibody VU 3C6 significantly restrained such activity ( P <0.01 ) ,which indicated the CTL response was specific to VNTR. The equiva-lent concentration of specific antibody against VNTR in the VC group (2810.2±308.3 ) mg/L was signif-icantly higher than the V group (2323.5±238.3) mg/L and the C group (2130.9±138.5) mg/L (P < 0.01 ),which implied that the C144 gene could enhance the antibody response induced by the vaccine. Conclusion The VNTR specific CTL response and antibody response induced in mice immunized with new MUCl-VNTR-C144 DNA vaccine for pancreatic cancer can be enhanced significantly by the C144 gene. Key words: Pancreatic carcinoma; DNA vaccine

Clinical Features That Affect Indirect-Hemagglutination-Assay Responses to Burkholderia pseudomallei

Melioidosis, a disease endemic to northern Australia and Southeast Asia, is caused by the soil saprophyte Burkholderia pseudomallei. The indirect hemagglutination assay (IHA) is the most frequently used serological test to help confirm exposure to the causative organism. However, despite culture-confirmed disease, patients often have a negative IHA result at presentation and occasionally fail to seroconvert in serial testing. We retrospectively examined results for all patients with culture-confirmed melioidosis from our laboratory between January 1996 and August 2008. One hundred forty patients had a recorded IHA titer at presentation, 71 of which were positive at a titer of 1:40 or greater. Fifty-three patients went on to have subsequent IHAs 1 month or more after presentation. The relationships between IHA responses and clinical features were examined. The presence of bacteremia was significantly associated with a negative IHA at presentation. The coexistence of diabetes was associated with the presence of a positive IHA at presentation. In total, 14 patients (26%) demonstrated persistently negative IHA titers upon serial testing. No clinical factors were found to be significantly associated with this phenomenon. Supplementary testing using melioidosis-specific immunoglobulin G by EIA demonstrated different effects, with only Aboriginal or Torres Straits Islander ethnicity being significantly associated with a positive EIA at presentation. Reasons for these findings are examined, and directions for future research are discussed.

一重症心身障害児施設における麻疹ウイルス抗体価と麻疹感染免疫に関する一考察

Neutralizing antibody (NA), hemagglutination inhibition antibody (HI), and antibody assayed by IgG-enzyme immunoassay (EIA) against measles were tested as a set on sera of 119 out of 120 patients institutionalized for severe motor and intellectual disability and correlations among the 3 types of antibody titers were examined. 1) NA, HI, and EIA titers correlated positively. 2) If NA, HI, and EIA antibody titers were 4 (2(2)) or more, NA, HI, and EIA serum titers were positive. If they were 8 (2(3)) or more, all sera were positive for NA. If light absorption was 4 or more, serum HI was also positive for EIA. 3) Nine cases were certified as positive for measles infection, and 3 of these were vaccinated 2-3 years after infection. All 9 had positive NA and EIA even 15 years or more after infection. Of 29 cases certified as negative for measles infection and injected with measles vaccine, 21 (72.4%) had positive NA and 16 (72.4%) had positive HI. In EIA, 28 of the 29 (96.6%) showed positive. The only EIA-negative case was also the only one negative for both NA and HI, i.e., an 18-year-old man suffering from chromosomal aberration, 21-ring trisomy, and suspected of being a low responder to measles antigens, including NA, HI and EIA antigens. 4) The above facts suggest that EIA is a more sensitive test for positive history of wild measles virus or measles vaccine virus contact, making it possible to detect measles or measles vaccine injection. To prevent nosocominal infection, it is important to know whether institutionalized individuals are immune to measles, but many have no clear history of measles or measles vaccine injection, especially those 40 years old or older. 5) Institutionalized individuals 40 years old or more numbered 45. Their antibody titers against measles were positive, 82.2% in NA, 48.9% in HI and 91.1% in EIA. The high positive EIA rate suggests that most would sufferered from measles before institutionalization, because they had little chance of measles vaccination as children and little possibility of measles infection in the institution at nosocomial infection currence in 1983, which was limited in other ward, and no nosocomial infection of measles has been experienced in this institution during this more than 20 years.

