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PubMed1984

A two-site sandwich radioimmunoassay of beta 2-microglobulin with monoclonal antibodies.

Among 21 batches of monoclonal antibodies raised against beta 2-microglobulin (anti-beta 2m), 6 reacted with soluble beta 2m, as well as with beta 2m present in association with HLA heavy chains on the surface of leukocytes. The remaining 15 anti-beta 2m antibodies bound with soluble beta 2m, but failed to react with beta 2m on the cell surface. No monoclonal anti-beta 2m antibodies revealed precipitin lines when they were tested against beta 2m in immunodiffusion. When 2 anti-beta 2m antibodies with different specificities were mixed together, however, they developed a precipitin line against beta 2m. Based on these observations, there was only 1 epitope on beta 2m that was available for the binding with a monoclonal anti-beta 2m antibody with either specificity. This allowed the development of a solid-phase radioimmunoassay in which beta 2m in test specimens was sandwiched between immobilized anti-beta 2m of 1 specificity and radiolabeled anti-beta 2m of a heterologous specificity in a single step. Monoclonal antibodies with at least 2 different specificities would be required for developing a sandwich-type immunoassay of polypeptides, such as beta 2m, that do not display 2 or more epitopes of the same specificity.

Animals↗
PubMed1984

Antigen and antibody attachment in ELISA for Aspergillus fumigatus IgG antibodies.

Four commercial antigen extracts of Aspergillus fumigatus were evaluated for use in a rapid enzyme-linked immunosorbent assay (ELISA) for anti-A. fumigatus IgG. Initial binding of both somatic and culture filtrate preparations to a polyvinyl chloride solid phase was concentration dependent and increased with incubation time. Antigen binding to the solid phase was reproducible. Binding of A. fumigatus precipitin-positive serum to bound antigen was rapid. All four A. fumigatus antigens demonstrated similar dose-response curves when tested against pooled sera containing a high titre of A. fumigatus antibodies. Detectable activity in precipitin test-negative sera decreased rapidly with dilution. All the antigen preparations were found to be suitable for ELISA procedures and permit the rapid determination of IgG antibodies to A. fumigatus.

Alkaline Phosphatase↗
PubMed1982

Litomosoides carinii infection in cotton-rats before and after treatment with diethylcarbamazine and suramin: evolution of free serum antigens.

Immunoelectrodiffusion was used to detect free serum antigens after treatment with diethylcarbamazine (DEC) and Suramin (S) in 30 cotton-rats infected with Litomosoides carinii. Precipitin lines between an antiserum produced by an adult worm extract and the test serum indicated the presence of free antigens in the serum. The number of precipitin lines was between one and 3. Before treatment only 13.3% of infected rats were positive compared with 83.7% after treatment. There was a difference in the delay before the appearance, the persistence and, possibly, the nature of antigens liberated by DEC and S treatment. With DEC release occurred earlier than with S and the antigens persisted for a shorter time. With combined DEC + S therapy the results were similar to those obtained with DEC and S used alone.

Animals↗
PubMedDate not supplied

Early diagnosis of human fascioliasis by the detection of copro-antigens using counterimmunoelectrophoresis.

We have reported the use of partially purified Fasciola gigantica worm antigens (peaks II and III separated by Sephadex G-200) in counterimmunoelectrophoresis (CIEP) for the specific and early detection of antibodies in sera of fascioliasis patients. In the present study, we report the use of hyperimmune serum from rabbits immunized with these partially purified worm antigens in CIEP for the detection of parasite antigens in saline extracts of patients' stools. This test was conducted on stool specimens from 144 patients with systemic or intestinal parasitic infections, 5 from persons with spurious Fasciola infections, and 10 from healthy negative controls. All saline extracts from fascioliasis patients developed 2-5 precipitin bands. No precipitin band developed with stool extracts of patients with spurious Fasciola infection or other parasitic infections, or from negative controls. This assay proved to be simple, rapid, sensitive, and specific for the diagnosis of early as well as established fascioliasis infections.

Animals↗
PubMed1984

Classification of Agkistrodon species in China.

