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PubMed1980

Physiological and serological characteristics of 48 Pasteurella multocida cultures from rabbits.

Forty-eight Pasteurella multocida cultures collected from rabbits over a 56-year period were examined to determine their physiological characteristics and to determine their serological types in the gel diffusion precipitin test. Generally, the physiological characteristics from 30 tests were typical for P. multocida. There were a few atypical variations in the fermentation of lactose and maltose and variations in trehalose, dulcitol, xylose, sorbitol, and glycerol. Seven cultures did not produce indole, and four cultures did not produce detectable amounts of hydrogaen sulfide. In obliquely transmitted light, 26 cultures formed large, slightly iridescent, mucoid colonies, 17 cultures had iridescent colonies, and 5 cultures had blue colonies. Heat-stable antigens from the 48 cultures reacted with antisera prepared from P. multocida type cultures presenting serotypes 1, 3, 4, 12, and 15. Antigens from 15 cultures reacted slightly with antisera from more than one serotype. Overall, gel precipitin reactions involving serotype 3 (25%) and serotype 12 (66.7%) were the most prevalent.

Animals↗
PubMed1986

Evaluation of cross-reactions in Histoplasma capsulatum serologic tests.

Cross-reactivity in Histoplasma serologic tests was evaluated by using sera from patients with histoplasmosis and other infections. Serum samples from 127 of 134 (95%) patients with histoplasmosis were judged positive by complement fixation tests, and 121 (90%) showed H bands, M bands, or both by immunodiffusion. Of these 134 patients, cross-reactions were seen to Blastomyces dermatitidis in 53 patients (40%), to Coccidioides immitis in 20 patients (16%), and to Aspergillus fumigatus in 3 patients (2%) by complement fixation. Serum samples from 5 of 99 patients (5%) with other fungal infections and from 5 of 46 patients (11%) with tuberculosis had M precipitin bands by the Histoplasma immunodiffusion test, whereas none of the 123 sera from patients with other bacterial, Mycoplasma, or viral infections showed H or M precipitin bands. In the complement fixation test, positive reactions were observed in 16 of 90 patients (18%) with other fungal infections, in 14 of 41 patients (34%) with tuberculosis, and in 18 of 105 patients (17%) with other bacterial, Mycoplasma, or viral infections. Positive reactions were seen by radioimmunoassay in 54 of 110 patients (49%) with other fungal infections, in 23 of 46 patients (50%) with tuberculosis, and in 35 of 123 patients (28%) with with other bacterial, Mycoplasma, or viral infections. These results demonstrate a wider range of cross-reactions in Histoplasma serology than has been previously recognized, and the cross-reactivity was greatest when observed by radioimmunoassay. Caution should be exercised in the interpretation of serologic data from patients with suspected fungal infections.

Antibodies, Fungal↗
PubMed1976

Counterimmunoelectrophoresis of staphylococcal antibody.

Modifications of the staphylococcal counterimmunoelectrophoresis technique were evaluated to determine how variations in the procedure affected results. Neither a buffer pH range of 7.8 to 9.0 nor buffer molarity of 0.015 or 0.025 when tested in combinations caused appreciable differences. However, use of different agar preparations or delay in addition of antigen to the test slide altered the location of the precipitin band. Agarose was found to be more sensitive in determining the serum precipitin titer and provided a better photographic record than either Ionagar or Noble agar.

Agar↗
PubMed1993

Bacteroides fragilis strains express multiple capsular polysaccharides.

Previous studies by our group have demonstrated that the capsule of Bacteroides fragilis type strain NCTC 9343 consists of two chemically distinct polysaccharides, designated PS A and PS B. These polysaccharides can be isolated as an aggregate from the surface of the organism and give a complex multiprecipitin profile when they are reacted with homologous antiserum in an immunoelectrophoresis assay. Following structural analysis of PS A and PS B, we have determined that the complex precipitin profile is formed as a result of the differing electrophoretic and antigenic properties associated with each of these polymers. Presently, we have examined the capsular polysaccharides of 13 other strains of B. fragilis according to methods used for the prototype strain. The capsules of these strains were extracted, partially purified, and analyzed by immunoelectrophoresis at pH 7.3. Following reaction with homologous polyclonal antisera, each of the capsular preparations tested yielded a complex precipitin profile similar to that of the prototype strain. When reacted by immunoelectrophoresis with polyclonal antiserum to 9343 or with monoclonal antibodies to PS A and PS B, these capsular preparations appeared to be antigenically diverse; some preparations (50%) showed complete or partial cross-reaction. These results suggest that the dual polysaccharide motif seen with the prototype strain is a common feature of B. fragilis strains. In addition, the antigenic heterogeneity of B. fragilis capsular polysaccharides could be used for the development of a serological typing scheme.

