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Topic:Complement Fixation
Complement fixation is an immunological reaction involving the binding of complement proteins to antibodies that have attached to specific antigens. In horses, this process is part of the innate immune response, contributing to the identification and elimination of pathogens. The complement system consists of a series of proteins that, when activated, enhance the ability of antibodies and phagocytic cells to clear microbes and damaged cells. This system also promotes inflammation and attacks the pathogen's cell membrane. The complement fixation test is a diagnostic tool used to detect the presence of specific antigens or antibodies in equine serum by observing the consumption of complement proteins. This page compiles peer-reviewed research studies and scholarly articles that explore the mechanisms, regulation, and diagnostic applications of complement fixation in equine immunology.
Preliminary findings for an inactivated African horsesickness vaccine using binary ethyleneimine. Investigation studies on inactivated African horsesickness vaccine using binary ethyleneimine were conducted. The inactivation process of virulent type-9 strain using the above inactivant revealed complete virus inactivation at 18, 48 and 84 h post-treatment with inactivant concentrations of 0.004, 0.003 and 0.002M, respectively, without detection of residual virus. An inactivant concentration of 0.003M is recommended and no changes in viral antigenic properties were noticed in complement fixation test. The physical parameters in oil-emulsion vaccine using the incomplete Freund's adjuvant, wer...
Inhibition of equine complement activity by polysulfated glycosaminoglycans. The ability of polysulfated glycosaminoglycans (PSGAG) to inhibit the complement cascade was evaluated. The role of complement in inflammation and infection has been well documented. Inhibition of the complement cascade by PSGAG could explain why intra-articularly administered PSGAG diminish diarthrodial joint inflammation and potentiate septic arthritis in horses. Hemolytic complement testing was performed to evaluate the effect of PSGAG on the equine classical and alternate pathways of complement, using rabbit erythrocytes as the target cells. Concentration of PSGAG between 0.2 mg/ml and 0.6...
Prevalence of complement-fixing antibody to the African horse sickness virus in domestic animals in Nigeria. The occurrence of antibodies against the African horse sickness virus was investigated in 246 domestic animals (horses, donkeys, camels, dogs) in various regions of Nigeria by means of the complement-fixing rate. 34% of the sera tested were positive: 75% in donkeys, 68% in horses, 19% in camels, and 9% in dogs. Among the horses, those of 6 to 15 years of age had higher than average prevalence rates than the other age groups. Stallions from the northern regions had higher prevalence rates than mares generally and stallions from other regions. These findings are important for the epidemiology of...
[Chlamydia as the cause of abortions in horses]. Between 1982 and 1989 59 equine fetuses were investigated for chlamydiae using animal experiments and embryonated eggs. Chlamydiae were isolated from 16 fetuses (27.1%) originating from 8 studs. The macroscopical lesions of the fetal organs were characterized by extensive haemorrhages. The histological picture shows severe lesions of the blood vessels of different organs. In 6 studs in which chlamydiae had been isolated, blood sera of clinically healthy and pregnant mares were investigated for antibodies during 1989 and 1990. Antibody titres between 1:10++ and 1:40 were detected by using compl...
Antibodies in horses, mules and donkeys following monovalent vaccination against African horse sickness. A total of 256 sera collected from three species of domesticated equidae in four different Spanish provinces were examined 1-4 months after the administration of attenuated monovalent African horse sickness virus (AHSV) serotype 4 vaccine. Approximately 10% of the sera were negative by ELISA, virus neutralization, agar gel immuno-diffusion and complement fixation tests. Similar negative reactions were recorded with sera from two ponies after experimental primary vaccination. The rapid rise in antibodies in sera from these two ponies, after a second dose of vaccine, suggested they would probabl...
Development of an avidin-biotin dot enzyme-linked immunosorbent assay and its comparison with other serological tests for diagnosis of glanders in equines. A dot enzyme-linked immunosorbent assay (dot ELISA) was developed for diagnosis of glanders in equines. The test was based on the detection of IgG antibodies to Pseudomonas mallei antigens bound to nitrocellulose coated on plastic strips (dipsticks), the reaction being amplified by an avidin-biotin system with biotinylated anti-horse IgG and horseradish peroxidase-avidin D. Sera from 810 normal, six naturally infected and 48 sensitized equines were tested by this assay, and results were compared with complement fixation, indirect haemagglutination and counter-immunoelectrophoresis tests. Dot E...