The wide geographical distribution of Agkistrodon and the slight morphological differences among the snakes of the genus Agkistrodon in China have posed a problem to taxonomists. We have employed polyacrylamide gel electrophoresis and immunological diffusion techniques for comparison of the venoms of different species and subspecies of Agkistrodon from various localities. The electrophoretic patterns of the proteins of the venoms were different from each other, but showed certain relations within species and subspecies. We used Ouchterlony double diffusion of a rabbit antiserum against the purified "neurotoxin" from the venom of Agkistrodon blomhoffii brevicaudaus (from the Zhejiang Province of China) on the various venoms of Agkistrodon. Precipitin lines formed with immunological identity between the same species, partial identity between closely related species and no precipitin line between different species. Combining experimental data, morphological characteristics and geographical distribution, we propose that the genus Agkistrodon (sensu stricto) in China consists of seven species and subspecies: (1) Agkistrodon blomhoffii brevicaudus Stejneger, (2) A. b ussuriensis Emelianov, (3) A. intermedius (Strauch), (4) A. saxatilis Emelianov, (5) A. shedaoensis Zhao, (6) A. strauchii Bedriaga, (7) A. monticola Werner. Agkistrodon acutus (Guenther) has recently been changed to a new genus, Deinagkitrodon, established by Gloyd in 1978.

Animals↗
PubMed1987

Observations on white and yellow venoms from an individual southern Pacific rattlesnake (Crotalus viridis helleri).

Biochemical differences in white and yellow venoms produced in the separate venom glands of an individual southern Pacific rattlesnake (Crotalus viridis helleri) were investigated. Compared to the yellow venom, the white venom contained fewer low molecular weight components and was considerably less toxic. Although the exact LD50 was not determined, the white venom did not produce toxic effects in mice when injected i.v. at concentrations up to 10 mg/kg. The i.v. LD50 of the yellow venom was approximately 1.6 mg/kg. Both white and yellow venoms had hemorrhagic activity, but the white venom caused less intradermal hemorrhage in mice. No L-amino acid oxidase activity was measured in the white venom and protease and phospholipase A2 activities of the white venom were much less than in the yellow venom. The white and yellow venoms both produced myonecrosis at 1, 3 and 24 hr after i.m. injection into mice, however, there were some qualitative differences in the myonecrosis produced. When the venom samples were reacted against Wyeth's polyvalent (Crotalidae) antivenom using immunodiffusion, three precipitin bands formed against the yellow venom, whereas only one formed against the white venom. When reacted against an antiserum to myotoxin alpha from C. viridis viridis venom, both the white and yellow venoms produced one precipitin band each.

Amino Acid Oxidoreductases↗
PubMed1988

Toxins in the skin secretion of the oriental catfish (Plotosus lineatus): immunological properties and immunocytochemical identification of producing cells.

Antiserum against toxin I, one of the lethal factors in the skin secretion from the oriental catfish (Plotosus lineatus), was used to examine immunological properties of the toxic factors and identify toxin-producing cells by an immunocytochemical technique. In immunodiffusion tests, the antiserum formed a precipitin line with toxin I while it formed no precipitin line either with another lethal factor (toxin II) or with a hemolysin. Lethal and edema-forming activities of toxin I were neutralized by the antiserum but lethal activity of toxin II and lytic activity of the hemolysin were not. These results suggested that toxin I can be antigenically distinguished from both toxin II and hemolysin. In immunocytochemical tests using the antiserum, club cells in the epidermis were positively stained, indicating that toxin I is produced in the club cells. Interestingly, venom glandular cells surrounding the dorsal and pectoral spines were also stained. The venom glandular cells appear to produce toxin I or a toxin with the same antigen determinants as toxin I.

Animals↗
PubMed1983

Serologic cross-reactions among streptococcal group A, A-variant, and C polysaccharides.