HIV-2 diagnosis and quantification in high-risk patients

Current diagnostic assays for HIV-1 do not always test for the presence of HIV-2 in the United States. We present the case of a patient from Cape Verde, who was admitted to our hospital with rapidly deteriorating neurological function and multiple white matter lesions on MRI likely secondary to progressive multifocal leukoencephalopathy (PML). Initially, the patient had a positive EIA for HIV, but a negative HIV-1 Western Blot and no viral load detected on a branched-DNA assay. A repeat viral load by reverse transcriptase methodology (RT-DNA) detected 121,000 copies and an HIV-2 Western Blot was positive. The case highlights an extremely rare presentation of HIV-2 with severe neurological disease. We discuss the different tests available for the diagnosis and monitoring of HIV-2 in the United States.

Diagnostic performance of a rapid antigen test for RSV in comparison with a 19-valent multiplex RT-PCR ELISA in children with acute respiratory tract infections

A rapid and reliable diagnosis of respiratory syncytial virus (RSV) infection in childhood is very important for clinical management. In this study we compared a rapid antigen test (enzyme immunoassay, EIA) for the detection of RSV with a reverse transcriptase polymerase chain reaction (RT-PCR) [19-valent multiplex RT-PCR enzyme-linked immunosorbent assay (ELISA)] to assess the diagnostic performance. Furthermore the diagnostic value of the EIA in terms of age and season relation was analyzed. A total of 400 nasopharyngeal or tracheal secretions from pediatric patients with clinical signs of lower respiratory tract infection were included. The specimen had to be taken in a standardized manner within 72 h after admission. Specimens were tested in parallel with the EIA and the multiplex RT-PCR ELISA. The RT-PCR technique was used as the target assay. The EIA reached a sensitivity of 58% and a specificity of 90% for all samples tested. For patients < [corrected] 1 year the post-test probability for a positive EIA was 91% during the RSV season; a negative test result decreased disease probability from 53 to 25%. For older patients a positive test raised disease probability from 23 to 45% during the RSV season. Negative test results did not markedly change disease probability.

Evaluation of the Dako IDEIA norovirus EIA assay for detection of norovirus using faecal specimens from Australian gastroenteritis outbreaks

New techniques for detection of norovirus, a major cause of gastroenteritis, require ongoing evaluation. The aim of this study was to use material from gastroenteritis outbreaks in Victoria, Australia, to evaluate the sensitivity and specificity of the Dako IDEIA norovirus EIA assay, using both photometric and visual analysis. A total of 130 faecal specimens from 41 gastroenteritis outbreaks were tested for norovirus by electron microscopy (EM), a two-round multiplex reverse transcription-polymerase chain reaction (RT-PCR) method and the IDEIA norovirus assay. All specimens with sufficient amplified product were sequenced to determine their norovirus genotype. In addition, six well-established RT-PCR protocols were used to test four EIA-positive, multiplex RT-PCR/EM-negative specimens. Also, a range of RT-PCR protocols was used to test a specimen positive for GII only by the multiplex RT-PCR but positive for GI and GII by the EIA. The effect of multiple freezing-thawing cycles on EIA positivity was tested on seven additional specimens. A further seven specimens, known to contain the gastroenteritis viruses sapovirus, adenovirus, astrovirus and rotavirus were also tested by the IDEIA norovirus assay. The IDEIA norovirus assay gave a single-specimen sensitivity and specificity of 66% and 85%, respectively (visual analysis compared with the multiplex RT-PCR), 63% and 88% (photometric analysis compared with the multiplex RT-PCR), 65% and 87% (visual analysis compared with the multiplex RT-PCR and/or EM) and 62% and 90% (photometric analysis compared with the multiplex RT-PCR and/or EM). None of the four EIA-positive specimens negative by the multiplex RT-PCR and/or EM was positive by any of the six alternative RT-PCR protocols. The specimen positive for GI and GII by EIA but for GII only by the multiplex RT-PCR was not positive for GI by any of the alternative RT-PCR protocols. A minimum of three specimens per outbreak had to be tested by the EIA to ensure that norovirus-positive outbreaks (multiplex RT-PCR and/or EM) were classified as positive for norovirus by the IDEIA norovirus assay (visual or photometric analysis). However, one specimen from a norovirus-negative outbreak (multiplex RT-PCR and/or EM) for which four specimens were provided was positive for norovirus by the IDEIA norovirus assay. Seven norovirus genotypes were identified by open reading frame 1 sequencing analysis and specimens from all seven norovirus genotypes (as well as an EM-positive/multiplex RT-PCR-negative specimen) were detected by the IDEIA norovirus assay by both visual and photometric analysis. Repeated freezing-thawing cycles (up to six) for faecal specimens did not reduce the sensitivity of the EIA assay but could render an EIA-negative specimen EIA-positive. The specimens positive for sapovirus, adenovirus, astrovirus and rotavirus were EIA-negative. The IDEIA norovirus assay lacks the sensitivity and specificity to ascribe a particular result to a particular specimen, but could be useful for detecting norovirus in a gastroenteritis outbreak where specimens are plentiful, although it is difficult to avoid a risk of false positives. Since visual analysis can be used for result assessment almost as reliably as photometric analysis, the test kit would be useful for laboratories lacking specialist equipment such as a photometric microplate reader.