One way protection between equid herpesvirus 1 and 4 in vivo. Two groups each of six sibling ponies were exposed to sequential infections with equid herpesvirus 1 or 4 (EHV-1 or EHV-4) at four or five month intervals. Two exposures to EHV-4 did not significantly reduce virus shedding or pyrexia when the ponies were subsequently exposed to EHV-1. However, two sequential infections with EHV-1 completely protected against challenge with EHV-4. Virus neutralising antibody in each group did not increase until 21 days after primary exposure and was subtype specific. However, complement fixing antibody rose within seven days after inoculation with EHV-1, and 14...
Detection of African horsesickness (AHS) in recently vaccinated horses with inactivated vaccine in Qatar. Two 7-year old Arabian racing horses were reported to show typical AHS symptoms in Qatar and died shortly after. The horses had been vaccinated with formol inactivated vaccine approximately 10 days before the onset of the disease. Blood samples from these horses were collected and AHS virus isolated from one sample after intracerebral (i.c.) inoculation into suckling mice. The virus identity was confirmed by complement fixation test (CFT) using the virus antigen and reference type 9 of AHS virus hyperimmune serum. The serotype of the isolated virus was identified by serum neutralization test (...
Population data and a fourth allele for equine complement component 3 (C3). The C3 polymorphism of equine serum or plasma revealed by agarose gel electrophoresis can be diagnosed with protein stain following acid protein fixation. In addition to the three alleles previously described (C31, C32, C33), a fourth allele (C34) was found. Population data for 25 domestic breeds and Equus przewalskii are presented.
Laboratory diagnosis of African horse sickness: comparison of serological techniques and evaluation of storage methods of samples for virus isolation. Five serological methods of diagnosing African horse sickness were evaluated, using a battery of serum samples from experimental horses vaccinated and challenged with each serotype of African horse sickness virus (AHSV1 through AHSV9): agar gel immunodiffusion (AGID), indirect fluorescent antibody (IFA), complement fixation (CF), virus neutralization (VN), and enzyme-linked immunosorbent assay (ELISA). The 5 tests were also compared using a panel of field samples, convalescent equine sera with antibodies to domestic equine viral diseases, and sera from horses awaiting export. The ELISA describ...
Complement activity and selected hematologic variables in newborn foals fed bovine colostrum. Serum complement activity and selected hematologic variables were evaluated in 5 newborn foals fed bovine colostrum (principal group) and 6 foals allowed to nurse their dam (control group). Also, bovine colostrum was evaluated for anti-equine antibodies. Precolostral serum hemolytic and conglutinating complement activities were low and increased similarly in foals of both groups to reach adult values between 1 and 3 weeks after birth. Bovine colostrum strongly agglutinated, but did not hemolyse principal foals' RBC and blood containing all known equine blood group alloantigens. Hemolysis was n...
Complement fixing antibodies against arboviruses in horses at Lagos, Nigeria. Sixty-two sera horse collected from two stables at Lagos, Nigeria, were tested for complement fixing antibody to 8 arbovirus antigens; Chikungunya, Igbo-Ora, Yellow fever, Wesselsbron, West Nile, Potiskum, Uganda S and Rift Valley fever. Ten per cent of the horse sera examined contained CF antibody to one or more of the test antigens and indicated considerable arbovirus activity in the two stables. Reactions with flavivirus antigens were most common and the highest antibody titres were obtained with Wesselsbron and Yellow fever viruses. Eleven per cent of the sera tested reacted with alphaviru...
Opsonins in uterine washings influencing in vitro activity of equine neutrophils. Uterine washings were found to promote neutrophil mediated killing of Streptococcus zooepidemicus. Depletion of complement and/or specific antibody from the washings significantly reduced bactericidal activity. Phagocytosis of yeast by uterine washings was complement dependent. Inhibition of the classical pathway significantly reduced opsonic activity indicating that, in addition to direct activation via the alternate pathway, antibody may also be involved in yeast phagocytosis.
Chemotactic response of equine polymorphonuclear leucocytes to Streptococcus equi. Streptococcus equi infection in horses is characterised by intense infiltration of lymph nodes by polymorphonuclear leucocytes (PMNs) suggesting a potent chemotactic response to the organism or its products. Equine PMNs were separated using Ficoll-Hypaque medium and used in an assay of chemotaxis under agarose to study the components of S equi involved in this response. Results showed that complement-derived chemotactic factors generated by activation of the alternative complement pathway were important in chemotactic responses to S equi. Both whole bacteria and peptidoglycan preparations were...
Serologic response of Babesia equi-infected horses as measured by complement-fixation and indirect fluorescent antibody tests. Both the complement-fixation test (CFT) and the indirect fluorescent antibody test (IFAT) were conducted on weekly serum samples from nine Arab geldings for 28 days before and 256 days after their exposure to Babesia equi of European origin. On an average the IFAT became positive 8 days before the CFT and showed higher relative serum titer increases. Both test procedures successfully detected infection and neither showed an appreciable drop in titer during this time frame, with the exception of the CFT, which showed a transient drop immediately following treatment with imidocarb. A test conduc...
