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Evaluation of a commercial latex agglutination test for detecting antibodies to cytomegalovirus in organ donors and transplant recipients.

A commercial latex agglutination test was evaluated for use in detecting CMV antibody in organ donors and transplant recipients. When compared with indirect and competitive enzyme-linked immunosorbent assays (ELISAs) and complement fixation tests (CFT), the test gave concordant results with 96.6% of sera. The latex agglutination test was more sensitive than CFT but less sensitive than ELISA. The major advantage of the latex agglutination test was its simplicity and rapidity. Taking only 8 min to perform, it is very suitable for testing sera from organ donors, since the time available for such tests is short because of the finite ischaemic time, particularly for liver and heart transplantation.

Antibodies, Viral↗

Identification of distinct antigenic determinants on dengue-2 virus using monoclonal antibodies.

Monoclonal antibodies directed against antigenic determinants of the New Guinea C strain of dengue-2 virus were obtained from lymphocyte hybridomas produced by fusing immune mouse lymphocytes with mouse myeloma cells. Hybridoma cell culture supernatants were screened by using a radioimmunoassay employing detergent-solubilized dengue-2 infected cell antigens. Monoclonal antibodies in ascitic fluids induced by 22 selected hybridomas were characterized by the hemagglutination-inhibition, plaque reduction neutralization, immunofluorescence, and complement-fixation tests. Both type-specific and broadly cross-reactive antibodies were observed, and immunoglobulin subclasses IgG1 and IgG2a were represented in both groups. At least three distinct antigenic determinants on the virion were defined using these antibodies. A single hybridoma produced antibody which recognized a dengue-2 virus type-specific determinant and exhibited high titered neutralization but had a low titer by hemagglutination inhibition. Four preparations reacted with a type-specific determinant and exhibited hemagglutination inhibition but did not neutralize. Seventeen hybridomas produced antibodies which were broadly cross reactive in all tests. Only two preparations reacted by complement fixation with dengue-2 antigens; both were cross reactive. Immunofluorescence specificity or cross reactivity correlated with neutralization and/or hemagglutination-inhibition. The dengue-2 virus type-specific antibody useful for identification of dengue-2 infected cells by immunofluorescence has been deposited in the Hybridoma Cell Bank of the American Type Culture Collection.

Animals↗

H and M antigens of Histoplasma capsulatum: preparation of antisera and location of these antigens in yeast-phase cells.

Antiserum has been prepared in rabbits against the H and M antigens of H. capsulatum with immunoelectrophoretic precipitin arcs used as vaccines. The antiserum is specific for H. capsulatum in the immunodiffusion test and can be used as reference serum for identifying antibodies to these antigens in sera from suspected cases histoplasmosis. We found that (1) hand m antigens are not located on the surface of yeast-phase cells and (ii) complement fixation releases the antigens reactive in the complement fixation test from yeast-phase cells.

Antibodies, Fungal↗

Application of an enzyme-linked immunosorbent assay in the final stages of a bovine brucellosis eradication program.

Bovine field serums from the Australian brucellosis eradication program were used to compare 2 enzyme linked immunosorbent assays (ELISA) with the complement fixation test (CFT) and Rose Bengal test (RBT). One ELISA used an anti-bovine IgG horseradish peroxidase conjugate (ELISA 1) and the other a monoclonal anti-bovine Ig alkaline phosphatase conjugate (ELISA 2). When compared with the CFT, the ELISA 2 like the ELISA 1 lacked specificity in B. abortus vaccinated herds but the ELISA 2 was more specific than the ELISA 1 in previously infected herds and equally as specific as the ELISA 1 in nonvaccinated Brucella free herds. In this study the ELISA 2 proved more sensitive than the CFT, RBT and ELISA 1 particularly in herds where B. abortus biotype 2 was present. The value of using the ELISA 2 in conjunction with the CFT in an eradication program is discussed.

Agglutination Tests↗

[Passive hemagglutination test for individual or epidemiologic diagnosis of human brucellosis].

Coupling of an extract from B. abortus strain 99 to sheep red blood cells by chromium chloride allowed to develop a highly specific and reliable passive hemagglutination test (H). Indeed, H was positive in 99.8 per cent sera from 968 known Brucella-infected heifers, while tube agglutination revealed only 59 per cent and complement fixation 90.9 per cent positive cases. H was positive in 153 sera from 778 proven or suspected human brucellosis where agglutination and/or complement fixation tests were positive, and in 21 sera negative to conventional tests. H demonstreated remote or undiagnosed human B. abortus infections in 28.6 per cent of 18 367 human sera randomly examined in a rural area. Therefore, a simple easy-to-perform passive hemagglutination test could replace a battery of conventional tests for the diagnosis of individual brucellosis, and epidemiologic surveys. The present study points to a high prevalence of undiagnosed brucellosis in man, specially in those area where cattle are raised.