Detection of Aspergillus Galactomannan Antigenemia To Determine Biological and Clinical Implications of Beta-Lactam Treatments

Detection of Aspergillus galactomannan (GM) in serum with the Platelia Aspergillus enzyme immunoassay (EIA) is useful for diagnosing invasive aspergillosis. From May 2003 to November 2004, 65 patients who did not develop aspergillosis had at least two positive sera while receiving a beta-lactam treatment (GM index [GMI], >or=0.5). Of the 69 treatment episodes scored, 41 consisted of a beta-lactam other than piperacillin-tazobactam (n=29), namely, amoxicillin-clavulanate (n=25), amoxicillin (n=10), ampicillin (n=3), or phenoxymethylpenicillin (n=2). In all cases, antigenemia became negative 24 h to 120 h upon stopping the antibiotic. Monitoring of 35 patients, including 26 with hematological malignancies, revealed three antigenemia kinetic patterns: each was observed with any drug regimen and consisted of a persistent GMI of >2.0 (65.7%), >0.5, and <or=1.5 (25.7%) or a variable GMI (14.3%) from the onset of antibiotic therapy. All available drug batches given to 26 patients cross-reacted with the EIA. Galactomannan titration in batches failed to predict the GM titers in the five patients studied at cumulative doses of ampicillin or amoxicillin-clavulanate, regardless of the time lapse between serum sampling and infusion period. Our results show that beta-lactams other than piperacillin-tazobactam may lead to false presumption of aspergillosis. The resulting kinetic patterns of GM antigenemia are variable, and sampling serum prior to the next beta-lactam dose may not decrease GMI below the threshold. Consequently, testing of suspected antibiotic batches remains the only indicator of possible false EIA positivity.

Dark ground microscopy and treponemal serological tests in the diagnosis of early syphilis

Objectives: To evaluate the use of dark ground microscopy (DGM) and treponemal serological tests in the diagnosis of primary (PS) and secondary (SS) syphilis. Methods: A retrospective case note review...

Immunoblot as a supplemental test to detect antibodies to hepatitis C virus in blood donors

Supplemental tests using Immunoblot are recommended to improve specificity of anti-HCV by ELISA. In Brazil immunoblot is not officially recommended. Aiming to identify EIA false-positive rate 70 positive EIA anti-HCV blood donors were submitted to 3rd generation immunoblot at Hemocentro of Mato Grosso State where polymerase chain reaction tests are not performed. There were 44 (62.9%) immunoblot-positive, 22 (31.4%) negative and 4 (5.7%) indeterminate. Anti-HCV immunoblot can distinguish blood donors with false-positive ELISA from those who need medical assessment. Our data suggest that immunoblot could be useful in Brazilian blood banks where molecular biology tests are not available.

Longevity of antibody responses to a Salmonella typhi-specific outer membrane protein: interpretation of a dot enzyme immunosorbent assay in an area of high typhoid fever endemicity.

The objective of this study was to investigate the longevity of positive dot enzyme immunosorbent assay (dot EIA) results for IgM and IgG to a Salmonella typhi outer membrane protein in Malaysian children with enteric fever. The patients were children one month to 12 years of age with clinical evidence of typhoid fever, positive blood or stool cultures for S. typhi, and/or a positive Widal test result who were admitted over a two-year period to General Hospital (Kota Bharu, Malaysia). These patients received standard inpatient treatment for enteric fever including chloramphenicol therapy for 14 days. Dot EIA tests were performed as part of clinical and laboratory assessments on admission, at two weeks, and then at 3, 6, 9, 12, 15, 18, and 21 months postdischarge. Assessment of the longevity of positive dot EIA IgM and IgG titers was done by Kaplan-Meier analysis. In 94 evaluable patients, 28% were dot EIA IgM positive but IgG negative on admission, 50% were both IgM and IgG positive, and 22% were IgM negative and IgG positive. Mean persistence of IgM dot EIA positivity was 2.6 months (95% confidence interval = 2.0-3.1 months) and that of IgG was 5.4 months (4.5-6.3 months). There were no significant differences between the three subgroups. Thus, positive IgM and IgG results determined by dot EIA within four and seven months, respectively, following documented or suspected enteric fever in a child from an endemic area should be interpreted with caution. In other clinical situations, the dot EIA remains a rapid and reliable aid to diagnosis.

