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Bovine leukocyte cultures achieved good growth after 72 hours of cultivation. Foot-and mouth disease (FMD) virus type 0 multiplied in such cultures to a titre of 10(6) TCID50/0.1 ml. The cell culture-grown virus was found to be suitable for the complement fixation test (CFT) after purification and concentration with calcium phosphate.
Traditionally, plasma for the production of the human varicella-zoster immunoglobulin (VZIG) has been selected on the basis of the complement-fixing antibody (CFA) titre. Since immune individuals may lack CFA to varicella-zoster virus (VZV), non-CFA may be of importance in protection. In a search for a simple and reliable method for potency determination, 24 VZIG preparations were quantified by enzyme-linked immunosorbent assay (ELISA), the complement-fixation test (CFT), the indirect fluorescent antibody test to acetone-fixed (IF) and viable (FAMA) VZV-infected cells, respectively. The antibody titres obtained by the various methods were compared. Arranged in order of decreasing agreement, the correlation coefficients (r) of the regression equations between the variables were 0.62 for CFT and FAMA, 0.50 for CFT and ELISA and 0.26 for CFT and IF in a log2 plot. There was complete agreement between the titres obtained by the commercially available Enzygnost Varicella/Zoster kits (Behring Institute, Marburg, F.R. Germany) and the ELISA microtitre plates produced at our institute (r = 1). The regression equation lines for ELISA/CFT and FAMA/CFT titres tended to be parallel to each other, while the line for IF/CFT titres had a less steep slope. Similar titration curves were obtained for VZIGs fractionated by two different methods. Furthermore, the titration curves of serum pools from varicella and zoster convalescents, respectively, had a similar shape below delta OD = 0.4. Generally, a steeper slope was observed above delta OD = 0.4. As antibody detectable by ELISA seems to correlate with protection and the method is sensitive, specific, reproduceable, simple to carry out and easily automated, it may be suitable for the potency determination of VZIGs.
The association of Campylobacter pyloridis with antrum gastritis and peptic ulcers was described. We investigated antral biopsies from 180 patients who underwent gastroscopy. By culture or Gram stain or both, we found overall 98 (54%) of them to be positive for C. pyloridis. In the various groups the following percentages were found to be positive: normal antral mucosa 3% (n = 30); moderate superficial antrum gastritis, 49% (n = 83); severe superficial antrum gastritis, 86% (n = 44); duodenal ulcer, 83% (n = 54); and gastric ulcer, 72% (n = 18). A serological screening that used a complement fixation test yielded the following results: highest rates of positive complement fixation titers were seen in patients with severe gastritis and those with duodenal ulcers, both with 79%; the lowest incidence was in a group of 20 blood donors, with 5%. Positive complement fixation titers in gastritis patients also correlated well with characteristic patterns on immunoglobulin G and A immunoblots, while there was no specific reactivity observed on immunoglobulin M immunoblots.
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A commercially available absorbed ELISA for the diagnosis of Johne's disease (JD) (paratuberculosis) in cattle, the Johne's Absorbed EIA, was compared with the conventional complement-fixation test (CFT) used in Australia. Stored plasma from 3 Victorian dairy herds with a history of JD, sera from specimens submitted from animals showing clinical signs of JD and sera from the US National Repository for Paratuberculosis Specimens were used to determine the sensitivity of each test. The EIA detected 48.8% of 43 Australian animals with subclinical JD, while the CFT detected only 12 (21.4%) of 56 subclinically affected cattle. Of 150 subclinically infected US cattle, the EIA detected 47.3% and the CFT detected 52.0%. The EIA detected 59.7% of animals which at the time of sampling were shedding Mycobacterium paratuberculosis in their faeces, but showed no clinical signs of JD, while the CFT detected 57.3%. The EIA correctly identified 88.2% of 136 histologically confirmed clinical cases, and the CFT detected 83.4%. The specificity of each test was determined by testing sera collected at slaughter from animals residing in a known JD-free area of Australia, and from samples from the US National Repository of Paratuberculosis Specimens collected from certified-free herds in Wisconsin. The EIA was found to have a specificity of 99.8% when 998 Australian animals were used as the test population, and 99.0% when 196 US animals were used. The specificity of the CFT using Australian samples was 96.9% and 95.2% using American samples.
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The results from studies to measure lytic complement (C') in sera of different animal species were reviewed. The traditional system, using sheep red blood cells (RBC) and rabbit antibody, was confirmed as the most sensitive to measure C' levels in man, monkey, dog, guinea pig, and rat serums. Sera C' from horse, cow, and sheep were found to be best assayed using rabbit RBC, whereas C' from goat, cat, and rabbit were best assayed with human RBC. Antibodies and C' from the same species usually mediated lysis of foreign RBC, but this lysis occurred more readily with some RBC targets than with others and may be associated with the presence of natural antibodies in the test sera. The effects of the species origin of a C' source in immunologic reactions in vitro and in vivo are discussed.
A detailed investigation into the effect of modifying the incubation temperature of the complement-fixation (CF) test is described. For varicella-zoster virus cytomegalovirus and rubella virus, increasing the incubation temperature progressively increased the sensitivity of the CF test to reach a maximum at 15 degrees C, at which temperature the geometric mean titre of seropositive samples was significantly greater than that found at 4 degrees C. For these three viruses, each serum shown to contain IgG antibodies by ultrasensitive radioimmunoassay procedures was detected by CF following incubation at 15 degrees C. No false-positive reactions occurred at 15 degrees C, but it was our impression that anticomplementary activity was enhanced at this temperature. Significant increases in antibody titre at 15 degrees C were also seen when measles virus, respiratory syncytial virus, adenovirus and Mycoplasma pneumoniae were employed as CF antigens. The results demonstrate that the CF test should be performed at 15 degrees C if optimum sensitivity is to be achieved. The ability of the test to detect significant rises in antibody titre was not impaired at the higher incubation temperature.
A method is described for preparation of an antigen from E. ovis-infected blood first lysed by freezing and thawing, and then followed by sonic disruption of the centrifuged deposit. It was then used in a complement fixation test to demonstrate antibodies in sheep infected with E. ovis. The antigen was not anti-complementary and it retained its potency for long periods when stored concentrated, at about -20 degrees C. Repeated freezing and thawing also has had no effect on the potency of the concentrated antigen. The testing of 668 serums from commercial flocks where E. ovis infection has been diagnosed microscopically has shown the CFT to be a valuable diagnostic aid mainly on a flock sample basis, though the test could be of value on an individual sample basis if the sample were taken from a recently infected sheep.
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