Animals↗

Mycoplasma antibody in Guillain-Barré syndrome and other neurological disorders.

Counterimmunoelectrophoresis (CIEP) was used to determine precipitating antibodies to Mycoplasma pneumoniae retrospectively in sera from 100 patients with Guillain-Barré syndrome (GBS), 125 medical and neurological controls, and 40 normal individuals. Sera from 7 patients produced precipitin lines. These positive cases included 5 patients with GBS, 1 with acute cerebellar ataxia, and 1 with acute disseminated encephalomyelitis. A complement-fixation test performed with the same antigen showed titers to M. pneumoniae of 1:512 or greater in these 7 sera. In contrast, sera from other patient controls and normal individuals were negative by CIEP and had only low mycoplasma complement-fixation antibody titers. No distinguishing clinical features separated the 5 seropositive GBS patients from the whole group except for their young age, which parallels that for human mycoplasma infection in general. Additional laboratory findings consistent with acute mycoplasma infection were demonstrated in 6 of the 7 seropositive patients.

Adolescent↗

HISTOPLASMOSIS IN MONTREAL DURING THE FALL OF 1963, WITH OBSERVATIONS ON ERYTHEMA MULTIFORME.

Although Montreal is in an endemic area, significant clinical histoplasmosis with systemic manifestations has been, until recently, infrequently diagnosed. However, since the autumn of 1963, 31 cases of clinically significant histoplasmosis have been seen by the authors. These were divided into two groups: (1) patients in whom the diagnosis was established on the basis of histological and/or cultural demonstration of the fungus; (2) patients in whom the diagnosis was based on a positive histoplasmin skin test, a complement fixation antibody titre of 1:32 or greater and compatible clinical and radiological findings. An additional group of 11 patients who presented with erythema multiforme was investigated and a heretofore unrecognized relationship between histoplasmosis and erythema multiforme was established.

Adolescent↗

Solid-phase enzyme immunoassay for determination of antibodies to cytomegalovirus.

A solid-phase enzyme immunoassay for the determination of immunoglobulin H (IgG) and IgM antibodies to cytomegalovirus is described. The enzyme immunoassay gave reliable and consistent results which were in concordance with those obtained by the complement fixation test and the indirect immunofluorescence test. Antibodies to herpes simplex and varicella-zoster viruses did not interfere in the enzyme immunoassay for cytomegalovirus IgM antibodies. In a few patients with IgM antibodies to Epstein-Barr virus, cytomegalovirus IgM antibodies were also detected. False-positive cytomegalovirus IgM antibody results were observed in sera containing both the rheumatoid factor and cytomegalovirus IgG antibodies. This rheumatoid factor interference was overcome by the absorption of sera with polymerized human gamma globulin. The absorption did not affect true cytomegalovirus IgM antibody titers. Also described is a simple enzyme immunoassay that makes possible a more sensitive detection of the rheumatoid factor than the latex agglutination test.

Antibodies, Viral↗

Preparation and standardization of an Australia antigen antibody of equine origin.

A horse has been immunized with Australia antigen (Au/SH) purified 20-fold by a procedure employing gel filtration of Cohn fraction IV derived from an Au/SH-positive human plasma pool. Hyperimmunization was initiated by the intramuscular injection of 20 ml of a mixture of equal parts of purified Au/SH and complete Freund's adjuvant. The 20-ml volume was divided into four 5-ml doses, two of which were administered on each side of the horse's neck. Booster doses of antigen alone were given as follows: 10 ml intravenously 30 days later and 5 ml intramuscularly on each of days 77 and 205. Au/SH antibody formed readily, beginning on day 17, and was demonstrated by the agar gel double-diffusion technique and the complement fixation test during the subsequent 6 months. Antihuman plasma protein antibodies were effectively removed from the horse serum by one absorption with 1 to 3 volumes of normal human plasma. Abrupt rises in anticomplementary activity observed shortly after the third and fourth antigen injections, when the horse had developed elevated and steady levels of Au/SH antibody, could possibly be due to formation of antigen-antibody complexes. After optimal conditions were determined, an Au/SH antibody reagent pool which met official requirements was prepared. It was found equally suitable for the agar gel double-diffusion, complement fixation, and counterimmunoelectrophoresis test procedures.

Animals↗

False-positive complement-fixation serology in histoplasmosis. A retrospective study.

A review of the records of 79 consecutive patients who had a positive immunodiffusion or complement-fixation test for histoplasmosis showed 28 (35%) with false-positive serologic results in relation to their clinical significance. Twelve patients (15%) had complement-fixation titers of 1:32 or greater, without cultural or histological evidence of active infection. No common clinical or laboratory finding could be correlated with the presence of false-positive tests. The latter finding correlates well with the incidence of false-positive complement-fixation titers (12%) in the normal random patient population.

Adult↗

Anaplasmosis in Uganda. I. Use of dried blood on filter papers and serum samples for serodiagnosis of anaplasmosis--a comparative study.

