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Elevated agglutination titres in plasma of Biomphalaria glabrata exposed to Echinostoma paraensei: characterization and functional relevance of a trematode-induced response.

Production of elevated haemolymph agglutination titres by Biomphalaria glabrata following exposure to Echinostoma paraensei miracidia was investigated, to characterize this parasite-induced response and to understand its functional relevance. Both the dose of infection (1, 10 or 100 miracidia per snail) or the number of separate exposures to infection (between one and three, over a 4 or 8 day interval) were varied, and assuming a threshold dosage (10 miracidia per snail or higher) was exceeded, titres of juvenile snails peaked at 8-16 times the values for unexposed control snails, regardless of the exposure regimen. Adult snails, which are relatively refractory to infection, have slightly higher resting titres than juveniles, but exhibit only a 2- to 4-fold increase in titre following exposure. Juveniles exposed to infection but lacking demonstrable infection had lower titres than snails with confirmed infections. Exposure to infection increased heterogeneity of plasma agglutinins and provoked production of unique specificities not found in unexposed snails. However, the overall pattern of agglutination responses for snails with successfully developed parasites did not differ from those in which parasite development was unsuccessful. Agglutinating activity was inhibitable by several different monosaccharides, although plasma from infected snails was relatively unaffected by N-acetyl-glucosamine or N-acetyl-galactosamine. Wounding of snails provoked no change in plasma agglutination activity. As the highest agglutination titres were produced in snails with successfully developing parasites and agglutinin composition did not differ between snails with successful or unsuccessful parasites, the functional relevance of the response remains enigmatic. The production of unique agglutinins following exposure deserves additional study.

Agglutination↗

A comparison of commercial Mycoplasma gallisepticum antigens in the rapid serum agglutination test.

Twenty-three chickens, aged 9 weeks, were housed in contact with a similar group of chickens which were infected with M. gallisepticum. The onset and development of the serum agglutinin responses which were detected by 3 commercial antigens using rapid serum agglutination tests up to 20 weeks of age, differed greatly both between the infected and in-contact groups and, within the latter group, between the antigens. Few reactions, which were weak and transient, were detected in 2 similar groups of chickens, neither infected with M. gallisepticum but 1 vaccinated at 12 and 17 weeks of age with killed Pasteurella multocida oil adjuvant vaccine.

Agglutination Tests↗

Identification and visualization of the sexual agglutinin from the mating-type plus flagellar membrane of Chlamydomonas.

Sexual agglutinins located on the flagellar membranes of Chlamydomonas gametes mediate a mating-type-specific adhesion reaction that brings complementary gametes together for zygotic cell fusion. We identify the mating-type plus agglutinin, using a combination of biochemical and genetic analysis, as a glycopolypeptide with an apparent molecular weight of greater than 10(6) by SDS-polyacrylamide gel electrophoresis. Its core polypeptide migrates as a approximately 480-kd species, and it is estimated to be present in approximately 30 copies per gametic flagellum. The agglutinin is present in the wild type, in a mutant that agglutinates but cannot fuse, and in a complementing diploid, whereas it is absent from four nonagglutinating mutants and from a noncomplementing diploid. Electron microscopy shows the purified agglutinin to be a highly asymmetric molecule, 220 X 4 nm. To our knowledge, this is the first reported purification and visualization of a membrane-associated cell-cell recognition protein.

Agglutinins↗

Expression of lectin receptors on the surface of granulocyte-macrophage progenitor cells (CFU-gm).

