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PubMed1976

Lipid vesicle-cell interactions. I. Hemagglutination and hemolysis.

The interaction of lipid vesicles (liposomes) of several different compositions with erythrocytes has been investigated. Lecithin liposomes, rendered positively charged with stearylamine, exhibit potent hemagglutination activity in media containing low concentrations of electrolytes. The hemagglutination titer is found to be a linear function of the zeta potential of the lipid vesicles. Hemagglutination is reduced when the surface potential of the cells is made more positive by pH adjustment or enzyme treatment. Similarly, hemagglutination is reduced by increasing concentrations of electrolytes. Hemagglutination is examined theoretically and is shown to be consistent with vesicle-cell interactions that are due to only electrostatic forces. Vesicles containing lysolecithin in addition to lecithin and stearylamine cause lysis of erythrocytes, provided the lipids of the vesicles are above the crystal-liquid crystal phase transition temperature. In addition, hemolysis requires close juxtaposition of the vesicle to the cell membrane; vesicles precoated with antibodies exhibit severely diminished hemolytic activities, only a small fraction of which can be attributed to a reduction in hemagglutination titer. Evidence is presented indicating that a single vesicle is sufficient to lyse one cell. With regard to hemagglutination and hemolysis, lipid vesicles of simple composition mimic paramyxoviruses such as Sendai virus.

Cell Aggregation↗
PubMed1951

Studies on host-virus interactions in the chick embryo-influenza virus system. IV. The role of inhibitors of hemagglutination in the evaluation of viral multiplication.

The role of inhibitors of hemagglutination in the evaluation of host-virus interactions in the chick embryo-influenza virus system has been analyzed. Comparisons were made between materials (allantoic fluids and membrane suspensions) derived from in vivo (growth curve) experiments at hourly intervals after inoculation, and from in vitro tests in which normal allantoic fluids and membrane suspensions were incubated with virus at 37 degrees C. for various periods of time. In both instances large amounts of virus were added to the systems, resulting in comparable concentrations of the agent. The seeds employed were either fully active or irradiated by ultraviolet light to the extent that the virus lost its capacity to increase but kept its interfering and hemagglutinating properties. The various materials were assayed for (a) the hemagglutinating titers of the virus present in the systems before and after heating to 56 degrees C.; (b) the concentration of inhibitor in the materials at various stages of incubation after heating to 70 degrees C. for 30 minutes as measured by the hemagglutination-inhibition reaction with native or heated test virus (30 minutes 56 degrees C.); and (c) the degree of adsorption of the hemagglutinins present in the materials onto chicken red cells at 0 degrees C. and their subsequent elution at 37 degrees C. The effects of receptor-destroying enzyme (RDE), treatment with sodium periodate, or high speed centrifugation on the inhibitory activities were studied in some of the tests. The essential results which indicate certain sources of error in the evaluation of host-virus interactions as well as means for studying virus activity at the early stages of the infectious process, were as follows: 1. Though some inhibitory effects on hemagglutination were noticeable in the allantoic fluid during the 1st hour after inoculation they were, as a rule, no longer apparent after this interval, and treatment with RDE did not increase the hemagglutinin titers. Thus, the interpretation of growth curve data concerning allantoic fluids hardly seems to be affected by inhibitor. On the other hand, striking effects were noted with the membrane suspensions of growth curve experiments in that RDE shortened the latent period to 2 hours and the titers in the first few positive samples (4 to 5 hours) increased) whereas in later harvests no such effect was noted. Under these conditions complement-fixation antigens and hemagglutinins made their appearance in the tissues simultaneously and not as previously reported the former prior to the latter. However, the infectivity showed increments only several hours after these two activities had become measurable. Thus the hypothesis of the stagewise development of influenza virus is still supported by these data. 2. Using the inhibition of hemagglutination technic it was found that the inhibitor in allantoic fluid rapidly decreased as a result of the action of active and irradiated virus, but destruction was never complete. In the membranes of the in vivo series only active seed led to loss of inhibitor, again without complete destruction, beginning at the time complement-fixing antigen and hemagglutinins became measurable. Irradiated seed was without effect in vivo whereas, in the in vitro tests it equalled the activity of the active virus. The implications of this difference in the effectiveness of active and irradiated seed in vivo with regard to the understanding of the mode of viral multiplication are discussed. 3. Although many factors may influence the shape of adsorption-elution curves it is felt that at 0 degrees C. the extent of adsorption is directly related to the amount of inhibitor present in the systems. In the early hours after inoculation the degree of adsorption was relatively small but it increased gradually with the time of incubation. The inhibitor of adsorption was destroyed by RDE and NaIO(4) and was only partially sedimentable by high speed centrifugation. In every respect studied its properties corresponded with the findings obtained with inhibitors in the hemagglutination-inhibition technic. Although the difference in the rapidity of inhibitor destruction as measured by the various technics might suggest a multiplicity of inhibitors it is felt that it rather denotes a greater sensitivity of the adsorption technic as compared to the others.

