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Persistent levels of serum antibodies to bovine C-type virus were demonstrated by the complement fixation test in cattle of a leukosis herd during an observation period of one and one half years. Using the same method, no antibodies were detected in a control herd.
A specific diagnostic complement-fixation test for hepatitis A antibody in human serum was described employing livers of marmosets infected with CR326 strain human hepatitis A virus. Persons with hepatitis A, but not hepatitis B, developed hepatitis A CF antibody shortly after the onset of illness and this persisted thereafter. Good agreement was noted in the development of CF and neutralizing antibodies in hepatitis A cases. Hepatitis A was shown to occur in a person with hepatitis B antigenemia and hepatitis B occurred in persons with hepatitis A antibody. Most persons with hepatitis A who were tested, but none of those with hepatitis B, developed increased anticomplementary activity in their sera at the time of onset of illness. At least one patient with hepatitis A developed antibody against normal liver that persisted. The possible inplications of this in relation to pathogenesis and to non-specific diagnostic tests in hepatitis were discussed. A limited epidemiologic study of a family outbreak of hepatitis in Costa Rica and of a group of young adults in our epidemic country acquire their infections at an early age and are immune thereafter; persons in areas of relatively low incidence may proceed into adulthood without experience with hepatitis A. The CF test should provide an excellent tool for diagnosis and for epidemiologic investigation of hepatitis A and should be of considerable value to detect hepatitis A virus in attempts to propagate the virus in cell culture.
The value of the complement fixation (CF) test as a means of assessing the immune response of fowls to Newcastle disease virus was examined using several strains of virus. A close degree of correlation was found between the CF test and the simpler haemagglutination-inhibition (HI) test. Like the HI test, the CF test only gave an indication of the immune status and was not a direct measurement of immunity.
A complement fixation (CF) test for neonatal calf diarrhea has been developed with Nebraska calf diarrhea virus (NCDV) as antigen. The CF antigenicity of NCDV appeared in BK cell cultures for the first time 3 days after inoculation. Usually, the unconcentrated BK cell culture fluid infected with NCDV possessed poor CF antigenicity. The fluid concentrated by ultracentrifugation and ultrafiltration and with polyethylene glycol 6000 revealed a high-titered CF antigenicity to antiserum against NCDV. The antigen was not sensitive to lipid solvents, but was relatively resistant to heating. It was recognized that the concentrated fluid of cell culture infected with NCDV was available as antigen in the CF test for the serological demonstration of infection with a calf reovirus-like agent. The wide prevalence of diarrhea among Japanese calves with the calf reovirus-like agent was revealed from the results of CF and neutralization tests of NCDV.
A slide flocculation test for Chagas' disease is described, which uses a lyophilized, stable antigen obtained by formalin and ultrasonic treatment of culture forms of Trypanosoma cruzi. The test was compared with other tests for the serodiagnosis of American trypanosomiasis and showed a high sensitivity, positive results being obtained in every case of acute or chronic Chagas' disease. In sera from blood donors and from normal individuals with negative T. cruzi antigen complement fixation tests a specificity of 96% was found. False positive flocculation tests were seen, especially in cases of South American blastomycosis and in a few cases of acute toxoplasmosis. Since it is easy and quick to perform, the slide flocculation test can be recommended as a screening procedure, especially for blood banks.
Serums from 263 cattle suspected of having paratuberculosis on the basis of clinical signs, were tested for antibodies to Mycobacterium paratuberculosis with a complement fixation test (CFT) employing a heat extracted, soluble M. avium antigen. Microscopic examination confirmed that 172 (65.4%) clinically affected animals had paratuberculosis, the remainder being disease-free. The specificity and sensitivity of the CFT was 92.3% and 74.4% respectively. Phenol treatment of serums before testing was compared with no treatment and was found to have no significant effect on the CFT titres. Results obtained are discussed in relation to the cause of false negative and false positive reactions.
The technical procedures for a simple quantitative micro-complement (C) fixation test are described. Major advantages of the present technique compared with the previously described method are: a) a simple measurement of the residual hemolytic activity of C by counting the radioactivity released from 51Cr-labeled sensitized sheep erythrocytes (51Cr-EA); b) an increased sensitivity of the test, brought about by the use of a relatively small number of 51CR-EA per reaction volume; and c) an increased specificity of the test, achieved by maintaining a constant amount of C available for the specific antigen--antibody reaction.