Serologic cross-reactions among streptococcal groups A, A-variant (A-V), and C cell wall polysaccharides were found previously in studies employing capillary or quantitative precipitin techniques. Similar cross-reactions occur with radioimmune precipitation using extrinsically labeled 125I-streptococcal antigens. This study was performed to determine the degree of cross-reactivity when intrinsically labeled 14C-polysaccharide antigens were used in the radioimmune precipitin assay. Unadsorbed antisera from rabbits immunized with group A streptococci bound 3-5% as much 14C-A-V antigen as homologous A carbohydrate but undetectable amounts of C polysaccharide. Similarly, A-V antisera bound 3-5% as much 14C-A or 14C-C carbohydrate as A-V antigen. Group C antiserum bound 1-2% as much A and A-V antigens as C carbohydrate. Thus, less than 3% of intrinsically labelled 14C-A or 14C-C carbohydrate represents exposed A-V antigenic sites, i.e., exposed polyrhamnose backbone. Otherwise, group A, C, or A-V carbohydrates failed to exhibit heterologous determinants. Anti-peptidoglycan antibodies in antisera did not result in serologic cross-reactivity. These data suggest that formamide-extracted streptococcal group-specific polysaccharides, intrinsically labeled with 14C, may possess greater group specificity than 125I-carbohydrates and yield only negligible cross-reactivity with heterologous antisera. This degree of cross-reactivity does not appear to be sufficient to account for the persistently elevated serum levels of antibody to group A carbohydrate in patients with rheumatic heart disease.

Acetylgalactosamine↗
PubMed1975

Comparison of honeybee venoms and their components from various sources.

The use of honeybee venoms and their components may assist in the elucidation of the pathophysiology of reactions to honeybee stings. This initial study compared venoms from various sources by chemical and biological assays, and significant variations were observed. Ten different bee venoms were compared by nitrogen analysis, mouse toxicity, hyaluronidase content, and antigenicity. Based on mouse toxicity, hyaluronidase content, and gel diffusion analysis, two groups of bee venoms could be differentiated. Venoms in one group, Group A, were more toxic, contained hyaluronidase, and showed an additional precipitin band. All venoms contained mellitin as a major fraction, which formed nonimmune precipitin bands during gel diffusion analysis. Gel filtration chromatography and dialysis separated the venoms into components that were then identified by enzyme assays, rat mast cell degranulation, hemolytic activity, and gel diffusion analysis. The venoms within Group A showed similar components, some of which, most noticeably hyaluronidase, were not present in Group B. Dialysis showed that a large portion of the venom could pass through a cellophane membrane including a portion of the phospholipase A. Heterogeneous molecular weights were found for phospholipase A by both gel filtration and dialysis, and may reflect variation in carbohydrate content. It appears that bee venom variability for whatever reason, a heterogeneous MW antigen, and a non-immune precipitable component require careful consideration in any study involving this venomm. These studies have yielded relatively pure, identified bee venom components which can be employed in further studies investigating reactions to honeybee stings.

Animals↗
PubMed1978

Humoral and cellular immune responses in Aspergillus fumigatus pulmonary disease.

This study was designed to evaluate immunologic differences between aspergilloma (A) and allergic bronchopulmonary aspergillosis (ABPA), comparing the results to atopic and nonatopic control subjects. Humoral studies included skin tests with common inhalant antigens and Aspergillus fumigatus. Total and specific IgE and other immunologlobulin levels and serum precipitins were evaluated against A. fumigatus. Cellular immunity was studied with routine skin testing and phytohemaqglutinin-induced lymphocyte blast transformation. Antigen-induced blast transformation was also carried out with the use of serial dilutions of A. fumigatus. All patients with ABPA were atopic and had marked elevations of IgE. None of the patients were atopic and they had normal IgE levels. Immediate and late skin reactivity to A. fumigatus was low in control and A groups but high in 2 patients with ABPA. IgG antibody against A. fumigatus was generally greater in the ABPA group. Both ABPA and A had serum precipitating antibody against A. fumigatus. The atopic controls had elevated IgE levels and immediate skin test reactivity to A. fumigatus, and one also had weak serum precipitins against A. fumigatus. IgE antibody against A. fumigatus was generally higher in ABPA than A. ABPA and A patients had elevated stimulation indices (SI) to A. fumigatus. No stimulation could be detected with cells from control subjects. This study indicates that both T and B cell sensitization may play a role in the development of or as a response to aspergillus-related pulmonary disease.

Antibody Formation↗
PubMed1982

Grain fever syndrome induced by inhalation of airborne grain dust.