Animals↗
PubMed1998

Hemagglutinating activity of extracellular alkaline metalloendopeptidases from Vibrio sp. NUF-BPP1.

Alkaline metalloendopeptidase (metalloprotease) AP1 (48 kDa) from Vibrio sp. isolated from the intestine of a five-barred goatfish (Parupeneus trifasciatus) was reported in our previous paper to produce AP2 (36 kDa) by releasing a peptide fragment (molecular mass of about 12 kDa) from the C-terminal end of AP1 by autodigestion. AP1 strongly agglutinated fish (flounder, Paralichthys olivaceus) and rabbit erythrocytes, and weakly chicken erythrocytes. In contrast, AP2 had no significant hemagglutinating activity toward any erythrocytes tested, except for weak activity on flounder erythrocytes, suggesting that the C-terminal region of AP1 may be required for the strong hemagglutinating activity. The optimum temperature for the hemagglutinating activity of AP1 was found to be lower than that for the proteolytic activity. At acidic pHs (below pH 7.5), the hemagglutinating activity of AP1 decreased, and its pH profile resembled that of the proteolytic activity. The hemagglutinating activity of AP1 was not observed in the presence of o-phenanthroline or synthetic and proteinous substrates, but different kinds of saccharides and lipids had no effect. While the proteolytic activity of AP1 was not affected by CaCl2, the hemagglutinating activity of AP1 decreased with increases in CaCl2 concentrations. These results suggested that the hemagglutinating activity of these proteases (AP1 and AP2) was most likely caused by their proteolytic action on erythrocyte cell surfaces.

Animals↗
PubMed1992

Role of crl in avian pathogenic Escherichia coli: a knockout mutation of crl does not affect hemagglutination activity, fibronectin binding, or Curli production.

This study determined the role of crl in the production of curli by, the hemagglutination activity of, and fibronectin binding by avian pathogenic Escherichia coli chi 7122. Curli, an extracellular structure that binds fibronectin, was recently described (A. Olsén, A. Jonsson, and S. Normark, Nature [London] 338:652-655, 1989). The crl gene product was hypothesized to be the subunit monomer of curli and to bind fibronectin. E. coli HB101, which does not contain crl, binds fibronectin and produces curli when harboring a plasmid containing the crl gene. We show that HB101 hemagglutinates chicken erythrocytes when harboring the crl gene and that chi 7122 hemagglutinates chicken erythrocytes, binds fibronectin, and produces curli. Hemagglutination activity, fibronectin binding, and curli production are best expressed by chi 7122 and HB101 harboring the crl gene when the bacteria are grown on colonization factor antigen agar at 26 degrees C. The expression of hemagglutination activity, fibronectin binding, and curli production by both strains is decreased by growth on this agar at an increased temperature, of an increased osmolarity, or in an anaerobic atmosphere. This results indicates that the crl gene plays a role in the expression of the three phenotypes in HB101 and possibly in chi 7122 as well. We inactivated crl in chi 7122 by allele replacement in the expectation of abolishing hemagglutination activity, fibronectin binding, and curli production. The mutation was verified by Southern blot analysis and by a polymerase chain reaction, and there was no evidence of a second crl gene in chi 7122. However, the mutant of chi 7122 lacking crl hemagglutinated chicken erythrocytes, bound fibronectin, and produced curli at wild-type levels. This result indicates that crl plays a nonessential role in the expression of these three phenotypes in chi 7122.