Antibodies, Bacterial↗
PubMed1995

Comparison of commercially available enzyme immunoassay with traditional serological tests for detection of antibodies to Coccidioides immitis.

A newly released commercially available enzyme-linked immunosorbent assay (ELISA) was evaluated for its ability to detect immunoglobulin M (IgM) and IgG antibodies against the tube precipitin and complement fixation (CF) antigens of Coccidioides immitis. The ELISA was compared with more traditional diagnostic assays, CF, latex agglutination (LA), and immunodiffusion (ID). When the IgM-specific portion of the ELISA was compared with LA, there was an agreement of 81.8%, a specificity of 75.0%, and a sensitivity of 84.6%. For the determination of the presence of IgG antibodies, the results of the IgG-specific part of the ELISA were compared with the combined results of ID and CF. After resolution of discrepant results, there was an agreement of 95.6%, a specificity of 98.3%, and a sensitivity of 92.6%. When the results of the IgG- and IgM-specific portions of the ELISA combined were compared with the results of the three traditional assays (CF, LA, and ID) there was an agreement of 96.7%, a specificity of 98.5%, and a sensitivity of 94.8%. The ELISA proved to be a reliable assay for the detection of antibodies against the tube precipitin and CF antigens and did not suffer from the objectivity required to interpret the results of the traditional assays and anticomplement interference associated with the traditional assays.

Antibodies, Fungal↗
PubMed1998

Antitoxin-in-membrane and antitoxin-in-well assays for detection of toxigenic Corynebacterium diphtheriae.

The Elek culture plate precipitin test is routinely used for the detection of exotoxin from toxigenic strains of Corynebacterium diphtheriae. Recently, the World Health Organization standardized this test to ensure accuracy, reliability, and reproducibility. In this study, we further modified the standard Elek test by using the antitoxin-in-membrane (AIM) and antitoxin-in-well (AIW) approaches. In the AIM tests, each strain was stabbed and streaked backwards and away from a point approximately 7 mm from the edge of a sterile cellulose acetate-cellulose nitrate filter membrane disk (pore size, 0.45 microm; diameter, 25 mm) containing 25 IU of diphtheria antitoxin. For AIW tests, a central well (diameter, 5 mm) containing 9 microl of antitoxin (4.5 IU) was surrounded by eight equidistant stab-streaks of each strain placed 10 mm from the well. In both methods, precipitin bands of identity typically were noted after 24- and 48-h incubations at 37 degrees C. Both toxigenic and weak toxigenic strains gave clear and reproducible results. Compared with the standard Elek test, the AIM and AIW tests each use 50% less medium and 75 and 87% less antitoxin, respectively. AIM has the potential to test up to 14 isolates and AIW has the potential to test up to 24 isolates on the same plate. Furthermore, clearer positives were noted with weak toxigenic strains. In a blinded test of 209 verified C. diphtheriae isolates, a 99.5% agreement with the standard Elek test was obtained overall. Both modifications conserve reagents and medium, permit the simultaneous testing of a larger number of strains, and may be particularly suitable for reference laboratories or hospitals involved in diphtheria epidemic settings.

Antitoxins↗
PubMed1977

Immunological procedure for the rapid and specific identification of Coccidioides immitis cultures.

