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PubMed1992

Demonstration of the humoral immune response of horses to Babesia caballi by western blotting.

Babesia caballi-infected or normal equine erythrocytes were solubilized in sodium dodecyl sulfate (SDS) buffer and analyzed by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. Antigens were allowed to react with sera from horses experimentally or field-infected with B. caballi and with sera from non-infected horses. Major babesial antigens recognized by immune sera had apparent mol. wts of 141, 112, 70, 50, 48, 34, and 30 kDa. The polypeptides at 50 and 48 kDa were recognized earliest and throughout infection, but also weakly by 3/100 equine sera tested negative and 1/33 sera tested false positive by the complement fixation test (CFT) and immunofluorescence antibody test (IFAT). Thus, further characterization and purification of B. caballi antigens are required to identify target antigens for an improved enzyme immuno assay. Until such an assay is available, Western blotting can provide a specific tool for the diagnosis of B. caballi infections, particularly in cases of contradicting CFT and IFAT results.

Animals↗
PubMed2002

Effects of vaginal Brucella ovis infection of red deer hinds on reproductive performance, and venereal transmission to stags.

AIMS: To investigate the effects of vaginal Brucella ovis infection on the reproductive performance of red deer (Cervus elaphus) hinds. To determine whether stags may become infected with B. ovis by venereal transmission from mating infected hinds. METHODS: Thirty mixed-age red deer hinds serologically negative for B. ovis antibodies were synchronised for oestrus on 22 March 2000. B. ovis was inoculated into the vagina of each hind at oestrus and again, 18 days later. At oestrus, hinds were randomly allocated to six groups, each joined with a 16 month-old red deer stag seronegative for B. ovis, for 55 days. Hinds were blood sampled and scanned for pregnancy using rectal ultrasonography at monthly intervals. Six pregnant and four non-pregnant hinds were slaughtered pre-calving and three hinds were slaughtered post-calving. Reproductive tracts and foetuses were examined grossly, histologically and microbiologically. Calves were identified and blood sampled within 3 days of birth. Hinds and calves were blood sampled in February and May 2001 and vaginal swabs were collected from hinds for B. ovis culture. Blood was collected from stags, 5 and 19 days after mating and semen was collected for B. ovis culture. The 17 remaining hinds were mated in 2001 to two mixed-age wapiti (Cervus canadensis) stags. Both stags were blood sampled after mating. Sera were tested in a B. ovis complement fixation test (CFT) and enzyme-linked immunosorbent assay (ELISA). RESULTS: All 30 hinds developed B. ovis antibody levels, measurable using either the CFT or ELISA, but these did not remain elevated. There was no evidence of infection, either by gross pathology, histopathology or microbiological culture in the ten hinds or six foetuses slaughtered pre-calving. All remaining 20 hinds produced normal calves, 15 of which survived until weaning. Three hinds experienced dystocia and gave birth to dead calves and two calves died within 4 days of birth. One hind which had dystocia was euthanased. Samples from this hind and from 3/5 dead calves showed no evidence of B. ovis infection. B. ovis was cultured from the vagina of 1/19 hinds 48 weeks after inoculation, at which time B. ovis CFT and ELISA results for this hind were negative. Most calves had B. ovis serum antibodies at 1-3 days of age but levels were negligible when sampled at 10-15 weeks of age. Foetuses and dead calves were all seronegative. Three of the five red deer stags used for mating became infected with B. ovis. The two wapiti stags used to mate the remaining 17 hinds the following year remained seronegative. CONCLUSIONS: B. ovis is unlikely to have significant detrimental effects on the reproductive performance of red deer hinds. Venereal transmission via the vagina of hinds is a possible route of transmission between stags. It is possible that survival of the organism in the vagina of some hinds could create difficulties in disease control programmes. CLINICAL RELEVANCE: B. ovis infection of hinds at the time of mating is unlikely to cause significant reproductive losses. Venereal transmission of B. ovis between stags via the hinds may occur when groups of hinds are joined with more than one stag.

Journal Article↗
PubMed1993

Coxsackievirus B1-based antibody-capture enzyme-linked immunosorbent assay for detection of immunoglobulin G (IgG), IgM, and IgA with broad specificity for enteroviruses.