Animals↗
PubMed2004

Inhibition of hemagglutinating activity by monoclonal antibody against Porphyromonas gingivalis 40-kDa outer membrane protein.

Periodontitis is a chronic inflammatory disease of periodontal tissues that results in alveolar bone loss, and Porphyromonas gingivalis, which has a high hemagglutinating activity, has been implicated as an important pathogen in the development of periodontitis. This bacterium has a high hemagglutinating activity. We previously succeeded in gene cloning the 40-kDa outer membrane protein (OMP) from P. gingivalis 381. Although recombinant (r) 40-kDa OMP itself did not show hemagglutinating activity, its polymeric form, constructed with a cross-linking reagent, significantly expressed that activity. Furthermore, an affinity-purified antibody against r40-kDa OMP inhibited the hemagglutinating activity of P. gingivalis vesicles. In the present study, in order to clarify the pathological role of 40-kDa OMP and develop passive immunotherapy, we examined the inhibitory effect of monoclonal antibodies (MAbs) against r40-kDa OMP on the hemagglutinating activity of P. gingivalis vesicles. The MAbs reacted with r40-kDa OMP, the outer membrane fraction, vesicles, and P. gingivalis cell extracts, and significantly inhibited the hemagglutinating activities of the polymeric r40-kDa OMP as well as of P. gingivalis vesicles. These findings suggest that MAbs against 40-kDa OMP may be useful for the development of passive immunotherapy and for assessing treatment for periodontal diseases caused by P. gingivalis infection.

Animals↗
PubMed1965

THE DETERMINATION OF ANTIBODY TO GROUP A STREPTOCOCCAL POLYSACCHARIDE IN HUMAN SERA BY HEMAGGLUTINATION.

A hemagglutination method employing tanned human O erythrocytes sensitized with purified Group A streptococcal polysaccharide has been developed to measure A polysaccharide antibody in antistreptococcal rabbit and human sera. The reaction of sensitized RBC with known Group A streptococcal rabbit antisera, inhibition of hemagglutination with A polysaccharide, and abolition of hemagglutination and hemagglutination-inhibition by exposure of polysaccharide antigen to A enzyme, which destroys the specific immunologic determinant of A carbohydrate, were utilized to demonstrate the specificity of this hemagglutinating system. The results of A polysaccharide antibody determinations on sera from normal persons, patients convalescent from streptococcal infection, and patients with non-suppurative streptococcal sequelae are presented.

Animals↗
PubMed1981

Lipid-mediated hemagglutination and its relevance to lectin-mediated agglutination.

Oleic acid and dioleoyl phosphatidic acid at low concentrations (20 and 0.5 mu g/ml, respectively) agglutinate rabbit and rat erythrocytes, while dioleoyl phosphatidylcholine is not hemagglutinating up to 0.5 mg/ml. Palmitic acid is not a hemagglutinin and dipalmitoyl phosphatidic acid is a very poor one. A polar lipid fraction obtained from calf thymocytes and a commercial preparation of gangliosides also exhibit pronounced hemagglutinating activity. Modification of the erythrocytes by either trypsin or neuraminidase causes a marked increase in agglutination only with oleic acid, whereas glutaraldehyde fixation of the cells significantly decreases agglutination with oleic acid, dioleoyl phosphatidic acid and calf thymocyte lipids. None of the lipids tested agglutinate freshly drawn human and sheep erythrocytes, but agglutination occurs following fixation of the sheep cells with glutaraldehyde. Lipid-mediated hemagglutination is strongly inhibited by fetuin and bovine submaxillary mucin (0.5 mg/ml). Defatted bovine serum albumin, also at 0.5 mg/ml, inhibits agglutination by oleic acid, whereas agglutination by other lipids is only poorly inhibited if at all. Monosaccharides at concentration up to 0.25 M do not inhibit the hemagglutinating activity of the lipids. Comparison of the hemagglutinating properties of lipids and lectins raises the possibility that the agglutinating activity of crude biological extracts which is not inhibited by mono- or oligosaccharides may be due to lipid constituents. Since agglutination by lipids is species specific, they may serve as mediators in intercellular recognition. The mechanism of lipid-mediated hemagglutination is discussed in terms of current concepts of the fusogenic activity of these compounds.