It has been emphasized that specific bindings between membrane glycoproteins and membrane lectin-like substances are important in cell-to-cell interactions. We explored the surface of granulocyte-macrophage precursor cells (CFU-gm) by the differential agglutination technique. Enrichment of CFU-gm in the agglutinated fraction, containing the cells which have lectin receptors, from marrow treated with soybean agglutinin (SBA), peanut agglutinin (PNA) and concanavalin A (Con A), suggests the presence of reactive galactosyl and mannosyl residues on the surface of CFU-gm. On the other hand, wheat germ agglutinin (WGA), phytohemagglutinin (PHA) and ulex europaeus agglutinin (UEA), which bind to reactive N-acetylglucosamine, N-acetylgalactosamine and fucose, respectively, did not specifically agglutinate CFU-gm. Thus, reactive groups containing galactosyl and mannosyl structures on the surface of CFU-gm may possibly play a role in the process of cell-to-cell interactions between CFU-gm and marrow stromal cells.

Animals↗

The role of carboxylic acids in EDTA-dependent panagglutination.

The investigation of a patient blood sample showing a discrepancy between cell grouping and serum confirmation demonstrated a serum agglutinin which reacted with all red blood cells tested when exposed to EDTA. This reaction was 4+ macroscopic at room temperature, 2+ macroscopic with hemolysis at 37 degrees C in albumin, and 1+ macroscopic in the anti-human globulin phase. Agglutination was abolished following dithiothreitol treatment of the patient's serum or following saline washing of the EDTA-exposed test cells. The agglutination reaction was not limited to EDTA, but could be produced with polycarboxylic acids (citrate, L-tartrate, succinate) and monocarboxylic acids (acetate, lactate, propionate, valerate, butyrate). Non-carboxylic acids and low molecular weight ketones or alcohols failed in the agglutination reaction. This study reports an additional example of an IgM "EDTA dependent agglutinin" and demonstrates the dependence on carboxyl groups for its agglutinating activity.

ABO Blood-Group System↗

Lectin separation of nonlymphoid suppressor cells induced by total lymphoid irradiation.

Suppression of the mixed lymphocyte reaction (MLR) exerted by splenocytes derived from mice treated with fractionated total lymphoid irradiation (TLI, 200 rds x 8) was analyzed by various criteria in order to characterize the phenotype of the cell type(s) responsible for suppression. TLI-induced suppressor cells could not be eliminated by removal of cells bearing surface immunoglobulin, Thy-1, Lyt-2 and TL, and thus could not be ascribed to lymphocytes of the B or T cell lineage. Suppressor cells were large, and nonadherent to nylon wool, Sephadex G-10 and plastic surfaces. Suppressor activity of TLI splenocytes was predominantly located in fractions of cells bearing receptors for soybean agglutinin (SBA), peanut agglutinin (PNA) or both lectins. SBA+, PNA+, sequentially agglutinated (SBA followed by PNA) SBA+PNA+ and (PNA followed by SBA) PNA+SBA+ suppressor cells were radioresistant upon exposure to 1000 rds in vitro. Cells bearing the receptor for PNA but lacking that for SBA (PNA+SBA-) had sharply reduced suppressor activity. However, a radiosensitive PNA- suppressor cell subset was also documented in the spleen of TLI-treated mice. Thus, suppressor cells could best be physically separated from nonsuppressors by the SBA lectin. SBA+ suppressor cells were found, by scatter analysis, to include the population of large cells characteristic of TLI splenocytes, whereas SBA- cells were much smaller and almost exclusively devoid of suppressive capacity. The PNA receptor was found to further dissect the SBA+ suppressor cells into two distinct subpopulations: radioresistant SBA+PNA+ cells and radiosensitive SBA+PNA- cells. In summary, we suggest here the presence of at least two suppressive populations induced by TLI: radioresistant SBA+, PNA+, SBA+PNA+ or PNA+SBA+ cells, and radiosensitive PNA- and SBA+PNA- cells. Similar subsets of MLR suppressor cells can be isolated from normal bone marrow cells and splenocytes of nude mice, suggesting that suppression is mediated by large, immature, nonlymphoid cells which might migrate from shielded bone marrow compartments into the spleen of TLI-treated mice.

Animals↗

A new cold agglutinin from Achatina fulica snails.