An investigation into the clinical pathological changes and serological response in horses experimentally infected with Babesia equi and Babesia caballi. Serologically negative horses, as determined with the indirect fluorescent antibody test (IFA), were infected with Babesia equi and 60 days later with Babesia caballi. The only clinical signs of disease observed in these animals were a febrile reaction and slight icterus. Haematological changes included a drop in haematocrit and haemoglobin concentration, as well as lowered platelet counts. The serum concentrations of albumin, iron and phosphorus were lowered. Mildly elevated serum bilirubin and fibrinogen concentrations were observed. Antibody titres were determined with the IFA and complemen...
The use of a passive hemolysis system to evaluate the complement activities of six mammalian species. A passive hemolysis assay system was developed which permitted comparisons of the hemolytic activities of complement (C) from six species. This system employs a single antigen and an antiserum raised in one species. Thus, variations resulting from different target antigens and those inherent in using antibodies (of different affinities and isotypes) raised in a variety of species were minimized. Of the erythrocytes (E) examined, those from horses and guinea pigs were most susceptible to lysis, and either would be suitable, as a tentative choice, for measuring C activity of a previously unstudi...
Genetic restriction of cytolysis during equid herpesvirus 1 subtype 2 infection. Six Welsh Mountain pony foals were experimentally infected with a subtype 2 isolate of Equid Herpesvirus 1 (EHV-1) and subsequently examined for T cell mediated cytotoxicity against both subtypes. Cytotoxicity was not observed at 3 or 7 days after primary exposure but virus-specific, and genetically restricted, cytotoxicity of EHV-1-labelled autologous skin fibroblasts could be demonstrated 7 and 21 days after the animals were given a second exposure to live virus. Killing of subtype 2 antigen-labelled targets was more efficient than subtype 1 coated cells. This finding was paralleled by the o...
Cellular and humoral defence mechanisms in mares susceptible and resistant to persistent endometritis. Both random and directional migration of blood neutrophils from 9 mares susceptible to persistent endometritis were significantly less (p less than 0.05) than neutrophils from 8 resistant mares. Serum from susceptible mares had significantly more (p less than 0.01) chemotactic activity than serum from resistant mares. Although phagocytosis of yeast blastospores by blood neutrophils from 4 resistant and 3 susceptible mares was similar, uterine neutrophils from susceptible mares were significantly worse (p less than 0.01) at phagocytosis than uterine neutrophils from resistant mares. Uterine was...
The immunological response of foals to Rhodococcus equi: a review. Normal horses of all ages regularly show evidence of having responded immunologically to R. equi, thus adding serological support to epidemiological evidence that this organism is a normal intestinal inhabitant. More animals from "diseased" farms show a stronger antibody response when compared with foals from "healthy" farms. Various serological tests have been used to detect evidence of infection and to relate antibody level to severity of disease. Anti-R. equi IgG antibody levels, as measured by ELISA, are raised significantly during natural infection. Clinical severity of pneumonia can be c...
Vesicular exanthema of swine virus: isolation and serotyping of field samples. Virus isolation was attempted from 262 field samples of vesicular material collected during the outbreaks of vesicular exanthema of swine in the U.S.A. from 1952-54. Using primary swine kidney culture, viral cytopathogenic agents were isolated from 76.3% of the samples. However, an overall recovery rate of 82.1% was obtained after samples negative in tissue culture were inoculated intradermally in susceptible swine. All vesicular exanthema of swine virus isolates were identified as serotype B51 using complement fixation and serum neutralization tests. Two isolates did not react with antisera t...
Complement-induced equine neutrophil adhesiveness and aggregation. Equine neutrophils (PMN) were isolated from citrated normal blood by density gradient separation on Ficoll-Hypaque to greater than 96% purity and 98% viability and an average of 3.78 x 10(7) PMN/ml. The agonist C5a des Arg was used in serial dilutions of whole zymosan-activated equine plasma (ZAP) or was partially purified from ZAP by column chromatography. Purified equine PMN exhibited rapid aggregation following incubation with C5a des Arg which was further dependent on the availability of divalent cations, especially Mg++. The microfilament disruptive agent cytochalasin B (5 micrograms/50 m...