Animals↗
PubMed1984

Multiphasic interactions of Mycoplasma pulmonis with erythrocytes defined by adherence and hemagglutination.

The mechanism(s) of interaction between Mycoplasma pulmonis and eucaryotic cells was studied by adherence to and hemagglutination of erythrocytes. Simple and complex carbohydrates and glycoproteins were unable to inhibit either adherence or hemagglutination, indicating that neither was a lectin activity. Both interactions appeared to be hydrophobic due to their requirement for salt and their sensitivity to temperature. Hemagglutination, but not adherence, was inhibited by both trypsin and glutaraldehyde treatment of the mycoplasma, suggesting that adherence and hemagglutination are qualitatively different. The erythrocyte receptor sites for the two activities were also separable since hemagglutination, but not adherence, required trypsinization of erythrocytes. The hemagglutinin was shown to be an integral mycoplasma component and not a broth contaminant. Once removed, hemagglutinating activity could not be replenished by incubation in serum or broth at 4 degrees C, but could be regenerated during protein synthesis under nonreplicative conditions. Thus, a mycoplasma membrane protein was detected which was capable of interacting with opposing membrane surfaces through hydrophobic interactions. Consequently, a multiphasic model of M. pulmonis-eucaryotic cell interactions was proposed.

Animals↗
PubMed1999

Essential functional role of the polysaccharide intercellular adhesin of Staphylococcus epidermidis in hemagglutination.

Hemagglutination of erythrocytes is a common property of Staphylococcus epidermidis strains, which is related to adherence and biofilm formation and may be essential for the pathogenesis of biomaterial-associated infections caused by S. epidermidis. In three independent biofilm-producing, hemagglutination-positive S. epidermidis isolates, interruption of the icaADBC operon essential for polysaccharide intercellular adhesin (PIA) synthesis by Tn917 insertions led to a hemagglutination-negative phenotype. An immunoglobulin G fraction of antiserum to PIA greatly reduced hemagglutination. Purified PIA led to a 64-fold decrease of hemagglutination titers of these strains; however, it did not mediate hemagglutination by itself. These observations define PIA as the hemagglutinin of S. epidermidis or at least as its major functional component.

Adhesins, Bacterial↗
PubMed1966

Cimulative hemagglutination by Mycoplasma pneumoniae and other agglutinins.

John, T. Jacob (University of Colorado School of Medicine, Denver), Marlene Stahl, and Vincent A. Fulginiti. Cumulative hemagglutination by Mycoplasma pneumoniae and other agglutinins. J. Bacteriol. 92:1002-1004. 1966.-The phenomenon of cumulative hemagglutination, or hemagglutination by two agglutinins, each in subagglutinating concentration, was demonstrated by use of four different systems, namely, horse serum and Mycoplasma pneumoniae, horse serum and measles antigen, M. pneumoniae and measles antigen, and parainfluenza 2 virus and M. pneumoniae. Cumulative hemagglutination appears to be the mechanism by which a horse-serum diluent causes high hemagglutination titers of M. pneumoniae, since both contain hemagglutinins against vervet erythrocytes. It was also shown that antibodies against either one of the two antigens may cause inhibition of such hemagglutination.