An electrophoretically homogeneous agglutinin was purified from the albumin gland of Achatina fulica snails using asialofetunin-Sepharose 4B as an affinity column. The agglutinin was found to be temperature sensitive; it agglutinated rabbit and human umbilical cord erythrocytes only at low temperature. It was found to be specific for methyl-beta-D-galactoside, and the best inhibitor was N-acetyllactosamine.

Animals↗

Establishment of assay kits for the determination of microheterogeneities of alpha-fetoprotein using lectin-affinity electrophoresis.

Diagnostic kits for determination of alpha-fetoprotein (AFP) carbohydrate chain microheterogeneity were developed using lectin affinity electrophoresis with Lens culinaris agglutinin-A (LCA-A) and erythro-agglutinating phytohemagglutinin-E4 (PHA-E4). Separated AFP bands by electrophoresis were detected with high sensitivity by antibody-affinity blotting and immunoenzymatic amplification. Densitometry was used to apportion lectin reactive AFPs. The within-run S.D. for proportions of AFP bands was below 3%. Band intensity was linearly related to AFP concentration between 2 and 200 ng/ml. Profiles of lectin reactive AFPs were compared in serum samples from 55 patients having liver diseases. The average values of lectin reactive AFPs for chronic hepatitis and liver cirrhosis patients were both below 13%, but those of hepatocellular carcinoma patients were above 25%. Correlation of data with disease states suggests that the methods can greatly facilitate the discrimination between benign and malignant liver diseases.

Carcinoma, Hepatocellular↗

Amines protect in vitro the celiac small intestine from the damaging activity of gliadin peptides.

Proteins and peptides responsible for the celiac small intestinal lesion inhibit both the enterocyte recovery of in vitro cultured flat celiac mucosa and the in vitro development of fetal rat intestine. They also agglutinate K 562 (S) cells. Using these three in vitro systems (cultured human celiac and rat fetal intestine and cell agglutination), it is shown that several small-molecular-weight amines, mostly the polyamines spermidine and spermine, prevent and reverse K 562 (S) cell agglutination induced by gliadin peptides, whereas they do not prevent cell agglutination induced by concanavalin A and wheat germ agglutinin. Some of these amines also protected in vitro developing fetal rat intestine and flat celiac mucosa from the damaging effect of gliadin peptides. This protective effect may be related to the trophic activity exerted by amines on the intestine and/or the effect of amines on the functions of intestinal brush border or intracellular membranes involved in the intestinal handling of gliadins.

Abdomen↗

Examination of the immunoglobulin classes involved in the serological response of pregnant sheep to Aspergillus fumigatus.

Pregnant sheep inoculated with Aspergillus fumigatus conidia developed agglutinating and precipitating antibodies for mycelial antigens. The agglutinins were initially exclusively of the IgM class, but were later supplemented by IgG antibodies, although IgM production was usually sustained throughout the serological response. Precipitins active in the immunodiffusion test were of the IgG class. They developed later in the immune response than agglutinins and declined more rapidly. The precipitins and IgG agglutinins were more closely associated with recent active infection than IgM agglutinins.

Animals↗

Lectin binding as a biological test in vitro for the prediction of functional activity of human spermatozoa.

Adequate acrosome reaction is one of the essential events that has to occur in successful mammalian fertilization. The purpose of the present study was to assess the acrosome reaction in human spermatozoa by means of iodine-labelled lectins concanavalin A and peanut agglutinin ([125I]Con A and [125I]PNA). Six sperm samples in the control (fertile) group were compared with 24 samples obtained from infertile patients. The acrosome reaction in both groups was induced in vitro by adding follicular fluid. Iodine-labelled lectins were bound to the sperm surface, and alteration in the binding capacity for [125I]PNA and [125I]Con A after induced acrosomal reaction was the main parameter for the prediction of acrosome reaction and fertilizing ability. It is hoped that with the availability of this test, in-vivo data may be accumulated.