An immunological procedure for the rapid and specific identification of Coccidioides immitis isolates has been developed. The specificity of the procedure is based on the fact that C. immitis produces antigens that are not produced by morphologically similar fungi. The procedure involved the transfer of heavy mold-form inocula to flasks that contained small volumes of brain heart infusion broth. Shake cultures were grown at 25 degrees C on a gyratory shaker at 150 rpm. The concentrated supernatant antigens so obtained from broth cultures were tested in parallel with coccidioidin by a microimmunodiffusion technique against C. immitis antiserum. The ability of the immunological procedure to identify C. immitis cultures was evaluated by testing the supernatant antigens of 166 unknown isolates, many of which, by gross or microscopic examination or both, resembled C. immitis or belonged to the family Gymnoascaceae. Each culture was also identified by conventional laboratory procedures. Comparative evaluation showed that the immunological test for C. immitis was 100% sensitive and specific. The diagnostic precipitin bands that were produced by the interaction of the C. immitis supernatants and antisera were shown to consist of (i) a heat-stable antigen comparable to that active in the tube precipitin test and (ii) two heat-labile antigens, one of which was associated with complement fixation reactivity. With pure cultures, this immunological procedure permitted the identification of C. immitis isolates within 5 days.

Antigens, Fungal↗
PubMed1979

Counterimmunoelectrophoresis test for immunoglobulin M antibodies to group B coxsackievirus.

A counterimmunoelectrophoresis test was developed for immunoglobulin M (IgM) antibodies to group B coxsackievirus (CB) types 1 through 5. The IgM precipitin line could be identified and differentiated from the IgG line by treating sera with 2-mercaptoethanol. Antigen purity was demonstrated by single precipitin lines occurring only to the homologous antigen when tested with type-specific hyperimmune rabbit sera. Serum pairs from 19 of 22 patients with documented CB type 1,3,4, and 5 infections were positive for IgM antibody to the infecting serotype, whereas 2 of 7 pairs from CB type 2 patients were positive. Heterologous IgM antibodies were present in sera from 14 fo 29 CB patients. Of the 14 patients with heterologous IgM antibodies, 12 also had greater than or equal to 4-fold rises in whole serum neutralizing antibody to heterologous serotypes. Only three control sera from 72 patients with coxsackievirus group A, echovirus, or other viral infections had IgM antibody to CB serotypes.

Antibodies, Viral↗
PubMed1971

Studies of colonic carcinoma antigens.

Extracts of human colonic adenocarcinomata were made with perchloric acid and shown to contain antigens which were not present in pooled samples of normal colonic mucosa or in healthy colonic mucosa from patients providing the tumours. Antisera prepared against the tumour extracts gave two precipitin lines on immunodiffusion, and antiserum prepared in Montreal according to the methods devised by Gold and Freedman (1965a and b) also gave two lines of precipitation when tested against some of these extracts; one of these precipitin lines gave a reaction of identity with that given by the antiserum produced here. The results suggest that perchloric acid may extract two antigens from colonic carcinomata.

Adenocarcinoma↗
PubMed1979

Antibodies to germinating and yeast cells of Candida albicans in human and rabbit sera.

Two major antigenic components, I and II, were detected by double immunodiffusion in sonic extracts of the germinating (G) or yeast (Y) cells of the dimorphis organism, Candida albicans group A. Component I may be a heterogeneous mixture of antigens which are stable to heating and phenol. Component II is more homogeneous but is labile to heat and phenol. Rabbit antisera, showing only precipitin to component II or certain human sera at high dilution, were found to react with G cells to give an immunofluorescence which was confined to the germ tubes. This suggested that component II is localised on the germ tubes, whereas no immunofluorescent reaction against the yeast cells could be detected under the same conditions although component II was as readily extracted from these cells as from G cells. This suggested that component II might exist in a cryptic state in the Y cells. In support of the latter contention it was shown that live Y cells did not absorb precipitin to component II nor were they capable of providing these antibodies in rabbits. Using both human and rabbit sera, it was shown that the antigenic specificity of the immunofluorescence assay where Y cells were used was related to component I and that where G cells were used it was related to both components I and II.

Adolescent↗
PubMed1982

Protein contents and binding modes of immunoglobulin-amylase complexes.