Antibodies↗
PubMed1968

Heterogeneity of antibody response to Salmonella lipopolysaccharide measured by passive hemagglutination and hemolysis in mice.

The complement-requiring passive hemolysis test with Salmonella typhimurium lipopolysaccharide-coated sheep erythrocytes is more sensitive for antibodies directed against the lipopolysaccharide than is the passive hemagglutination test. The hemagglutinating and hemolyzing antibodies produced in Swiss mice by hyperimmunization, either with or without Freund's adjuvant, were distributed in both the light and heavy fractions isolated by sucrose density gradient fractionation and gel filtration. IgM fractions, whether tested by hemagglutination or hemolysis, were sensitive to 2-mercaptoethanol (0.15 m). On the other hand, IgG hemolytic antibodies were more sensitive to 2-mercaptoethanol than were IgG hemagglutinating antibodies. The resistance of IgG hemagglutinating activity amounted to about 72 to 95% of the total IgG recovered, whereas the resistant portion of the IgG hemolytic activity was approximately 40 to 53%. It is suggested that, although mercaptoethanol sensitivity is not a definitive test for IgM antibody, its use in connection with the hemagglutination test gives at least an approximation of the IgG antibody, whereas the hemolysis test gives a better approximation of maximal measurable antibody against Salmonella lipopolysaccharides.

Animals↗
PubMed1990

Hemagglutination properties of Streptococcus suis.

A total of 49 strains (23 reference strains and 26 field isolates) of Streptococcus suis were tested for their ability to agglutinate erythrocytes from different animal species. Ten different hemagglutination patterns were established. Thirty-three strains (67%) did not agglutinate any of the erythrocytes tested; sixteen strains (33%) agglutinated erythrocytes from one or more animal species. Different strains belonging to the same capsular type presented different hemagglutination patterns. No correlation was found between the tissue origin and/or the virulence (evaluated in 4-week-old mice) of different field isolates and their hemagglutination activity. Hydrophobic surface properties were also evaluated. All S. suis strains studied appeared to possess a hydrophilic cell surface. Morphologically similar fimbriae were observed on hemagglutinating as well as on nonhemagglutinating strains of S. suis. This study provides evidence that certain strains of S. suis possess hemagglutinating properties which do not appear to involve hydrophobic interactions. The possible role of fimbriae in hemagglutination remains unclear.

Animals↗
PubMed1992

Modified indirect hemagglutination test for detection of treponemal antibodies in finger-prick blood.

A modified indirect hemagglutination test for the detection of treponemal antibodies was developed for use with finger-prick blood. By using paired serum and absorbed finger-prick blood from 58 patients from an area previously endemic for yaws and 12 patients without yaws, the modified hemagglutination test was compared with a hemagglutination test for Treponema pallidum and the fluorescent treponemal antibody absorption test. The modified hemagglutination test showed 100% specificity and an overall agreement of 96.5% with the hemagglutination test for T. pallidum and 94.8% with the fluorescent treponemal antibody test. The modified hemagglutination test appears to be a simple and economical test that is suitable for use in large epidemiological surveys for yaws.

Adult↗
PubMed1986

Rubella virus antigens: localization of epitopes involved in hemagglutination and neutralization by using monoclonal antibodies.

Monoclonal antibodies (MAbs) against the rubella virion were used to locate epitopes involved in hemagglutination and neutralization. The MAbs exhibiting high-level hemagglutination-inhibiting activity were shown by Western blot analysis to be specific for the E1 polypeptide; this is consistent with the presence of the hemagglutinin on the E1 polypeptide. Some of the E1-specific MAbs also neutralized viral infectivity. However, hemagglutination-inhibiting activity and neutralizing activity did not always correlate. Three distinct functional epitopes were identified on the E1 polypeptide by competition analyses: one which reacted with MAbs with high-level hemagglutination-inhibiting activity and with neutralizing activity, one which reacted with MAbs with low-level hemagglutination-inhibiting activity and with neutralizing activity, and one which reacted with MAbs with only hemagglutination-inhibiting activity. A MAb specific for the E2 polypeptide exhibited neutralizing activity. This E2-specific MAb and two E1-specific MAbs with neutralizing activity were capable of precipitating intact virus which indicates that at least three epitopes involved in neutralization are accessible on the surface of the virion.