Neutrophil phagocytic and serum opsonic response of the foal to Corynebacterium equi. This study was undertaken to examine the neutrophil response to Corynebacterium (Rhodococcus) equi, and to assess the possibility of neutrophil immaturity or malfunction in predisposition to C. equi pneumonia in foals. Neutrophil phagocytosis of Corynebacterium (Rhodococcus) equi was studied in foals from birth to 6 months of age. Chemiluminescence (CL) and bactericidal assays were used to assay the phagocytic response of peripheral blood neutrophils to C. equi in vitro. Results of in vitro bactericidal and CL assays indicate that foal neutrophils are able to ingest and kill C. equi, however a...
Complement-mediated hemolysis of horse erythrocytes treated with equine infectious anemia virus. Horse erythrocytes treated with equine infectious anemia virus hemagglutinin were found to be lysed after incubation with fresh horse serum at 37 degrees C. Fresh guinea pig serum induced more efficient hemolysis than horse serum. Direct immunofluorescence test revealed the adsorption of complement factors on the surface of the erythrocytes. Calcium and magnesium ions were necessary for the hemolysis to take place. Antibody against equine infectious anemia virus enhanced the virus-induced complement-mediated hemolysis. These observations indicated that the classical pathway of complement activ...
Phagocytosis of horse erythrocytes treated with equine infectious anemia virus by cultivated horse leukocytes. Horse erythrocytes treated with equine infectious anemia virus hemagglutinin were phagocytized by cultivated horse leukocytes (mainly macrophage-like cells and partly polymorphonuclear cells) after incubation with fresh horse serum but not with inactivated horse serum. The phagocytosis began as soon as the erythrocytes were added to the leukocyte cultures, and the majority of the reaction proceeded within 30 minutes. Addition of antiserum showed a slightly suppressing but no enhancing effect on the phagocytosis. Phagocytosis seemed to be caused by the recognition of the third complement compon...
A short, reliable, highly reproducible complement fixation test for the serological diagnosis of contagious equine metritis. A complement fixation test, using round-bottomed microtitration plates and an 8 channel microdiluter, based on that used for brucellosis by Herr, Huchzermeyer, Te Brugge, Williamson, Roos & Schiele, 1985, has been developed for use on the sera of horses to detect antibodies to the contagious equine metritis organism. The results with 2 known positive sera tested 116 times in 27 separate tests were reproducible for the most part within a twofold range. They seldom exceeded these limits and never exceeded a fourfold range. The test itself is capable of being carried out within 90 min. The test w...
Observations on the long term effects of Brucella abortus infection in the horse, including effects during pregnancy and lactation. Five mares and a stallion were studied from three to 30 months after experimental infection with Brucella abortus strain 544. The mares bred normally. No organisms were recovered from horses or from pregnant Friesian heifer contacts. Titres of serum antibody in the antiglobulin (Coombs) and complement fixation tests fell more slowly than those assessed by other tests. The serum of one foal yielded maternal antibody. An intradermal test was positive in infected adults only, and negative in all foals.
Dourine in Southern Africa 1981-1984: serological findings from the Veterinary Research Institute, Onderstepoort. The distribution of positive dourine cases found on the complement fixation test at the Veterinary Research Institute, Onderstepoort from 1981 to 1984, is recorded. Within the Republic of South Africa, foci of infection occurred in the Johannesburg, Pretoria, Potchefstroom, Rustenburg, Upington, Lichtenburg, Kroonstad, Louis Trichardt, Middelburg (Cape) and Mossel Bay state veterinary districts. In Bophuthatswana, Transkei, Lesotho, South West Africa and Swaziland, positive cases were also recorded. Anti-complementary activity of horse sera does not present a problem. In donkey and mule sera, ...
Experimental Babesia equi infection in mature horses. Nine 4-year-old Arabian geldings were experimentally infected with Babesia equi of European origin. All horses developed detectable parasitemia an average of 30 days after they were inoculated, which was accompanied by a decrease in PCV. The infections were generally mild with no animal deaths. All horses became serologically positive by the indirect fluorescent antibody test within an average of 23 days after they were inoculated and by the complement-fixation test 30 days after they were inoculated.
Serodiagnosis of experimental and natural Babesia equi and B. caballi infections. The sensitivity and specificity of the complement fixation (CF) test for the diagnosis of Babesia infections in equines was assessed, using the indirect fluorescent antibody (IFA) test as a reference. Antibodies were first detected between 11 and 20 days post infection (dpi) in the CF test and between 7 and 14 dpi in the IFA test in ponies infected experimentally with B. equi (USDA strain). The CF test became negative in four of five ponies 63-174 dpi although B. equi was demonstrated microscopically in two of these four ponies up to 364 and 455 dpi. The IFA test remained positive up to 476 dp...