The complement fixation test was applied to assess antibody responses in cattle infected with T. brucei, T. congolense, and T. vivax which had been treated and re-infected. After the first infection, the specificity of the results was low in T. congolense, fair in T. brucei, and high in T. vivax infected animals. After re-infection with the same trypanosoma species as used for previous infections, the specificity of CF results was high in all cases. After a foregoing T. vivax infection, specific titres of T. congolense and to a lesser degree of T. brucei infections were frequently masked by residual T. vivax titres. After treatment of trypanosoma-infected cattle the fasted drop of CF antibody titres occurred in T. congolense cases, followed by T. brucei infections. T. vivax titres persisted over prolonged periods.
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Studied were five flocks of 240 sheep each. It was found that prior to impregnation the positive complement-fixation test (c. f. test) reagents varied from 16 to 61 per cent with titers ranging from 1:50 to 1:10. Abortions, however, were established only in two of the flocks consisting of young sheep, aged 3 and 4 years, at the first or second lambing. The abortions took place in the fourth month of pregnancy. In one of these flocks having 43 per cent positive reagents there were 40 per cent returns, and 28 of the ewes miscarried, 12 of them being positive for toxoplasmosis. In the second flock there were 61 per cent positive reagents, and 9 of the ewes miscarried, 5 of them being positive for toxoplasmosis. In the remaining three flocks consisting of 5-year-old sheep there were no abortions. The ewes that miscarried and were positive for toxoplasmosis showed a rise in their serum titers: from 1:40 on the 15th day after the abortion to 1:80 on the 30th day. It is believed that in these sheep the abortions are not due to toxoplasmosis, and in sheep flocks with a high percent of positive toxoplasmosis reagnets there may not be abortions.
Anticomplementarity occurring in the complement-fixation (CF) test with soluble antigens of rickettsiae of the spotted fever (SF) group is caused by interaction of soluble antigens with guinea pig sera serving as the source of complement. The serum factor responsible for anticomplementarity is thermostable and cannot be removed by treatment of guinea pig serum with CO2 or nitrogen. An adverse effect of such serum was also observed upon its mixing with other guinea pig sera lacking anticomplementarity properties. An anticomplementarity factor present in the soluble rickettsial antigens was sensitive to treatment with potassium periodate.
A stabilized modification of the single radial complement fixation (SRCF) test in gel was developed for detecting various virus antibodies. The principle of the test is based on the use of a single-stage procedure with an agarose plate containing virus antigens and antibody-coated erythrocytes, and thin plastic film coated with complement. By filling the wells in the agar plate with a 1:4 diluted heat-inactivated sera and covering the agar surface with a complement film, a zone of unlysed cells surrounded by a hemolytic area appears after incubation overnight at 4 degrees C and then for 1-2 h at 37 degrees C, depending on the antibody titers. The SRCF antibody titer is calculated numerically from the square of the diameter of the unlysed cell zone. The stability of reagents could be significantly improved using thin complement film and several stabilizers. When this test was used for serodiagnosis of influenza, respiratory syncytial (RS), mumps, adeno virus type 3 and herpes simplex type 1 virus infections (using a total of 400 sera), excellent correlations were demonstrated for antibody titers between conventional complement fixation (CF) and SRCF titers. Furthermore, the expression of antibody titer as an SRCF unit with consecutive value, produced results sensitive to fluctuations in the antibody titers. The simplicity of the procedure, stability of the reagents, and excellent correlation with the conventional CF test might make this a useful test for routine serodiagnosis and seroepidemiological survey of various virus infections.
An enzyme-linked immunosorbent assay (ELISA) was evaluated and compared in parallel with the standard complement fixation test (CFT) for the diagnosis of bovine subclinical paratuberculosis. Bovine sera preabsorbed with the mixture of Mycobacterium phlei and kaolin suspension were assayed for antibody activities to the crude protoplasmic antigen of Mycobacterium paratuberculosis in the ELISA. ELISA antibody titer was expressed as ELISA antibody index (EAI) value: EAI = (At-An)/(Ap-An), where At, Ap and An are the absorbance values of a 1:200 dilution of unknown test sera, a 1:400 dilution of positive control serum, and a 1:200 dilution of negative control serum. An EAI of 0.6 or greater was established as a reasonable cutoff point for a positive antibody titer by ELISA. Of the 156 sera from cattle with subclinical M. paratuberculosis-infection, 106 (67.9%) were positive by ELISA and 41 (26.3%) by CFT. Of the 3,880 sera from cattle in the herds which had no history or evidence of paratuberculosis, 3,875 (99.9%) were negative by ELISA, and 3,787 (97.6%) by CFT. Positive ELISA titers were detectable 1 to 5 months earlier than positive CFT titers in experimentally infected cattle, and 7 to 10 months earlier in naturally infected cattle. These results indicate that the ELISA should replace the CFT as the routine test of choice for the diagnosis of bovine paratuberculosis.