The protein contents and binding modes of immunoglobulin-amylase complexes were investigated. Amylase staining after immunoelectrophoresis permitted identification of IgA-lambda type in three cases, IgA-kappa type in two cases and IgG-kappa type in one case. The precipitin line of Fab fragment of IgA in four patient's sera after papain digestion was found to have amylase activity. Sucrose density gradient ultracentrifugation studies showed that these complexes were divided into three groups: Group I complexes were dissociated in acid pH, and at pH 7.0 the immunoglobulins, which were isolated at the acid pH, became rebound to serum amylase; Group II complexes were dissociated by acidification, but did not reform at pH 7.0; Group III complexes were unaffected by acidification. We suggest that the immunoglobulin-amylase complexes belonging to groups I and II result from an antigen-antibody reaction. Precipitin curves resulting from the addition of increasing quantities of the patient's serum containing 9 S or 11 S immunoglobulin-amylase complexes to specific antisera revealed that these complexes contained not only monoclonal IgA, but also alpha-1-antitrypsin (alpha 1 AT) and albumin. These IgA complexes could have bound non-specifically to alpha 1 AT and albumin, forming the larger macroamylase complexes.

Aged↗
PubMed1986

Relationships between radiographic change, pulmonary function, and bronchoalveolar lavage fluid lymphocytes in farmer's lung disease.

Ninety four dairy farmers were investigated by chest radiography, pulmonary function tests, and bronchoalveolar lavage. They were divided into five groups--1: 11 subjects with acute farmer's lung; 2: 25 subjects with previously diagnosed farmer's lung who had stayed on their farm; 3: 15 farmers with previously diagnosed farmer's lung who had left the farm; 4: 23 precipitin positive symptomless farmers; 5: 20 precipitin negative symptomless farmers. The study evaluated the relationships between radiographic changes measured with a scoring system derived from the International Labour Office (ILO) classification, the results of pulmonary function tests, and bronchoalveolar lavage fluid. Thirty eight subjects had radiographic evidence of interstitial pulmonary infiltrates. Group 1 had the highest percentage of lymphocytes recovered by bronchoalveolar lavage (mean 66.3 (SD 19.2]. For all subjects carbon monoxide transfer factor (TLCO) and total lung capacity were negatively correlated with radiographic changes (r = -0.45 and -0.30; p less than 0.001 and less than 0.01 respectively). TLCO was also negatively correlated with radiographic change in group 2 (r = -0.59, p less than 0.005). The percentage of lavage lymphocytes was correlated with radiographic changes for all subjects (r = 0.36, p less than 0.001), but this correlation was not seen within groups. This study shows good correlation between radiographic abnormalities, pulmonary function changes and the cellular composition of bronchoalveolar lavage fluid.

Adolescent↗
PubMed1978

Susceptibility of chicks to infectious bursal disease (IBD) following vaccination of their parents with live IBD vaccine.

Vaccination of parent chickens with a commercial live infectious bursal disease (IBD) vaccine under field conditions at varying ages and by different routes resulted in variable susceptibility to the disease in their chicks. There was little correlation between the methods of vaccination and the levels of immunity in the chicks. There was some evidence that levels of transferred immunity decreased with the age of the parents. Of five flocks examined, the onset of susceptibility to IBD occurred at two days of age in two flocks and about one week of age in the other three. Four flocks were completely susceptible by the 27th day, the other at 31 days. Precipitins were demonstrated at day old in 100 per cent of chicks in three flocks and in 80 per cent in the other two. The precipitins had disappeared in all flocks by 17 days of age.

Age Factors↗
PubMed1990

Isolation and immunochemical characterization of fractions from membranes of Aspergillus fumigatus with protease activity.

