Comparison of Indirect Hemagglutination and Immunodiffusion Tests for Detecting Type II Leukosis (Marek's) Infection in S- and K-Line Chickens

The indirect hemagglutination and immunodiffusion tests were compared for detection of antigen and antibody to JM strain of leukosis virus infection between S- and K-line chickens. The indirect hemagglutination test was more sensitive than the immunodiffusion test for detecting the smallest amount of viral antigen and corresponding antibody in the plasma of infected chickens. The Cornell S-line had higher levels of antigen and antibody as compared with the Cornell K-line during the 20-week experimental period.     

Immunogenicity of Trypanosoma cruzi in different animal species

The immunogenicity of two fractions (1 500 F and 10 000 F) from epimastigotes of Trypanosoma cruzi as well as the supernatant from culture media (SF) were studied using hens, rabbits and opossums. For comparative purposes, sera from individuals with chronic Chagas' disease were also used. A similar, positive response was obtained for the fractions in all the animal species studied using indirect hemagglutination test. Supernatants from culture media were the least immunogenic. By double immunodiffusion test, it was possible to detect a positive response to a different number as well as to different antigens in the three animal species, but there was response to a common antigen by all the different animal species. The common antigen called here major, was present in all the fractions assayed. Human sera from individuals chronically infected showed a variable response. When assayed by double immunodiffusion technique, the major antigen could be detected in just a few samples.     

Estimation and Identification of Streptococcus dysgalactiae Protective Antibody in Sera of Rabbits and Cattle

Rabbit and cow anti-Streptococcus dysgalactiae sera were tested by bacterial agglutination, complement fixation, hemagglutination, and immunodiffusion for the presence of antibody. The results of these tests were compared with mouse-protection studies on the same serum to estimate which in vitro test would best reflect the in vivo protective capacity of serum. Identification of the antibody constituents responsible for the mouse protection, hemagglutination, and complement fixation titers were established by reacting whole and diluted antisera with mercaptoethanol before and after testing. Results indicate that the complement fixation test may be a more accurate indicator of IgG protective bovine and rabbit antibody, whereas the hemagglutination test may more readily reflect a wider range of protective antibody levels and IgM. The complement fixation test showed some shared responses to IgG and IgM in both the rabbit and cow, whereas the IgM components seemed to be the predominant factor influencing hemagglutination titers in the rabbit and more so in the bovine. Mouse protection tests with mercaptoethanol-treated cow and rabbit sera indicate that the protective capacity of these antisera is shared between IgM and IgG components.     

Detection of HBs antigen with latex particles coated with human antibody prepared by affinity chromatography on concanavalin A-sepharose.

Human HBs antibody was isolated by affinity chromatography on HBs antigen absorbed to concanavalin A linked to Sepharose 4B. When a human anti-HBs immunoglobulin preparation obtained by Cohn's cold ethanol fractionation method was used as a starting material, the antibody was concentrated about 10 times in terms of the passive hemagglutination titer with a recovery rate higher than 50%. Latex particles coated with human anti-HBs antibody thus prepared were proved to be useful in detecting HBs antigen in human blood samples. In its sensitivity and in rapidity of its performance, the antibody-coated latex agglutination test seems to be superior to conventional immunodiffusion techniques.     

Echinococcus granulosus: host lymphocyte transformation by parasite antigens.

Lymphocyte transformation, measured by in vitro tritiated thymidine incorporation, and indirect hemagglutination tests were carried out on hydatid patients and normal individuals using sheep and human hydatid fluid or scolex antigens. The hydatid patients showed statistically significant lymphocyte transformation with human and sheep hydatid fluid or scolex antigens when compared to normal individuals. The indirect hemagglutination tests resulted in high titers of antibody with sheep or human hydatid fluid antigens, while very low titers were obtained with scolex antigens. Unlike in the indirect hemagglutination test, the source of the antigen, scolex or fluid, was not of consequence in the lymphocyte transformation test. Furthermore, there was no correlation between the results of the serologic and lymphocyte transformation tests, since some patients with very high lymphocyte stimulation indices produced low indirect hemagglutination titers and vice versa. Similar results were obtained from rabbits which were immunized with sheep hydatid fluid or scolex extracts. The skin tests were of the immediate type of hypersensitivity reactions. Delayed skin reactions did not occur in spite of the presence of sensitized lymphocytes in the blood of the immunized rabbits.     