Utilization and cost of serologic tests for Lyme disease in Maryland.

To ascertain use of serologic tests for Lyme disease (LD) in Maryland, all laboratories registered with the State Health Department were surveyed. Results show that from 1992 to 1995, 17 laboratories performed 100,000 serologic tests costing $7.1 million on Maryland residents; 90% of these tests were EIAs. The proportion of positive EIAs increased from 3.4% in 1992 to approximately 7.0% in 1994 and 1995, and the percentage of positive second tests (Western blot, WB) fell from 7.9% to 5.0%-5.5%. The large number of EIAs performed in comparison with the low incidence of LD in the state results in a low predictive value of a positive EIA test. Therefore, the WB is indicated to confirm equivocal and positive EIA tests when characteristic clinical findings of LD are not present. The 30,000 tests for LD performed annually on Maryland residents at a cost of over $2 million in direct medical costs must be added to the public health burden of LD in this state.

Detection of infection with human immunodeficiency virus type 1 before seroconversion: correlation with clinical symptoms and outcome.

Early (pre-seroconversion) infection with human immunodeficiency virus type 1 (HIV-1) was identified in 50 of 267 participants in the Multicenter AIDS Cohort Study. These 50 men had a positive EIA result, which detected IgM antibody (n = 35), p24 antigen, or serum HIV RNA (n = 15) at their last "seronegative" visit. At that visit, the mean CD4 lymphocyte number (890/mm3 vs. 1038/mm3) was significantly lower than in men who subsequently seroconverted but had no evidence of early infection. The decline in CD4 cells was slower and the duration of AIDS-free time longer in the 19 men who were symptomatic in comparison to the 31 asymptomatic men with early infection, but differences were not significant.

Limited utility of alanine aminotransferase screening of hepatitis C antibody-screened blood donors.

The introduction of hepatitis C virus (HCV) screening has significantly reduced the frequency of posttransfusion hepatitis C. To examine the current added value of alanine aminotransferase (ALT) screening, all donor screening at two large blood centers was reviewed. From July 1991 through March 1994, 1,258,000 allogeneic blood donors were screened by enzyme immunoassay for anti-HCV: 343,000 donations by the first-generation test (HCV 1.0) and 915,000 donations by the second-generation test (HCV 2.0). Donations with positive EIA results were confirmed with a recombinant immunoblot assay. Of these donors, 1,637 (0.13%) were confirmed as HCV-positive and 21,666 (1.72%) had elevated ALT. To estimate the additional margin of safety due to ALT screening, all donors who seroconverted were reviewed, and those donors who had elevated ALT but were HCV negative on a previous donation were identified. One hundred eleven HCV seroconversions were observed: 19 seroconversions from HCV 1.0-negative to HCV 1.0-confirmed-positive, 82 apparent seroconversions from HCV 1.0-negative to HCV 2.0-confirmed-positive, and 10 seroconversions from HCV 2.0-negative to HCV 2.0-confirmed-positive. The number of apparent HCV 1.0-negative to HCV 2.0-positive seroconversions was much greater than expected, which reflected the increased sensitivity of HCV 2.0. Only 15 donors were identified who had an elevated ALT on a previous HCV-negative blood donation, and all of these were among those who apparently seroconverted from HCV 1.0-negative to HCV 2.0-confirmed-positive. Out of the 10 HCV 2.0-seroconverting donors, no donor was found who was initially HCV 2.0 negative with elevated ALT and later was HCV 2.0 positive; nor were such donors found among 4 additional HCV 2.0-seroconverting donors. With the introduction of HCV 2.0 screening. ALT appears to have little value as a surrogate test for hepatitis C, and ALT testing was unable to detect any donors who later seroconverted, as detected by HCV 2.0.