Two fractions exhibiting acid protease activity (AFPI and AFPII) were isolated by extraction of membrane vesicles of Aspergillus fumigatus with Triton X-100. These two fractions produced single bands in both polyacrylamide and sodium dodecyl sulfate polyacrylamide gel electrophoresis and showed apparent molecular weights of 73,000 and 43,000, respectively. Molecular weights determined by gel filtration in the absence and presence of Triton X-100 and sedimentation velocities in analytical ultracentrifugation indicated hydrophobic characteristics, since both fractions readily aggregated and complexed with Triton X-100; both exhibited elevated enzyme activities in the presence of Triton X-100. Carbohydrate content was 93% for AFPI and 85% for AFPII. The enzymatic fractions demonstrated different pH optima in the acid range as well as different temperature stabilities. Both protease fractions cross reacted in double immunodiffusion, while in crossed immunoelectrophoresis both demonstrated five precipitin peaks, each with similar patterns. AFPI demonstrated two additional precipitin peaks in crossed immunoelectrophoresis. As determined by crossed immunoaffinoelectrophoresis, the protease fractions demonstrated galactose and mannose residues. In biotin-avidin enzyme-linked immunosorbent assay both fractions reacted with allergic bronchopulmonary aspergillosis and aspergilloma sera. It can be concluded that two fractions with protease activity of A. fumigatus reported here may be of significance in Aspergillus-induced diseases.

Aspergillosis, Allergic Bronchopulmonary↗
PubMed2001

Longitudinal changes of sensitization to farming-related antigens among young farmers.

BACKGROUND: We have shown previously that the prevalence of allergic sensitization in Tyrolean farming students is high, with a clear relationship to occupational exposure. OBJECTIVES: It was the aim of this investigation to assess longitudinal changes in sensitization and a possible impact on lung function. METHODS: Of the 147 farming students in the original cohort, we could re-examine 42 after a mean interval of 4.0 years. All individuals completed a questionnaire and had spirometry, skin prick tests, total and specific IgE analysis, and testing for precipitating antibodies. RESULTS: As compared to the first study, there was a decrease in the frequency of IgE-mediated allergy (30.1 vs. 54.8%; p < 0.05) and in total serum IgE levels (107.4 +/- 254 vs. 157.8 +/- 304 U/ml; p < 0.001). Conversely, 3 individuals had developed precipitating antibodies de novo, and those who had initially had a positive precipitin test (n = 6) were still positive. A larger-sized estate, the lack of a hay dryer, and the presence of moldy hay were confirmed as risk factors for allergic sensitization. Although there was a slight overall decrease in forced vital capacity, no new cases of occupational lung disease were found. CONCLUSIONS: In young Tyrolean dairy farmers, the prevalence of precipitins is constantly high and rising while IgE-mediated allergy declines. Within 4 years, no clinically relevant impact of allergy on lung function was noticed.

Adolescent↗
PubMed1980

Application of single radial immunodiffusion for qualitative comparison of plant virus particles.

The three fundamental types of precipitin patterns obtained with plant viruses by double radial immunodiffusion (DRID) tests can also be obtained by single radial immunodiffusion (SRID) tests. Using virus antigens that were serologically identical or unrelated, the reaction patterns were readily recognized. However, the precipitin reaction corresponding to the spur in DRID tests using virus antigens that were partially identical serologically was difficult to distinguish. It seems, therefore, that when virus isolates are serologically closely related, DRID should be preferred to SRID. Different viruses in a mixed infection can be successfully distinguished by SRID and also by 'rocket' immunoelectrophoresis.

Antigens, Viral↗
PubMed1988

Isolation and purification of erythropoiesis inhibitory activity from uremic sera.

In vitro erythropoiesis inhibitory activity (EIA) was isolated and partially purified from sera obtained from a patient with chronic renal failure (CRF). EIA was purified from human sera by ammonium sulfate precipitation, column chromatography and PAGE electrophoresis, and the corresponding biological activity was assayed in erythroid colony (CFU-E) cultures. Ammonium sulfate precipitation revealed that the EIA resided in two high molecular weight fractions which yielded a single peak on G-200 Sephadex. PAGE electrophoresis demonstrated that the EIA fraction separated into 3 bands with a molecular weight range of 12,000-50,000 daltons. Antibody to EIA was prepared in rabbits, and Ouchterlony immunodiffusion assays employing the antibody material revealed that several CRF sera generated intense double precipitin bands when tested in the assay. In contrast, normal sera consistently yielded a weak precipitin band. These results suggest the presence of reduced amounts of EIA-like material in normal humans and elevated levels in CRF sera. The significance of EIA in the anemia of CRF is discussed.

Aged↗