Evaluation of three commercial enzyme-linked immunosorbent assays for diagnosis of Chagas' disease

Chagas' disease is a common cause of morbidity in Latin American countries. In Brazil, naturally occurring transmission of its etiologic agent, Trypanosoma cruzi, has been almost completely abolished through effective control programs aimed at the triatomid insect vector. Thus, transfusion of blood from infected donors has become the major route for contracting Chagas' disease due to the socioeconomically motivated migration of residents from areas where the disease is endemic to the larger urban centers. Therefore, the employment of screening tests is mandatory for all blood banks throughout the country. We compared the diagnostic performances of three commercially available screening assays used in routine testing in Brazilian blood banks: the Abbott Chagas antibody enzyme immunoassay (Abbott Laboratórios do Brasil, S?o Paulo), the BIOELISACRUZI kit (Biolab-Mérieux, Rio de Janeiro, Brazil), and the BIOZIMA Chagas kit (Polychaco S.A.I.C., Buenos Aires, Argentina). The evaluation was performed with sera obtained from chagasic patients and healthy residents of four different areas in Brazil where Chagas' disease is either endemic or emergent and where clinical manifestations of the disease and circulating parasite strains vary. The results obtained with each kit were compared to matched in-house enzyme-linked immunosorbent assay and immunofluorescence assay data obtained for each sample. Depending on the area under investigation, the three commercial kits produced specificity values between 93.3 and 100.0%, sensitivity values between 97.7 and 100%, and accuracies ranging from 93.6 to 100.0%.     

Impact of strain heterogeneity on Lyme disease serology in Europe: comparison of enzyme-linked immunosorbent assays using different species of Borrelia burgdorferi sensu lato

BACKGROUND: The gatekeeper function of the general practitioner (GP) in the pathway to specialized psychiatric services was investigated in this study, which is part of the Nordic Comparative Study on Sectorized Psychiatry. The question addressed in this paper is whether different sociodemographic and clinical factors as well as factors related to service utilization are associated with referral from the GP compared with self-referrals (including referrals from relatives). METHODS: The study comprised a total of 1413 consecutive patients, admitted during 1 year to five psychiatric centres in four Nordic countries. The centres included in this study were those that accepted non-medical referrals. Only new patients (not in contact with the service for at least 18 months) were included. RESULTS: Increasing age was the only sociodemographic factor significantly associated with referral by the GP. The clinical factors (psychosis, being totally new to psychiatry and being in need of in-patient treatment) and some treatment characteristics (planned out-patient treatment and involuntary in-patient treatment), were all significantly associated with referral by the GP. Some indication was found that self-referred patients have shorter episodes of care. CONCLUSIONS: The findings were remarkably stable across the different centres indicating a general pattern. This study extends previous work on the role of GPs in the pathway to specialized psychiatric services and indicates that the GP has an important gatekeeper function for the most disabled patients.     

Antibody responses of cows during an outbreak of neosporosis evaluated by indirect fluorescent antibody test and different enzyme-linked immunosorbent assays

Hairpin ribozymes with high cleavage activities were designed. An extra sequence was introduced at the 3'-end of the hairpin ribozyme to increase the binding to the substrate RNA, as compared to the wild-type hairpin ribozyme. A three-way junction (TWJ) was formed between the newly designed ribozyme and the substrate RNA. The complex with a solid TWJ showed less RNA cleavage activity than the wild-type hairpin ribozyme. However, the ribozyme with a TWJ with five unpaired bases or propandiol phosphate linkers had higher cleavage activity than the parent ribozyme without the TWJ. When a cis-cleavage system, in which the 5'-end of the substrate RNA was conjugated to the 3'-end of the ribozyme, was employed, the complex with the TWJ containing unpaired bases was also cleaved faster than the complex with the solid TWJ. This suggested that these differences in the cleavage activities were derived from the confirmation, and this was proven by nondenaturing gel electrophoresis. The TWJ hairpin ribozyme containing unpaired bases is able to bind strongly with substrate RNAs and to cleave them efficiently. Since the three-way ribozyme presented here is more active than the wild-type ribozyme, this type of ribozyme can serve as a more efficient tool to control RNA activities in vitro and in vivo.     