An antibody-capture enzyme-linked immunosorbent assay (ELISA) with coxsackievirus B1 as the antigen was evaluated for detection of immunoglobulin G (IgG), IgM, and IgA antibodies and showed broad specificity for enteroviruses. In total, 116 serum or cerebrospinal fluid samples from 62 patients were tested by ELISA and the complement fixation test (CFT). Additionally, 15 serum samples that contained poliovirus-specific IgM antibody were tested. Serum samples from 200 healthy blood donors were used for standardization of the assays. The sensitivity of the ELISA varied with time of serum sampling, with a relatively low sensitivity when serum was collected within 3 days after the onset of symptoms (23%; 5 of 22) but good sensitivity when serum was collected later (83%; 20 of 24). The sensitivity was better than that of the CFT. The ELISAs were broadly reactive as concluded from typing of virus isolates that were simultaneously obtained. The assay did, furthermore, detect antibody against poliovirus type 3. Sera that contained rheumatoid factor, antinuclear antibody, or cardiolipin antibody (by the Venereal Disease Research Laboratory test) did not react in this ELISA. Nonspecific reactivity did occur, however, in cases of infectious mononucleosis and in Mycoplasma pneumoniae infection. The enterovirus-specific ELISA is found to be simple to perform, more sensitive than the CFT, and far less laborious than the neutralization test.

Acute Disease↗
PubMed1984

Rapid, homogeneous phase, liposome-based assays for total complement activity.

A simple, rapid assay for determining total complement activity has been developed. Complement activity is quantitated spectrophotometrically by measuring the amount of liposome-encapsulated enzyme unmasked by the action of anti-Dnp antibody and complement on Dnp-tagged liposomes. The assay is homogeneous in nature and is nonisotopic. The activity of complement in guinea pig serum has been measured and shown to be proportional to complement concentration. The assay was modified to measure the complement-fixing titer of anti-Dnp antibody preparations. We have compared two monoclonal anti-Dnp antibodies (IgG1 and IgM) for their ability to fix complement. The IgM antibody preparation was 450-fold more effective than the IgG1 preparation in mediating complement-dependent damage to Dnp liposomes. In addition, the test was modified to measure complement fixation by soluble antigen-antibody complexes. This complement fixation format is capable of detecting 2 pmol Dnp antigen.

2,4-Dinitrophenol↗
PubMedDate not supplied

Seroprevalence of rubella and cytomegalovirus (CMV) infection in pregnant women from Kerala State.

Prevalence of rubella antibody in 536 and cytomegalovirus (CMV) antibody in 260 pregnant women attending an antenatal clinic, in India, was studied by means of hemagglutination inhibition (HAI) test (for rubella antibody) and complement fixation test (CFT-for CMV antibody). The antibody detection rate in pregnant women was 74.1% for rubella and 85.4% for CMV. The prevalence rate of CMV infection was higher during third trimester (89.6%), while rubella antibody was more prevalent during second trimester (77.5%). The geometrical mean titre (GMT) was 73 for rubella antibody and 57 for CMV. Further, rubella antibody was found to be more prevalent in the age group of 26-30 years (76.8%), while for CMV, the highest prevalence rate was seen in 21-25 years (89.2%). The present study indicates the potential possibility of primary rubella infection in 26.0% and CMV infection in 15.0% of the susceptible women during pregnancy and the development of congenital malformations in children born to the infected women.

Age Factors↗
PubMed1992

Prevention of prenatal toxoplasmosis by serological screening of pregnant women in Austria.

In 1975 Austria introduced an obligatory serological screening of pregnant women for toxoplasmosis. Every woman is tested for antibodies at the beginning of her pregnancy and, in case of seronegativity, again in the second and third trimester. Basic tests are--alternatively--Dye test (SFT) and Indirect Fluorescent Antibody test IFAT); for further clarification, complement fixation test (CFT) and, particularly, various tests for detection of specific IgM and IgA antibodies and, in certain cases, for circulating antigen are carried out. If a primary Toxoplasma gondii infection of the pregnant woman is suspected, immediate therapy--with spiramycin before the 16th week of gestation and with pyrimethamin plus sulfadiazin after the 15th week of gestation--is carried out. Before the introduction of the screening programme, the incidence of prenatal toxoplasma infections was 50-70 per 10,000 births, presently it is below 1 per 10,000 births. Seropositivity among pregnant woman has decreased from almost 50% at the end of the seventies, to 36.7% in recent years (1989-1991). The percentage of suspected primary infection during pregnancy has, however, in the same period increased from less than 0.4% to 0.83%.