To study the clinical and physiologic manifestations of the grain fever syndrome and the potentially pathogenic role of complement activation, 12 subjects (six grain workers and six healthy non-grain workers) underwent inhalation provocations with airborne grain dust. The clinical response was characterized by facial warmth, headache, malaise, myalgias, feverish sensation, chilliness, throat and tracheal burning sensation, chest tightness, dyspnea, cough, and expectoration. Fever developed in four grain workers and two controls. Leukocytosis, ranging between 11,700 and 24,300 leukocytes/mm3 with left shift, developed in five grain workers and five controls. There was no evidence of complement activation by the classical or alternate pathway. None of the subjects had serum precipitins to grain dust. The pulmonary response was characterized by a decrease in FEV1, FVC, MMF, Vmax50, and Vmax75, with significant rise in pulmonary resistance and consistent change in dynamic compliance but without changes in static compliance or diffusing capacity. Hence, grain dust inhalation induced diffuse airways obstruction without detectable parenchymal reaction. The airways response to high concentrations of grain dust inhalation were unrelated to the presence of immediate skin hypersensitivity. Although we cannot exclude the etiopathogenetic role of an immunologic reaction to grain dust, our data do not support the hypothesis that the grain fever syndrome is a precipitin-mediated allergic pneumonitis. More likely, the manifestations of grain fever probably reflect the host reaction to grain dust bacterial endotoxins and/or nonallergic mediator release by grain or grain dust constituents.

Adult↗
PubMed1984

The lectin reactivity and lectin-like activity of allergenic pollen extracts.

The binding of soluble components of pollen grains to plant-stigma receptors can be inhibited by concanavalin A. This lectin-like activity of pollen components is important in the genetic control of plant reproduction. Aqueous extracts of allergenic pollens also react with concanavalin A. Agarose gel-diffusion precipitates were used to survey and characterize the ability of allergenic pollen extracts to react with concanavalin A and other lectins. Concanavalin A alone precipitated with extracts of plantain, American beech, white ash, and corn pollens. Surprisingly, extracts of the pollen from certain plants also precipitated when the extracts were diffused against pollen extracts from other plants. Pollen extracts of alfalfa, white ash, American beech, burweed marsh elder, redtop grass, corn, plantain, orchard grass, and aspen reacted with one or more other pollen extracts. Extract precipitin activity was reliably obtained after extracting pollens for 20 min with pH 7.5, 0.05M Tris buffer in 0.2M of saline. Optimal agarose gel conditions for detecting the precipitin reactions were pH 8.5 to 9.0, 75 mM borate buffer made to an ionic strength of 1.5M with NaCl for concanavalin A pollen reactions and 0.015M with NaCl for pollen-pollen reactions. The presence of the borate ion was necessary for optimal detection of the agarose gel precipitates. Studies of the inhibition of the lectin-pollen and pollen-pollen reactions with specific mono and disaccharides revealed many similarities and differences between the two types of reactions. The high concentrations of glycerol used to stabilize pollen extracts also inhibit these reactions.

Concanavalin A↗
PubMed1985

A familial occurrence of allergic bronchopulmonary aspergillosis: a probable environmental source.

This is the first article of a familial occurrence of allergic bronchopulmonary aspergillosis in a father and daughter. Serologic studies revealed serum precipitins and elevated IgG or IgE antibody activity against certain Aspergillus species. Investigation of the home revealed an environmental source of exposure. Another family member demonstrated serum precipitins and an IgG antibody response to the isolated Aspergillus species without evidence of allergic bronchopulmonary aspergillosis.

Adult↗
PubMed1987

Isolation and identification of pollen allergens of Artemisia scoparia.

The allergenic proteins of Artemisia scoparia pollen were separated and identified with ammonium sulfate precipitation, ion-exchange chromatography, gel filtration, and RAST-inhibition techniques. The important allergenic component Artemisia VI b that constitutes 29% of total protein in the extract was purified to homogeneity. It was found to be an acidic protein with isoelectric point 3.8 and molecular weight of 14,300. It was rich in carbohydrate, but the carbohydrate portion did not appear to be important for allergenicity. In the crossed immunoelectrophoresis reference pattern of the whole pollen extract, 37 precipitin lines could be identified on the anodic side, whereas Artemisia VI b could be observed as a single precipitin line. Immunologically, the whole pollen extract of A. scoparia demonstrated shared antigenic and allergenic determinants with Ageratum conyzoides-pollen extract. The use of fast protein liquid chromatography in partial purification of allergenic components is also discussed.