Peptide-based OspC enzyme-linked immunosorbent assay for serodiagnosis of Lyme borreliosis

Sera from 210 patients with Lyme borreliosis (LB) were studied by an enzyme-linked immunosorbent assay (ELISA) based on a synthetic peptide (pepC10) comprising the C-terminal 10-amino-acid residues of OspC of Borrelia burgdorferi. We found that 36.3 and 45.0% of the serum samples from patients with erythema migrans (EM) and neuroborreliosis (NB), respectively, displayed immunoglobulin M (IgM) anti-pepC10 reactivities, while these samples rarely (相似文献   

Advances in the diagnosis of some parasitic diseases by monoclonal antibody-based enzyme-linked immunosorbent assays

Advances in diagnostic assays for parasitic diseases include the use of monoclonal antibodies (MAbs) in antigen capture and competitive inhibition enzyme-linked immunosorbent assays (C-ELISA). Antigen capture ELISAs for Anaplasma marginale and Cryptosporidium parvum provide direct detection of these parasites during clinical disease, and the C-ELISA format has been adapted for detection of anti-Babesia equi, anti-A. marginale and anti-bluetongue virus antibodies. False-positive results may occur when antigen preparations in other ELISA formats are contaminated with Escherichia coli, erythrocyte or cell-culture antigens. The C-ELISA format overcomes problems of antigen purity, since the specificity of the C-ELISA depends solely on the MAb used. For this reason, the C-ELISA format is highly suited for use with recombinant antigens. Also, the use of recombinant protein in diagnostic assays precludes the need to infect animals for antigen production when the antigen cannot be produced in cell culture.     

Serological diagnosis of influenza B virus infection: comparison of an enzyme-linked immunosorbent assay and the hemagglutination inhibition test

The sensitivity of an enzyme-linked immunosorbent assay (ELISA) for the detection of antibody to influenza B virus was compared with that of the hemagglutination inhibition test on acute- and convalescence-phase sera obtained from adults and children infected with influenza B virus. Two whole virus, tissue culture-grown antigen preparations were used in the ELISA, influenza B/West Virginia/81 and influenza B/Hong Kong/72. Four antigens were used in the hemagglutination inhibition test. These included the tissue culture-grown whole virus antigens that were used in the ELISA. In addition the standard egg-grown antigens, influenza B/Singapore/79 and influenza B/Hong Kong/72, were included for comparison. The ELISA antibody titer was significantly correlated to the hemagglutination inhibition antibody titer, and 10 of 10 adults and 17 of 21 children infected with influenza B had fourfold antibody increases as detected by ELISA with either antigenic type of tissue culture-grown whole virus. Increases in geometric mean antibody titers of 16- to 71-fold were detected by ELISA. Increases in geometric mean antibody titers of 3- to 10-fold were detected by hemagglutination inhibition depending on the type of antigen utilized. We found that ELISA with whole virus antigens was a sensitive and specific test for the detection of antibody to influenza B virus.     

Serologic diagnosis of canine and equine borreliosis: use of recombinant antigens in enzyme-linked immunosorbent assays

A key regulatory enzyme of the C4-photosynthetic pathway is stromal pyruvate,orthophosphate dikinase (PPDK, EC 2.7.9.1). As a pivotal enzyme in the C4 pathway, it undergoes diurnal light-dark regulation of activity which is mediated by a single bifunctional regulatory protein (RP). RP specifically inactivates PPDK in the dark by an ADP-dependent phosphorylation of an active-site Thr residue (Thr-456 in maize). Conversely, RP activates inactive PPDK in the light by phosphorolytic dephosphorylation of this target Thr-P residue. We have employed a His-tagged maize recombinant C4 PPDK for directed mutagenesis of this active-site regulatory Thr. Three such mutants (T456V, T456S, T456D) were analyzed with respect to overall catalysis and regulation by exogenous maize RP. Substitution with Val and Ser at this position does not affect overall catalysis, whereas Asp abolishes enzyme activity. With respect to regulation by RP, it was found that Ser can effectively substitute for the wild-type Thr residue in that mutant enzyme is phosphorylated and inactivated by RP. The T456V mutant, however, could not be phosphorylated and was, thus, resistant to ADP-dependent inactivation by RP.     

Application of double sandwich enzyme-linked immunosorbent assay for the diagnosis of foot-and-mouth disease in Saudi Arabia

A case of Hemangiopericytoma at the thigh is reported. The Hemangiopericytoma is a rare tumour made with pericyties. This neoplasia is usually benign and it is located in the soft tissues. Tumour is profusely irrigated. The most common locations of the Hemangiopericytoma are the lower limbs and the abdominal cavity. The Hemangiopericytoma is difficult to be recognized by histological criteria. The high number of relapses and metastasis involve to an extent surgical ablation of the tumour and its borders. Present literature is reviewed and the different diagnostic and therapeutic options are discussed.     