Multiple Group-specific Antigen Components of Avian Tumor Viruses Detected with Chicken and Hamster Sera

Multiple group-specific (gs) components of the avian leukosis-sarcoma viruses were detected by immunodiffusion (Ouchterlony) tests with sera from hamsters bearing tumors induced by sarcoma viruses and with sera from adult chickens immunized with avian sarcoma or leukosis viruses. Immune hamster sera detected up to four components, whereas chicken sera detected at least one. The hamster and chicken sera identified a similar antigen, as indicated by reactions of identity. Relatively few chicken sera containing neutralizing antibody to avian sarcoma or leukosis viruses reacted in immunodiffusion with the gs antigen. The gs components were released from the virion by various means of disruption, including freezing and thawing. Tests with tissues from normal chickens and from chickens with Marek's disease failed to demonstrate any reactions with hamster or chicken gs antiserum.     

Rapid, Sensitive Assay for Staphylococcal Enterotoxin and a Comparison of Serological Methods

Reversed passive hemagglutination was used to assay enterotoxin in culture filtrates and in food samples. With cells tanned and then sensitized with antitoxin globulin and preserved with either formaldehyde or pyruvic aldehyde, as little as 0.0007 mug of enterotoxin was detectable. The results of hemagglutination tests compared well with those obtained by quantitative precipitin tests or by immunodiffusion, but hemagglutination was 50 to 100 times more sensitive than the immunodiffusion technique. In addition, results of the hemagglutination test were available within a few hours, and neither elimination of interfering proteins from food extracts nor concentration of the sample, both of which are necessary for immunodiffusion, was required for this procedure.     

Seroimmunological and parasitological surveys of leucocytozoon caulleryi infection in chickens in several asian countries

Epizootiological surveys of Leucocytozoon caulleryi infection in chickens in Japan, Taiwan, Philippines, Singapore, Malaysia and Thailand were undertaken by means of the immunodiffusion test. The rate of infection of L. caulleryi confirmed by the examination of parasites in the peripheral blood of chickens coincided with that of positive antibody response in the immunodiffusion test. Antibodies against L. caulleryi were found in chickens in all the countries surveyed in the present investigation. The prevalence of L. caulleryi infection in chickens was confirmed by the immunodiffusion test. Several chickens in each country showed the presence of serum antigens of L. caulleryi at the times of serum sample collection. These results seemed to indicate that the immunodiffusion test is a method efficient enough to be applicable to the epizootiological surveys and diagnosis of L. caulleryi infection in chickens in the field. As a result, the antibodies or soluble antigens in the sera of chickens infected with L. caulleryi present, respectively, in each country may have the same immunological characters.     

Radial Immunodiffusion: a Simple and Rapid Method for Detection of Marek''s Disease Antigen(s)

A qualitative radial immunodiffusion technique is described which detects antigen(s) in feathers from live or dead chickens infected with Marek's disease herpesvirus. Antiserum, which is incorporated into a support medium, reacts with antigen(s) in the feather tip producing a radial precipitin ring. Antigen(s) was detected in 93.3% of experimentally inoculated chickens 21 days postinoculation and in 100% of infected birds subsequently tested through 6 weeks. No antigen was detectable in the feathers of uninoculated control chickens. The technique is simple and rapid to perform. Positive tests could be detected after 1 to 2 hours of incubation. Antigen detection by the radial immunodiffusion test correlated well with other criteria of infection. This technique should have application as a laboratory research tool and as an adjunct for a rapid flock diagnosis of Marek's disease.     