Austria↗
PubMed2004

Serological diagnosis of bovine brucellosis: a review of test performance and cost comparison.

The authors reviewed over 50 publications in which the sensitivity and specificity values of assays used for the detection of exposure to Brucella abortus had been examined. The sum of the sensitivity and specificity values for each test was averaged to give a performance index (PI) and allow for a comparison between the different methodologies. A score of 200 was perfect. Based on the PI, the buffered antigen plate agglutination test (BPAT) rated highest (PI = 193.1) among the conventional tests. This indicates better accuracy than the other conventional tests including the Rose Bengal test (PI = 167.6) and the complement fixation test (PI = 172.5). Overall, the primary binding assays, including the fluorescence polarisation assay (PI = 196.4), the indirect enzyme-linked immunosorbent assay (PI = 189.8) and the competitive enzyme-linked immunosorbent assay (PI = 188.2), were more accurate than the conventional tests, except for the BPAT. In addition, a fee comparison suggested that the primary binding tests were price competitive with conventional tests for the diagnosis of brucellosis and, therefore, had a better combined cost/efficiency rating.

Agglutination Tests↗
PubMed1986

Experimentally induced Brucella abortus infection in pregnant goats.

Pregnant goats in midgestation (7 to 16 weeks) were conjunctivally exposed to Brucella abortus strain 2308 to evaluate their applicability as an animal model for bovine brucellosis. Brucellae were isolated from uterine fluid and/or placental specimens of 10 of 12 does at parturition. Six of the 10 infected does delivered dead fetuses and 1 of the 10 delivered live, premature twins. Dead fetuses typically contained brucellae in multiple tissues, whereas brucellae generally were not isolated at birth from live kids. After parturition, B abortus was excreted in the milk and uterine fluids of the infected does. At necropsy (6 weeks after parturition), organisms in the doe were primarily in the uterus and in the lymph nodes that drained the mammary glands, uterus, and head. Brucella abortus was most often isolated from the cranial lymph nodes of neonates that had remained with their dam for 6 weeks after parturition. Serum anti-Brucella antibody concentrations were determined by use of standard tube agglutination, mercaptoethanol agglutination, Rivanol plate tests, card tests, complement fixation, hemolysis-in-gel tests, and an enzyme-linked immunosorbent assay. Serologic responses were detected 2 to 3 weeks after exposure and remained detectable until parturition. Antibody titers increased after parturition in does shedding B abortus at parturition. Anti-Brucella antibody was not detected in neonates before colostrum intake. The neonate's postcolostral titers were similar to those in the dam at the time of parturition. Milk anti-Brucella antibody was detected in milk (milk ring test) from infected and noninfected mammary glands.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1979

Immunogenic complexes obtained from Salmonella typhi-murium and Salmonella typhi Ty2 by the bacterial acetone powder method.

An immunogenic complex was isolated from Salmonella typhi-murium and another one from Salmonella typhi Ty2. Both were prepared by the bacterial acetone powder method which eliminated the cell wall, the DNA almost completely and the membrane phospholipids. The complexes were denominated "New Vaccines". The S. typhi-murium new vaccine induced, even at doses of 0.5 microgram dry weight per mouse, a high degree of protection against the challenge of the virulent microorganism. By immunoelectrophoresis, 21 antigen-antibody systems could be detected, two of them corresponding to O antigens. The S. typhi Ty2 new vaccine induced better protection than the standard vaccine (heat-phenol inactivated typhoid vaccine) when both vaccines were compared in the relative potency test. Moreover, the new vaccine had very low toxicity when inoculated in humans at doses of 1 microgram dry weight, able to elicite a high antibody titre (1/1,790 mean of 10 sera) in 75% of the tested population, estimated by the complement fixation test. In contrast, the standard vaccine induced a low antibody titre (1/222, mean of 5 sera) in 50% of the humans inoculated with 1 X 10(8) bacterial cells. The new vaccine did not induce undesirable effects whereas the standard vaccine induced an important inflammatory process in 100% of the cases, with intense local pain in 67% after 24 h post-first inoculation as well as other less severe symptoms.