Immunoglobulin A-specific capture enzyme-linked immunosorbent assay for diagnosis of dengue fever

Dengue fever (DF) is usually diagnosed by testing for dengue virus immunoglobulin M (IgM) by a capture enzyme-linked immunosorbent assay (ELISA) (MAC-ELISA). However, IgM can last for months, and its presence might reflect a previous infection. We have tested the use of anti-dengue virus IgA capture ELISA (AAC-ELISA) for the diagnosis of DF by comparing the results of MAC-ELISAs and AAC-ELISAs for 178 serum samples taken from patients with confirmed cases of DF. IgM appears more rapidly (mean delay of positivity, 3.8 days after the onset of DF) than IgA (4.6 days) but lasts longer; the peak IgA titer is obtained on day 8. The specificity and the positive predictive value of AAC-ELISA are 100%; its sensitivity and negative predictive value (NPV) are also 100% between days 6 and 25 after the onset of DF, but they decrease drastically when data for tests conducted with specimens from the first days of infection are included, because the IgA titers, like the IgM titers, have not yet risen. AAC-ELISA is a simple method that can be performed together with MAC-ELISA and that can help in interpreting DF serology.     

Development of enzyme-linked immunosorbent assays using recombinant borrelial antigens for serodiagnosis of Borrelia burgdorferi infection

The basic tenet of continuous quality improvement is that there is always room for additional improvement in the clinical care provided to patients. The opportunities for this improvement come from the analysis of information collected during the ongoing monitoring of important elements of care. The provision of clinical psychiatric care is seen as a complex process that is dependent on the effective functioning of all of the health and mental health care organization. The concept of continuous quality improvement is the most recent stage in a long process of defining and redefining the basic goals and tenants of medical and psychiatric quality assurance. The determination of the actual improvement of psychiatric and mental health care due to quality assurance is a substantial and important technical problem. The determination of the value of this improvement in mental health care is an even greater ethical and social problem.     

Determination of synapsin I and synaptophysin in body fluids by two-site enzyme-linked immunosorbent assays

Megalin, a 600 kDa membrane protein belonging to the IDL receptor family is highly expressed in the endocytic pathway of renal proximal tubules. In addition, this receptor is found in several other epithelia facing transcellular fluids but is also expressed in the parathyroid glands. Recent studies have established this protein as probably the most important receptor for endocytosis of macromolecules filtered in the renal glomeruli. The ligands reported to bind to megalin consist of a variety of different substances including albumin, vitamin-carrier complexes, proteinases and proteinase-inhibitor complexes, lipoprotein particles, receptor associated protein (RAP), different drugs and calcium.     

PCR enzyme-linked immunosorbent assay for diagnosis of leishmaniasis in human immunodeficiency virus-infected patients

A PCR enzyme-linked immunosorbent assay (ELISA) involving the use of bone marrow aspirates (BMA) and blood samples (BS) for the diagnosis of visceral leishmaniasis (VL) in human immunodeficiency virus-infected patients was developed with primers selected from the sequence of the small-subunit rRNA gene and compared with direct examination and in vitro cultivation. The PCR was optimized for routine diagnosis: processing of samples with lysis of erythrocytes without isolation of leukocytes, enzymatic prevention of contamination, internal control of the reaction, and ELISA testing in a microtitration plate hybridization. Of 79 samples (33 BMA and 46 BS) from 77 patients without VL, all the results were negative. Fifty-three samples (9 BMA and 44 BS) were obtained from 13 patients with VL: 6 samples drawn during anti-Leishmania treatment were negative whatever the technique used, and 47 samples (9 BMA and 38 BS) were positive with at least one technique. The sensitivities were 51% (24 of 47), 81% (38 of 47), and 98% (46 of 47) for direct examination, culture, and PCR, respectively. Thus, PCR ELISA is reliable for diagnosing VL in human immunodeficiency virus-infected patients, and blood sampling should be sufficient for the follow-up.     

Comparability and reliability of ELISA, immunofluorescence, and indirect hemagglutination assays for Trypanosoma cruzi and Trypanosoma rangeli

Serum samples and filter paper spot samples from 48 adults were assayed in three laboratories for Trypanosoma cruzi and Trypanosoma rangeli by ELISA, immunofluorescence (IF), and indirect hemagglutination (HA) in a controlled, blind experiment. IF (serum samples) attained higher levels of comparability necessary for validity than did ELISA (serum samples) in these laboratories. ELISA was inferior to IF and HA (spot samples) in testing for T. cruzi. ELISA was not reliable (intrasubject r = .51) and was not in agreement between two laboratories (kappa = .09). The experiment indicates that ELISA is not the best choice of assay for T. cruzi in epidemiologic surveys. Assays for T. rangeli by ELISA and IF lacked comparability to the extent that those two techniques could not be considered useful for T. rangeli.     