Serological diagnostics of imported malaria in Czechoslovakia

Two serologic techniques for malaria detection were compared in this study; the indirect fluorescent antibody (IFA) test used in 214 persons (38 Czechoslovak citizens returning from visits to tropical countries and 176 foreign visitors arriving to Czechoslovakia from areas endemic for malaria) and the indirect hemagglutination (IHA) test employed in 125 persons (29 Czechoslovak citizens and 96 foreigners). Comparisons revealed poor correlation between the IFA test and IHA test data. Of the two tests the IFA test appeared to be distinctly more reliable, more sensitive and more specific, the IHA test turned out to yield both false positive and false negative results. The antigen from Plasmodium gallinaceum gave lower IFA titres than P. falciparum antigen, but reacted with antibodies to all species of human plasmodia, and gave reliable test results. Positive serologic responses were appreciably more frequent in foreigners (46.0%) than Czechoslovak citizens (23.7%). The maximum percent positivity for malarial antibody was among individuals from tropical countries of Africa (74.6%), seropositivity in people from malaria endemic areas in Asia and Latin America was far less frequent (28.4% and 44.4%, respectively).     

Radial Immunodiffusion: a Simple and Rapid Method for Detection of Marek's Disease Antigen(s)

A qualitative radial immunodiffusion technique is described which detects antigen(s) in feathers from live or dead chickens infected with Marek''s disease herpesvirus. Antiserum, which is incorporated into a support medium, reacts with antigen(s) in the feather tip producing a radial precipitin ring. Antigen(s) was detected in 93.3% of experimentally inoculated chickens 21 days postinoculation and in 100% of infected birds subsequently tested through 6 weeks. No antigen was detectable in the feathers of uninoculated control chickens. The technique is simple and rapid to perform. Positive tests could be detected after 1 to 2 hours of incubation. Antigen detection by the radial immunodiffusion test correlated well with other criteria of infection. This technique should have application as a laboratory research tool and as an adjunct for a rapid flock diagnosis of Marek''s disease.     

Diagnosis of Corynebacterium pyogenes infection in pigs by immunodiffusion test with protease antigen.

A protease antigen was prepared from the culture supernatant of Corynebacterium pyogenes by concentrating with a flash evaporator and ultrafiltration. It was adjusted to the concentration of 32 units by the single radial immunodiffusion with a tentative standard serum. In the immunodiffusion test, the antigen of 4 units reacted enough with sera having an antibody titer ranging from 1 to 128. As a result, it was decided that the antigen of 4 units should be used in the immunodiffusion test for the detection of protease antibody. By the immunodiffusion test, protease antibody was demonstrated in about 35% of 443 sera from pigs collected at random. The antibody titer showed the distribution of 2 peaks. The summits of the two peaks were seen at 4 and 32 of antibody titer, respectively. The valley between the two peaks was seen at 16 of titer. From the result, a diagnostic criterion of the immunodiffusion test was decided provisionally as follows: above 16 of antibody titer is positive, 1 to 8 suspect, and less than 1 negative. On the other hand, protease antibody was demonstrated in sera from 13 of 14 pigs carrying abscesses from which C. pyogenes had been isolated. Its titer was 8 (in 2 pigs), 16 (in 1), 32 (in 3), 64 (in 6), and 128 (in 1). From these results, it was proposed that the immunodiffusion test with protease antigen be used for the diagnosis of C. pyogenes infection in pigs.     

Use of monoclonal antibodies for the production of a plague antibody erythrocyte diagnostic agent

Monoclonal antibodies to Yersinia pestis capsular antigen were fixed onto the surface of formulated sheep red blood cells. The preparation thus obtained was compared with commercial antibody erythrocyte diagnosticum in the passive hemagglutination test aimed at the search for the capsular antigen in the suspensions of Yersinia pestis museum cultures and in the antigen neutralization test aimed at the search for antibodies in the sera of wild and laboratory animals having had plague. Monoclonal erythrocyte diagnosticum proved to be suitable for the detection of both the capsular antigen and antibodies. The comparison of the results of the passive hemagglutination test and the enzyme immunoassay demonstrated the presence of very close relationship between them.     