Acrosome↗

Interactions between peroxide and salivary glycoprotein: protection by peroxidase.

The effect of ascorbic acid oxidation and of peroxide on salivary glycoproteins has been studied. No signs of depolymerization of the glycoproteins were observed as a result of treatment of whole saliva or the isolated glycoprotein fraction from human saliva with an ascorbic acid, hydrogen peroxide and cupric ion combination. Sialic acid was analyzed with two methods which gave different chromogens of the acid. The conclusion could be drawn from the results that the effect of ascorbic acid oxidation was a decarboxylation of sialic acid in salivary glycoproteins. This also destroys the bacteria-agglutinating activity of saliva. Full protection of the agglutinin activity is given by salivary peroxidase. It is suggested that one role of peroxidases in saliva is to protect biologically active proteins from the action of oxygen or its radicals.

Ascorbic Acid↗

Immune electron microscopy of cross-reactions between Mycoplasma pneumoniae and human erythrocytes.

Respiratory infection with Mycoplasma pneumoniae evokes immunoglobulin M autoantibody which agglutinates human erythrocytes at 4 degrees C (cold agglutinin) and is specific for I antigen. Cross-reactions between surface antigens of M. pneumoniae and human erythrocytes, previously examined by serological analysis, were examined by transmission and scanning electron microscopy. Ferritin-labeled human antimycoplasmal and rabbit antisera to erythrocyte membrane components reacted with antigens on the surface of both M. pneumoniae and erythrocytes. Adsorption of human erythrocytes to M. pneumoniae was blocked by the same antisera without ferritin label. It is proposed that the cross-reactive specificity lies in peripheral areas of the mycoplasmal cell, probably in a surface carbohydrate which has antigenic identity with erythrocyte glycoprotein.

Antigens, Surface↗

[Temperature-dependent pseudoleukocytopenia and in vitro studies on the underlying mechanism].

Spuriously low electronic white-cell count was obtained on EDTA-anticoagulated blood from a 54-year-old male suffering from liver cell carcinoma. In an attempt to understand better the phenomenon, in vitro studies were performed. The blood smear anticoagulated with EDTA revealed remarkable agglutinations of leukocytes, mainly neutrophils but not lymphocytes. The EDTA-treated blood showed maximal leukocytes-agglutinations at room temperature but no agglutination at 37 degrees C. The agglutinated leukocytes, moreover, were dispersed by incubating the blood at 37 degrees C. Although agglutination occurred in some degree in the blood anticoagulated with Na-heparin or Na3-citrate, it was much less pronounced compared to the EDTA-treated blood. Remarkable leukocytes-agglutinations were induced when EDTA-anticoagulated blood from healthy donor was mixed with serum from the patient at room temperature. These results suggest that the phenomenon of in vitro leukocyte agglutination and consequent pseudoleukocytopenia is due to leukocyte agglutinin in serum from the patient. The mechanisms were discussed comparing with pseudothrombocytopenia caused by platelet agglutinin.

Adolescent↗

The serological specificities of Salmonella typhi antigens.

Sera prepared with two different strains of Salmonella typhi were analysed against all the soluble antigens isolated from S. typhi 0901, S. typhi Ty2 and S. typhi Vi. Agar-gel diffusion against individual sera showed that, in all the sera, antibodies were induced against somatic antigens and free proteins. Absorptions of the sera with polysaccharides, split from the somatic antigens, removed the antibodies induced against the polysaccharide and its proteinic carrier in most of the somatic antigens of S. typhi 0901. The antibodies left in the absorbed sera reacted against the proteinic moieties of more complex somatic antigens of S. typhi and against free proteins from all the analysed strains. Only the absorption with proteins removed all the precipitating antibodies from the sera. Moreover, in incomplete absorptions with proteins, the first antibodies removed are the antipolysaccharides, since antibodies are never induced against the haptenic polysaccharide but against somatic conjugates; in these the proteinic moiety eventually varies with every batch of bacteria. The sera exhausted of precipitins still agglutinate the bacteria, thus confirming the assumption that agglutinins and precipitins may be different antibodies.