Evaluation of enzyme-linked immunosorbent assay for antinuclear antibodies

Antinuclear antibodies (ANAs) are clinically important indicators of collagen diseases. As corresponding antigens for ANAs vary considerably, patients with collagen diseases usually demonstrate several ANAs coincidentally, making difficult to detect the full spectrum of ANAs in each patient's serum. To design an efficient system for measuring ANAs, an enzyme-linked immunosorbent assay (ELISA) which adsorbs eight kinds of recombinant or purified antigens in each well of a multiwell plate was used and results were compared to those obtained with conventional assays by the fluorescent antinuclear antibodies (FANA), and double immunodiffusion (DID) methods. The positivity rates of 106 sera from patients with collagen diseases and 286 sera from healthy subjects were 92.5% and 5.5%, respectively. Sixty-one of 65 positive sera (93.8%) in the corresponding ANAs positive sera by DID or other conventional assay methods were positive by ELISA. Anti-SSA/Ro antibody could be detected with higher sensitivity by this assay method than with the FANA and DID method, but the sensitivities for anti-Scl-70 antibody and anti-centromere antibody were lower. Application of this ELISA method for measuring ANAs along with the FANA test may be beneficial for diagnosis of collagen diseases.     

Babesial antibody dynamics after cattle immunisation with live vaccines, measured with an indirect immunofluorescence test

The efficacy of vaccination of Argentinean cattle against babesiosis and anaplasmosis using live immunogens was tested to detect specific antibodies in samples obtained about 60 days after vaccination. Under these conditions a higher than expected proportion of cattle failed to show antibodies against Babesia bigemina. Therefore, a study was designed to evaluate if this failure was due to insensitivity of the routine test to detect antibodies to B. bigemina or to lack of infectivity of the live vaccine. Four groups (G) of cattle were each inoculated subcutaneously with 10 million Babesia bovis (vaccinal strain R1A), 10 million B. bigemina (vaccinal strain S1A) and 10 million Anaplasma centrale (strain M1). G1 and G2 consisted of ten Angus bulls 20-24 months old and ten Angus bulls 15-18 months old, respectively; G3 and G4 consisted of ten and 16 Holstein 1-month-old male calves, respectively. Blood samples were obtained on days 0, 10, 20, 30, 40, 50 and 60 after vaccination and the sera were analysed with an indirect immunofluorescent (IFA) test to detect antibodies to B. bovis (baseline dilution for a positive result 1:60) and B. bigemina (baseline dilution 1:120). Positive IFA titres were considered as evidence of the infectivity of the Babesia vaccinal strains contained in the vaccine. All Angus bulls were found positive to antibodies against both Babesia species, by day 20 (B. bovis) and day 30 (B. bigemina), whereas 10-25% of Holstein calves were negative throughout. The partial lack of vaccine infectivity in the calves was considered to be a consequence of innate resistance of young calves to Babesia. Antibody titres to B. bovis and B. bigemina declined by day 60 after vaccination. However, all cattle that were positive to B. bovis antibodies on day 50 were still positive to the IFA test 10 days later while 10%, 30% and 12% of cattle of G1, G2 and G3 that were positive to B. bigemina antibodies on day 50 after vaccination were found negative to the IFA test on day 60. In future, samples taken on days 40-50 will be used for detection of B. bigemina antibodies in vaccinated cattle, on day 60 for A. centrale and on either occasion for B. bovis. The reaction to the inoculation of B. bigemina S1A strain appears to lag behind the reaction to B. bovis R1A strain. It is not certain if this is a normal reaction to this B. bigemina strain or the result of interaction with the B. bovis strain.     

Improved method for serological testing in celiac disease--IgA anti-endomysium antibody test: a comparison between monkey oesophagus and human umbilical cord as substrate in indirect immunofluorescence test

To clarify the importance of the duration from the onset of a urinary abnormality until a biopsy is actually performed (UA-Bx time) in making a renal prognosis, we investigated 496 patients with IgA nephropathy (male/female: 222/274, mean age: 33.0 +/- 13.7 yrs, mean follow-up period: 10.8 +/- 4.3 yrs). All patients were found to have a urinary abnormality, including both hematuria and proteinuria, at clinical onset while demonstrating a normal renal function, and showing a serum creatinine level of < or = 1.2 mg/dl or a creatinine clearance level of > or = 80 ml/min. The UA-Bx time was divided into 3 groups: < 1 yrs (S-G), 1 < or = < 3 yrs (M-G), > or = 3 yrs (L-G). The severity of glomerular damage was divided into 5 groups based on the occupational rate of segmental sclerotic glomeruli. Based on a multivariate analysis of independent prognostic factors relating to renal death, the severity of glomerular damage was the most independent factor, while the UA-Bx time showed no risk for renal death. However, based on a multivariate analysis of the UA-Bx time regarding the timing of a renal biopsy, patients in L-G, which had the most glomerular damage, showed twice the hazard ratio as those in S-G or M-G and the difference was significant. These results thus indicate that because the glomerular damage is able to progress for 3 yrs or longer after the clinical onset of renal disease, a renal biopsy should therefore be performed within 3 yrs from the clinical onset in patients demonstrating both hematuria and proteinuria when such patients are also suspected of having IgA nephropathy.