Influenza A Neuraminidase Antibody Assay with Sensitized Erythrocytes

Erythrocytes sensitized with purified neuraminidase (Hong Kong) antigens were used for assay of influenza A neuraminidase antibodies. The neuraminidase indirect hemagglutination test was equal to the neuraminidase hemagglutination-inhibition (enhancement) test and appeared to be better than the neuraminidase inhibition test for detection of fourfold or greater antibody rises in paired sera from influenza patients or vaccinees. It was better than both tests for detection of neuraminidase antibody. The neuraminidase indirect hemagglutination test is simple to perform and has the advantage of direct antigen-antibody assay.     

A New Method for Strain Identification of Rhizobium trifolii in Nodules

An indirect haemagglutination test has been developed for the detection of strains of Rhizobium trifolii in nodules of subterranean clover plants. Preserved sheep red blood cells, coated with isolated specific rhizobial lipopolysaccharide, were used as the indicator of agglutination; these cells were agglutinated by specific antilipopolysaccharide antibody. Detection of lipopolysaccharide antigen in a suspension of nodular tissue was carried out by reacting the suspension with antilipopolysaccharide antibody prior to the addition of coated red blood cells. The presence of antigen in the suspension was indicated by an inhibition of agglutination. The test was more sensitive than agglutination and immunodiffusion in the detection of rhizobial lipopolysaccharide antigens, and could be used for the rapid screening of large numbers of nodules.     

Comparison of the Reliability and Sensitivity of Three Serological Procedures in Detecting Antibody to Yersinia pestis (Pasteurella pestis)

Three serological procedures, the agar-gel precipitin inhibition, the complement fixation, and the indirect hemagglutination tests, were used to detect and measure antibody to Yersinia pestis in the sera from 383 individuals. Although all three tests were useful in detecting plague antibody, the most reliable and sensitive test procedure was indirect hemagglutination.     

Enzyme-linked immunosorbent assay for efficient detection of antibody to bluetongue virus in pronghorn (Antilocapra americana)

An indirect enzyme-linked immunosorbent assay (ELISA), using cell-associated viral antigen, was developed for detection of antibody to bluetongue virus (BTV) in field-collected pronghorn (Antilocapra americana) sera. To test the applicability of the ELISA to seroepizootiologic studies, pronghorn serum samples from three Wyoming counties (USA) were tested. Bluetongue virus ELISA results were compared to those of the bluetongue immunodiffusion assay. Discrepant serum samples were retested for reaction to either BTV or epizootic hemorrhagic disease virus. The pronghorn BTV ELISA gave rapid, quantitative, objective results and should facilitate testing large numbers of sera for BT diagnostic and seroepizootiologic studies.     

Malaria: seroepidemiology and serologic diagnosis

Recent advances on the application of serologic methods employing the indirect hemagglutination test with a Plasmodium knowlesi antigen for the study of malaria epidemiology are outlined. Work in progress on the stabilization of malaria antigens and the preparation of gluteraldehyde sensitized cells were reviewed. Fluorescent antibody studies in progress are discussed and work on the cross reactivity of Babesia antigens with malaria is mentioned.     

Hemagglutination-Inhibition Method and Immunofluorescence Staining with Venezuelan Equine Encephalomyelitis Virus

Hemagglutination and fluorescent antibody (FA) are compared for the direct detection of virus devoid of host cells. A determination was made of the minimal number of tissue plaque-forming units of Venezuelan equine encephalomyelitis virus that could be detected by the hemagglutination technique. Similar concentrations of the virus in bovine albumin borate saline, Brain Heart Infusion broth (Difco), and demineralized water were tested by the FA technique. Somewhat higher concentrations of the virus in bovine albumin borate saline were used in the hemagglutination-inhibition test. The quantitative hemagglutination procedure employed for these studies was carried out at 37 C for 75 min with variations in concentration of goose red cells. As a result of lowering the red cell concentration, smaller concentrations of virus were detected. The direct FA staining procedure applied to slide preparations containing known numbers of tissue culture plaque-forming units of virus was negative. Adsorbed viral antigen on agglutinated goose erythrocytes was visualized by direct and indirect FA techniques.