Animals↗

Characterization of a rat salivary sialoglycoprotein complex which agglutinates Streptococcus mutans.

Rat saliva agglutinated Streptococcus mutans Ingbritt and NCTC 10449 and Streptococcus sanguis NCTC 7864 but not S. mutans NCTC 10921, GS 5, or LM 7, Streptococcus sobrinus 6715-13 or OMZ 65, or Streptococcus cricetus HS 6, as measured turbidometrically. The specificity of agglutination by rat saliva was the same as that by human saliva. Agglutination was associated with a mucin complex (rat salivary agglutinin complex [rS-A]) of sulfated sialoglycoproteins, with a trace of associated lipid and an apparent Mr of 1.6 X 10(6), isolated by gel-filtration Fast Protein Liquid Chromatography. The complex was dissociated in a high-ionic-strength buffer containing 6 M urea and then fractionated by gel filtration and anion-exchange Fast Protein Liquid Chromatography into four sulfated sialoglycoprotein components, designated rS-A-1Q1, rS-A-1Q2, rS-A-1Q3, and rS-A-2, with rS-A-1Q2 being polydisperse through differential glycosylation of the polypeptide backbone. The dissociation destroyed agglutination activity. The polypeptide backbones contained up to 42% serine plus threonine and up to 40% glycine plus alanine plus proline plus valine. The carbohydrate moiety of the rS-A sialoglycoproteins consisted of N-acetylgalactosamine, sialate, galactose, fucose, N-acetylglucosamine, and small amounts of mannose, with the predominant sugar being N-acetylgalactosamine. Agglutination was inhibited by 1 mM EDTA but was restored by 1.5 mM CaCl2. Agglutination was also inhibited by 5 mM CaCl2; nonimmune sera; cationic polymers; and wheat germ, lentil, soybean, and peanut lectins. However, agglutination was not affected by lipoteichoic acid, various anionic proteins, or various sugars. Neuraminidase treatment of rS-A did not affect activity, but tryptic digestion of S. mutans did prevent agglutination. The results are consistent with calcium bridging the negative groups within the rS-A complex and allowing the approach of rS-A to the bacterial cell surface to effect a specific conformational attachment.

Agglutinins↗

Nucleotide sequence analysis of the V regions of two IgM cold agglutinins. Evidence that the VH4-21 gene segment is responsible for the major cross-reactive idiotype.

Cold agglutinins are human autoantibodies, usually of the IgM class, which agglutinate RBC at low temperature. The major subset recognizes the I/i carbohydrate Ag, and many of these antibodies bear cross-reacting idiotypic determinants. An anti-idiotypic mAb that is specific for one of the idiotopes largely confined to cold agglutinins has been used to identify and monitor tumor cells that secrete these molecules in two patients. The tumor cells were immortalized with EBV and the idiotope-positive lines used to investigate the utilization of the VH and VL genes by these antibodies. Nucleotide sequence analysis of the two cold agglutinins (FS-1 and FS-2) revealed the utilization of a single common gene segment, VH4-21. Serologic analysis documented that only human antibodies utilizing the VH4-21 gene segment were reactive in the idiotope assay, other VHIV antibodies as well as a panel of antibodies derived from other VH families being negative. The DH, JH, VK, and JK gene segments of FS-1 and FS-2 were structurally distinct. These data suggest that the structural basis for the cross-reactive idiotope as well as cold agglutinin activity is the VH4-21 gene segment. A nucleotide change in H chain CDR1 of both cold agglutinins results in the substitution of an aspartic acid residue for glycine at position 31, suggesting that this amino acid might be critical to recognition of the red cell Ag.

Aged↗