Acetone↗
PubMedDate not supplied

[Detection of oncornavirus D-induced antigen in cells by a mixed hemadsorption method].

Further study of a new oncornavirus D-induced antigen was done. Its immunological specificity and localization in cells was determined by a sensitive method of mixed hemadsorption. A continuous HEp-2 cell culture was used as the source of the new as well as virus antigen. Adsorbed specific sera to the new antigen and to oncornavirus D obtained by the previously developed method were used for the antigens detection. This method demonstrated localization of both virus antigens and oncornavirus D-induced antigens not included into the virion structure on the surface of oncornavirus D-infected cells. Mixed hemadsorption test as well as complement-fixation test gave the results identical in their specificity indicating the presence in HEp-2 cells of a new non-virion antigen.

Animals↗
PubMed1984

Remarks on the diagnosis of Clonorchis sinensis infection.

A number of techniques have been developed for the diagnosis of Clonorchis sinensis infection. Although the most reliable technique for this infection are the various stool examinations in the laboratory, practical problems such as difficulties of stool collection and mass examinations need more applicable methods for epidemiological studies. During past decades some serologic and immunologic methods such as intradermal skin test, gel-diffusion technique, complement fixation test, haemagglutination test, fluorescent antibody test and enzyme-linked immunosorbent assay (ELISA) have been introduced for the diagnosis of clonorchiasis in Korea, but satisfactory results have not been obtained especially in field work because of the lack of available purified antigen. From this viewpoint it is essential to undertake careful isolation and purification of specific antigens. Recently ELISA has been well established as a micro-system for mass survey. In spite of the complexity of its procedures in contrast to the skin test the results obtained explain its importance. In this paper available serologic and immunologic methods for the diagnosis of clonorchiasis used at the Institute of Tropical Medicine, Yonsei University, Seoul, are reviewed, and the results from skin test and ELISA are compared and discussed.

Antibody Specificity↗
PubMed1975

Absence of Anaplasma marginale infection in American Bison raised in an anaplasmosis endemic area.

Blood was collected at slaughter from 132 adult American bison (Bison bison) raised in an anaplasmosis endemic area where the vector Dermacentor andersoni (equals venustus) is indigenous. Hematologic studies revealed no indication of clinical anaplasmosis. Card agglutination and complement-fixation tests on all bison serums were negative. Eleven anaplasmosis-susceptible calves each inoculated with 204 ml of blood pooled from 12 bison did not develop anaplasmosis. Results of this study indicate American bison have resistance to natural A. marginale infection.

Agglutination Tests↗
PubMed1984

An hemolysis-in-gel test for bovine brucellosis.

An hemolysis-in-gel test (HIGT) for bovine antibody against Brucella abortus was developed and evaluated. Sera to be tested were placed in wells in an agarose gel containing guinea pig complement and J-negative bovine erythrocytes coated with lipopolysaccharide prepared from B. abortus biotype 1. After incubation, zones of hemolysis were produced by positive sera. The activity of some positive sera was heat labile, but was restored to heated sera by addition of a crude preparation of the first component of bovine complement. The HIGT was compared with the complement fixation test (CFT), the buffered antigen plate test (BPAT), the standard tube agglutination test (STAT) and the standard plate agglutination test (SPAT), using sera from 1041 brucellosis-free cattle, from 51 cattle infected with B. abortus biotype 1 or biotype 4, and from six heifers vaccinated with strain 19. Three of 1041 sera (0.29%) from brucellosis-free cattle were HIGT-positive, ten (0.96%) were BPAT-positive, and none were positive in the CFT, STAT, or SPAT. The HIGT was more sensitive, and detected infection earlier than the other tests in the case of B. abortus biotype 1 infection, but was less sensitive for biotype 4 infection. In vaccinated heifers, HIGT reactions appeared later than reactions to the other tests, and persisted as long as 565 days. Further studies are needed to standardize the HIGT test conditions and to improve its sensitivity for B. abortus biotype 4 infections. Attempts to determine the effects of LPS antigen prepared from different biotypes of B. abortus are in progress.

Agglutination Tests↗
PubMed1986

Evaluation of laboratory diagnosis of toxoplasmosis by means of an ELISA-triple test. Detection of class specific IgG, IgM and circulating antigen.