An enzyme-linked immunosorbent assay (ELISA) and a complement fixation test (CFT) were applied to sera collected regularly from 60 cattle challenged with Brucella abortus strain 544. Of the 60 cattle, 48 were vaccinated with either B. abortus strain 19 or B. abortus strain 45/20 as calves or adults. The remaining 12 cattle were not vaccinated. Of the 27 sera from cattle found to be infected at slaughter, 9 showed aberrant reactions to the CFT in that a positive titer after challenge was delayed or transient. The performance of the ELISA in these nine cattle with aberrant reactions and the other cattle in the trial was as follows. After vaccination with strain 19 or strain 45/20, the number of weeks at which the ELISA was positive was significantly greater (P less than 0.001) than that for the CFT. The strain 19 vaccine also induced positive responses to the CFT and the ELISA for a significantly longer period (P less than 0.001) than did the strain 45/20 vaccine. For cattle with aberrant reactions, the number of weeks after challenge when the ELISA was positive was significantly greater (P less than 0.001) than that for the CFT. For noninfected cattle, the average number of weeks after challenge when the CFT was negative (35 of 40) was higher than that for the ELISA (18 of 40).
A panel of 68 serum specimens from 41 subjects exhibiting various immunological patterns to Mycoplasma pneumoniae as determined by detection of a 180 kDa protein in immunoblotting was used to compare five commercially available tests based on different methods: complement fixation test (CFT), microparticle agglutination (MAG), indirect immunofluorescence assay (IFA), enzyme-linked immunosorbent assay (Elisa), and latex agglutination (LA). The tests were performed according to the manufacturers' instructions. For the determination of immunity to M pneumoniae, the five tests were in good accordance with immunoblotting: sensitivity was 100% for all the five assays, specificity ranged from 95.6% (MAG) to 82.6% (Elisa) and overall agreement ranged from 98.2% (MAG) to 92.8% (Elisa). The comparisons of antibody rates obtained by the four quantitative tests (CFT, MAG, IFA, Elisa) showed correlation coefficients ranging from 0.87 (CFT-IFA) to 0.67 (CFT-Elisa). Six significant antibody rises demonstrated by immunoblotting patterns were detected by all the tests but Elisa in one case. As a whole, the commercial assays gave satisfactory results for routine determination of immune status to M pneumoniae: CFT was the cheapest test and MAG and LA were the easiest to perform.
The authors compared the sensitiveness of the complement-fixation reaction and the latex-fixation test in the demonstration of a migration phase of ascariasis in a non-specific host (rabbit). The sensitiveness of the latex-fixation test was higher by the value of one dilution of the geometric series, on an average.
A competitive enzyme-linked immunosorbent assay (ELISA) using a monoclonal antibody conjugated to horseradish peroxidase MA(A) and a complement fixation test (CFT) were applied to sera collected over a two-year period from 60 cattle challenged with Brucella abortus strain 544. Forty-eight of the cattle were previously vaccinated with B. abortus strain 19 (S19) or B. abortus strain 45/20 (45/20). After challenge 33 of the cattle remained uninfected and nine of the 27 infected cattle showed aberrant reactions by the CFT. The performance of the MA(A) ELISA was as follows: after vaccination, the MA(A) ELISA, like the CFT, was unable to differentiate infected cattle from those recently vaccinated with S19. After challenge the MA(A) ELISA gave results comparable with the CFT for those cattle with aberrant reactions. For the non-infected cattle there was a similar number of weeks after challenge when both tests were negative. It is suggested that the main advantage of the MA(A) ELISA when compared with the CFT lies in its relatively simple test procedure.
Studies were carried out to establish the optimal conditions for the indirect complement-fixation test (ICFT) and explore the possibility to use the test in differentiating the foot-and-mouth disease (FMD) viruses and checking the immunity obtained in survivals or in animals that had been vaccinated against FMD. It was demonstrated that the proper use of ICFT necessitates in turn the use of a minimal amount of antigen, which in the presence of 1-2 E homologous hyperimmune serum completely binds the 1 E complement. Results showed that the sera of survived or vaccinated animals contain specific antibodies of titers of 1:4 up to 1:256. The latter depended on the type of vaccine, the amount of antigen in the vaccinal dose, and the time period from the immunization or survival up to the time of taking the serum samples. It is pointed out that ICFT is a sensitive and specific method for the differentiation of FMD viruses and the study of animals' immunity against FMD.