The suitability of blood collected on filter papers in comparison with corresponding conventional serum samples in the diagnosis of bovine anaplasmosis was studied using the complement fixation test, DOT-ELISA, Western immunoblot and rapid card agglutination test. Dried blood on Whatman filter paper no. 1 was eluted in PBS 0.05% Tween 20 giving an initial dilution of 1:10. The reactivity of the eluted samples in both DOT-ELISA and Western immunoblotting were similar to those obtained with the corresponding straight serum sample dilutions. Filter paper samples gave lower reactivity in the remaining tests when compared with corresponding serum samples. There was no significant difference in the reactivity between the eluates from filter papers stored at temperatures ranging between 15.5 and 24 degrees C and those kept refrigerated. Storage at 15.5 to 24 degrees C did not significantly affect reactivity for up to six months. Eluates from filter papers stored for six months at 15.5 to 24 degrees C continued to give similar reactivity as those from freshly prepared filter papers in both DOT-ELISA and Western blot, and in the rapid card agglutination test. It is concluded that collecting blood on filter papers is a suitable technique for large scale seroepidemiological studies on anaplasmosis and offers many advantages in developing countries where transport and cold chain facilities are a major constraint.

Agglutination Tests↗

The intravenous administration of johnin in the diagnosis of paratuberculosis in practice.

The diagnostic value of the intravenous injection of 1.6 or 3.2 ml of johnin was evaluated in a total of 116 cattle. The sensitivity was 84% with clinically suspect cattle. A general reaction occurred frequently as well as elevation of body temperature (average 1.47 degrees C). The observance of a general reaction was a good additional source of information when evaluating the test. There was no significant difference in elevation of temperature between the cattle injected with 1.6 ml and those injected with 3.2 ml. A combination of diagnostic techniques such as the complement fixation test, intravenous johnin test, microscopic examination of the faeces and a biopsy examination of the rectal mucosa were necessary, in order to confirm the diagnosis of Johne's disease. In this manner a diagnosis can be confirmed with clinically suspect cattle in 96% of the cases. The sensitivity of the intravenous johnin test was considerably lower in cases of non-clinical Johne's disease.

Animals↗

The serological clostridium tumour test--application of discriminance analysis.

The microbiological tumour test is based on the detection of humoral antibodies to clostridial rods following intravenous injection of apathogenic clostridial spores. As serological methods we used the complement fixation test with a simultaneous short-time warm and long-time cold incubation as well as the passive haemagglutination test after absorption of clostridial group-specific antibodies. The evaluation of titre dynamics was rendered difficult by normal clostridial antibodies, by serological cross-reactions and, above all, by border-line reactions. The serological test results from 189 dogs investigated were evaluated in a multivariate analytical procedure. We determined the adequate combination of methods and antigens, thus improving the reliability of the test results and objectifying the predictive value of low sero-positive titres. On the basis of the computer-calculated individual weighting factors and the corresponding reference value it has become possible to evaluate and classify actual serological data emerging from the previous overall investigation.

Animals↗

Observations on the symptomatology and diagnosis of clinical cases of Johne's disease.

Two clinical cases of Johne's disease are reported. There was a total absence of diarrhoea in one case despite a clinical course of the disease of more than 18 months. Typical gut lesions found on necropsy were confined to the small intestine. The diagnosis was confirmed on histopathological examination, and the finding of acid fast staining organisms and a retrospective complement fixation test on stored serum. The result of an intravenous avian tuberculin test conducted on the second case was inconclusive. This was attributed to the animal's temperament.

Animals↗

Isolation and identification of viral agents in Argentinian children with acute lower respiratory tract infection.

From a total population of 1,002 children with acute lower respiratory infection (ALRI), identification of virus was achieved in 304 cases. In this survey, 1,000 nasopharyngeal aspirate and 13 lung tissue samples were tested in four cell lines (HEp-2, MRC-5, MDCK, and LLC-MK2) and by indirect immunofluorescence (IIF), while 242 paired sera were studied by complement fixation. Respiratory syncytial virus (RSV) was the most frequently detected agent (n = 183), followed by adenovirus (n = 28), parainfluenza (n = 5) and 3 (n = 16) viruses, and influenza A (n = 10) and B (n = 4) viruses. The sensitivity and specificity of IIF identification vs. isolation in cell culture were high for RSV (91.5% and 94.9%, respectively). However, IIF proved poorly sensitive for detection of adenovirus (sensitivity, 23.8%; specificity, 100%). The complement-fixation test proved the least effective, with a sensitivity of only 41.5%. Therefore, on the basis of our experience, it appears that the yield for positive diagnosis of virus is increased if both isolation in multiple cell lines and identification by means of IIF are used. Our survey provides for the first time reliable data on the viral etiology of ALRI in Argentina, as determined by three different methods.

Adenoviruses, Human↗