The combination of three parameters (IgG, IgM and circulating antigen) in a so-called 'ELISA triple test' was suggested for advantageous diagnosing of human toxoplasmosis. A qualitative assay was used with the following arbitrary assumptions: IgG antibodies reflect an infection, IgM antibodies reflect a recent (primary) infection and circulating antigens reflect an active infection. The three assays were performed simultaneously in one microtiter plate. This approach was tested with 1091 patient sera submitted for routine diagnosis. In comparison with conventional indirect immunofluorescence and complement fixation test it was observed that combinations indicating a recently acquired infection (combinations with IgM and/or circulating antigen) mainly paralleled low or negligible conventional antibody titers. No strict association was seen between particular combinations and certain clinical symptoms suggestive for toxoplasmosis. In conclusion it was stated that the triple test for support of clinical diagnosis has some advantages but that a strong need exists to be able to demonstrate exacerbation or reinfections which are generally not characterised by IgM antibody formation of free circulating antigens but may be the reason for elevated conventional antibody titers.

Antigens, Protozoan↗
PubMed1975

[Value of TPHA in the serological screening of lues].

In the present work, 21,400 not selected sera were tested with hemagglutination test (TPHA) and flocculation test (VDRL) with cardiolipin for screening of syphilis; sera that gave negative results against both tests showed remarkable agreement (97,81%). 468 sera gave positive results to least one of the tests employed in screening and were tested with the following procedures: fluorescent Treponemal antibody-absorption (FTA/ABS) test, complement fixation (by Kolmer, modified) with proteic (KP) and lipidic (KL) antigen; 100 of these sera were also tested with T. pallidum immobilizaiton (TPI) test. The agreement among the before mentioned tests and sera that gave positive results only to VDRL was significatively low; on the contrary, the agreement was remarkable in the case of those positive only to TPHA (99% with FTA/ABS, 83% with TPI, 52,5% with KP and 21,5% with KL). The results obtained in attesting sensitivity and specificity of TPHA indicate that it is a very valuable screening test, and suggest to the AA. to emphasize that TPHA/VDRL association a definitive conclusion in syphilis serological negativity.

Antibody Formation↗
PubMed1989

[The problem of homologous blood in transfusiology and its solution].

In cardiosurgery conducted in this country and abroad the development of homologous blood syndrome was observed. Its frequency comprises up to 2.5%, according to the authors' data. It has been established that the syndrome is the result of isoimmunologic incompatibility by the antigenic systems of blood plasma allogeneic proteins. The authors have proposed the testing of blood compatibility by the agglutination methods according to the erythrocytic antigen systems with the use of the complement-fixation test according to the protein-plasma antigen systems, due to these tests the development of homologous blood syndrome has been completely eliminated from their practice. The development of massive blood transfusion syndrome described by some transfusiologists has been rejected by the authors, it is considered by them as manifestation of insufficient blood compatibility of the test animals as a result of a wrong method of their isoimmunologic selection using the only cross-testing.

ABO Blood-Group System↗
PubMed1992

[Q fever antibody titer--follow-up study in cattle with special reference to pregnancy].

In 290 Q fever positive cattle from three 2000 head dairy farms in the former district of Erfurt (Thüringen) the course of titers was examined serologically over several months by means of the complement fixation test (CFT). In 47.2% of the cows serologically observed for 2 up to 28 months complement fixing antibodies against Coxiella burnetii could be demonstrated til the end of the investigation period. Repeated tests during pregnancy showed increase of antibody titers in the first 4 months and after a short decrease again from the 5. til the 7. month. By observing the antibody titers during several pregnancies each time a new increase comparable to a booster immunisation could be found. This may explain the persistence of Q fever antibodies in cows for several years. The results of this investigation suggest that from a high antibody titer it can not be concluded an abortion in a positive cow being caused by a Coxiella burnetii infection.

Animals↗
PubMed1998

Eradication of brucellosis from a cattle herd in the Amazonian region (short communication).

The authors intend to elaborate a brucellosis eradication program which could be used in the Amazonian region, an area characterised by extremely extensive animal husbandry practices. Under such conditions, eradication by selection is the only feasible approach. Brucellosis has been successfully eradicated from a herd with 22.1% prevalence of infection by two serological surveys using an indirect ELISA and the complement fixation test.

Agglutination Tests↗