Allergens↗
PubMed1988

Crossed immunoelectrophoretic studies of whole body extracts and venom from the imported fire ant Solenopsis invicta.

Although allergic reactions occur after imported fire ant (IFA) sting, currently, only IFA whole body extract (IFAWBE) is available for diagnosis and immunotherapy of IFA-sensitive individuals. We report our crossed immunoelectrophoretic studies comparing antigenicity and allergenicity of Solenopsis invicta IFAWBE and IFA venom (IFAV). Rabbits were hyperimmunized with IFAWBE prepared from S. invicta or with IFAV obtained from S. invicta by an electric shock method. Crossed immunoelectrophoresis with anti-IFAWBE detected at least 29 precipitin lines in IFAWBE and three lines in IFAV, but none in a synthetic venom, transpiperidine. Anti-IFAV detected six precipitin lines in IFAV and five lines in IFAWBE, but no lines were detected with transpiperidine. Crossed-line immunoelectrophoresis confirmed the IFAV origin of at least three of the peaks in IFAWBE. Crossed radioimmunoelectrophoresis with 11 IFAWBE RAST-positive sera elicited radiostaining with five antigens in IFAWBE that were probably IFAV associated. One of these allergens was recognized by all sera; the other allergens were recognized by 8, 7, 5, and 4, respectively, of the 11 sera. Four of the antigens present in IFAV preparations had allergenicity. These findings indicate that IFA allergens probably originate in IFAV, but that transpiperidine, a major constituent of IFAV, does not appear to be immunogenic. The results suggest that IFAV may be a more appropriate reagent than IFAWBE for laboratory testing and for clinical diagnosis and immunotherapy of IFA-sensitive individuals.

Animals↗
PubMed1990

Allergic Bipolaris sinusitis: clinical and immunopathologic characteristics.

Allergic Aspergillus sinusitis was first reported in 1983. We present the first three cases of allergic fungal sinusitis caused by the black fungus Bipolaris spicifera. The patients were young, atopic, and immunocompetent. All three patients demonstrated pansinusitis with nasal polyps and underwent multiple surgical procedures. Pathologic features included a characteristic mucoid exudate containing eosinophils, Charcot-Leyden crystals, and fungal hyphae. In two cases there was bony erosion revealed by computed tomography scan but no histologic evidence of direct fungal invasion into the mucosa or bony trabeculae. Immunologic features, including total eosinophil count, total serum IgE, immediate and late-phase skin response to B. spicifera, serum precipitins, and specific IgE and IgG to B. spicifera, are described. B. spicifera is a previously unrecognized cause of allergic fungal sinusitis that may be an underdiagnosed disorder. This diagnosis should be considered in atopic patients with nasal polyps and pansinusitis unresponsive to conventional medical therapy. Diagnostic criteria include characteristic histologic allergic mucin, culture identification of fungus, positive immediate cutaneous reactivity to fungal extract, positive serum precipitins, and elevated specific IgE and IgG antibodies.

Adolescent↗
PubMed1991

Variability in parameters of allergic bronchopulmonary aspergillosis in patients with cystic fibrosis.

Seventy-nine patients with cystic fibrosis (CF) were evaluated and were followed in a longitudinal, prospective fashion during a 6-year period for the development of immune parameters indicating Aspergillus fumigatus (Af) sensitization and allergic bronchopulmonary aspergillosis (ABPA). Although four patients developed frank ABPA, there was considerable variability in immune parameters in non-ABPA. Twenty-four patients became skin test positive to Af with none losing skin reactivity. Twenty-five patients developed serum precipitins to Af, whereas 12 patients lost their precipitins. Of 15 patients with an elevated total serum IgE of greater than or equal to 2 SD, five demonstrated a marked decline of at least 40%. Three of 16 patients with IgE-Af became negative, whereas eight of 27 patients lost their IgG-Af. None of these patients had received corticosteroid therapy that could have accounted for the findings. Thus, patients with CF frequently lose evidence of Af sensitivity spontaneously without corticosteroid intervention. The diagnosis of ABPA in CF should not be based solely on serology and skin test results, since at any point in time, patients with CF may demonstrate variable responses to Af.

Adolescent↗