Multi-center european evaluation of HIV testing on serum and saliva samples

to evaluate the reliability of HIV antibody testing on saliva. matched serum and saliva samples were collected from both seronegative (n = 344) and seropositive (n = 125) individuals in five European countries. Duplicate saliva samples collected with Omni-Sal devices provided by Saliva Diagnostic System (SDS) were pooled before analysis. all samples were analyzed by Recombinant HIV1 EIA Cambridge Bioscience and 2nd generation Abbott HIV 1&2 1A80. EIA procedures were adapted for saliva testing by modification of sample dilution and/or cut-off calculation. All saliva recording positive and/or doubtful EIA results were further analyzed by Western blot as a confirmatory method. EIA results obtained from sera analysis from both seropositives and seronegatives allowed for calculation of the tests' sensitivity (HIV1 Biotech: 99.2%-100%; Abbott: 100%) and specificity (both tests 100%). In the series of 125 saliva samples collected from seropositives, the EIA results were as follows: with Biotech (3 negative, 3 in the grey-zone and 119 reactive) and with Abbott (1 negative, 1 in the grey-zone and 123 reactive). One saliva sample found negative by both EIA tests, although fulfilling HIV1 WB criteria of positivity, was collected from an HIV2 infected person. Out of 125 saliva samples collected from seropositives, 121 produced positive Western Blot profiles, 4 were indeterminate and 1 was found negative whereas 125/125 sera were found positive. the reliability of HIV testing of saliva is dependent on the sensitivity of EIA tests and on the criteria used for the interpretation of Western blot tests as well. Although saliva testing offers numerous advantages for epidemiological purposes, it should not be recommended for diagnosis.

Three-year study of antibody to Borrelia burgdorferi in southern Spain.

The prevalence of anti-Borrelia burgdorferi antibodies was studied in Granada, Spain, between January 1991 and November 1993 in 354 patients with suspected Lyme disease (group 1); in 50 patients either with syphilis (n = 32) or without syphilis but with a positive Rapid Plasma Reagin test (n = 18) (group 2); and in 150 healthy subjects (group 3). In addition, intrathecal antibody production was evaluated by EIA in CSF samples obtained from 117 patients in group 1. Anti-Borrelia burgdorferi antibodies were detected by EIA in 58 patients (16.4%) in group 1, 29 (8.2%) of whom were positive by Western blot. Intrathecal antibody production was detected in one patient. In group 2, 8 (16%) patients had a positive EIA result, but none of these was confirmed by Western blot. western blot was negative for all subjects in group 3. The results of this study indicate that anti-Borrelia burgdorferi antibodies are not uncommon in our area, although Lyme disease is rare.

Ophthalmia neonatorum in a trachoma endemic area.

Chlamydia trachomatis can be directly transmitted by sexual or perinatal contact and indirectly transmitted by flies or fomites. Whether distinct epidemiologic forces among human populations or biologic characteristics of the organism are responsible for the different routes of transmission is uncertain. To determine if ophthalmia neonatorum and trachoma are linked epidemiologically, 38 infants with ophthalmia and 277 children with trachoma were studied for evidence of C. trachomatis infection using culture, antigen and DNA detection tests. The study was performed in a trachoma endemic area of central Kenya. Of infants with ophthalmia neonatorum, 8% to 9% had microbiologic evidence of ocular C. trachomatis infection. Of the children with trachoma, 31% had evidence of chlamydial infection. Ninety-two percent of the 59 identified strains causing trachoma belonged to the classic trachoma serovars (A, B, Ba and C). Neither of the two chlamydial strains recovered from infants with ophthalmia was a trachoma serovar. Mothers rarely (3%) had cervical C. trachomatis infection. This study does not support a major role for perinatally transmitted C. trachomatis infection in trachoma epidemiology.

Risk factors for severe disease due to Toxoplasma gondii in HIV-positive patients

The objective of this study was to assess the risk of toxoplasmosis in HIV-positive subjects as a basis for primary prophylaxis. A retrospective chart review of 400 consecutive patients was carried out and clinical and laboratory markers at first presentation and follow-up data on the occurrence of toxoplasmosis were recorded. Independent variables were identified, laboratory parameters were stratified, and estimates for the risk of toxoplasmosis and the impact of different variables on its occurrence were made using conventional statistical methods. An increased risk of toxoplasmosis was strongly associated with a positive Toxoplasma gondii IgG EIA in conjunction with a CD4+ cell cont below 0.15/nl (the estimated risk of toxoplasmosis was 20% and 35% after 12 and 24 months, respectively) or a history of one or more opportunistic infections (the estimated risk was 12% and 30% after 12 and 24 months, respectively). Toxoplasma gondii-seropositive patients with CD4+ cell counts below 0.15/nl and those with antecedent opportunistic infections are most likely to develop toxoplasmosis and thus might benefit from primary prophylaxis. The risk of disease probably outweighs the risk of medication in these subjects. Prospective clinical trials are needed to define the optimal choice of drugs.