Animals↗
PubMed1991

A single-amino-acid substitution in polyomavirus VP1 correlates with plaque size and hemagglutination behavior.

The plaque size and hemagglutination characteristics of five cloned wild-type strains of polyomavirus were determined. The strains fell into two groups, those with large or small plaques, each with distinctive hemagglutination behavior at different temperatures and pHs. The nucleotide sequence of VP1, the major capsid protein of the virus, was determined for each of the viral strains. The PTA (large-plaque) and RA (small-plaque) strains differed only at residue 92 of VP1, where there is a glutamic acid or glycine, respectively (R. Freund, A. Calderone, C. J. Dawe, and T. L. Benjamin, J. Virol. 65:335-341, 1991). The same amino acid difference in VP1 correlated with plaque size and hemagglutination properties of the other sequenced viruses. Mutagenesis converting amino acid 92 from glutamic acid to glycine converted the plaque size and hemagglutination behavior of the large-plaque PTA strain to that of a small-plaque strain. Furthermore, PTA and RA VP1 proteins produced in Escherichia coli behaved as their parental viruses did in hemagglutination assays. These results demonstrate that amino acid residue 92 of VP1 is involved in determining the plaque size and hemagglutination behavior of polyomavirus and strongly suggest that this region of the VP1 polypeptide interacts directly with cell receptors.

Amino Acid Sequence↗
PubMed1990

Isolation and hemagglutinating activities of bovine immunoglobulins reactive with melibiose.

Calcium-dependent and -independent bovine immunoglobulins (IgM and IgG) reactive with melibiose were isolated by affinity chromatography on melibiose-coupled Sepharose 4B. Hemagglutination and hemagglutination inhibition were carried out to study their carbohydrate specificities. Human and animal erythrocytes were agglutinated by these bovine IgM, whereas those were not by bovine IgG at the highest concentrations used. Neuraminidase- and pronase-treated erythrocytes were more strongly agglutinated by these IgM than untreated ones. On the other hand, melibiose-reactive human immunoglobulins isolated from AB serum showed strong hemagglutinating activities to rabbit erythrocytes. Hemagglutination of neuraminidase-treated human type B erythrocytes by calcium-independent bovine IgM reactive with melibiose was effectively inhibited by galactose, methyl alpha-D-galactopyranoside and melibiose, whereas that was not by methyl beta-D-galactopyranoside, lactose, and other substances at the highest concentrations used. Similar results were also obtained in hemagglutination inhibition with untreated rabbit erythrocytes and calcium-independent human immunoglobulins (IgM and IgG) reactive with melibiose. However, hemagglutination of pronase-treated human type A erythrocytes by calcium-dependent bovine IgM reactive with melibiose was not at all inhibited by these substances at the highest concentrations used. From these results, bovine serum is found to also contain antibodies with a specificity for alpha-linked galactosyl residues as found for human AB serum reported previously.

Animals↗
PubMedDate not supplied

[Hemagglutination test and the diagnosis of food allergy].

We have studied 50 children suspected to have food allergy. Their clinical diagnoses included the following: digestive trouble (prolonged diarrhoea or vomiting), abdominal pain, repetitive urticaria, angioneurotic edema, eczema. The aim of thie study has been to value the results obtained with the hemagglutination test according to Boyden, comparing them with skin tests carried out through intradermal techniques. 113 hemagglutination and skin tests with varying foods have been carried out. Nearly all the children have been tested with milk, white and yolk of egg, the most suspected foods, and also other foods depending on the data found through anamnesis. With milk (47 cases) we have obtained positivity in 12 hemagglutination tests, and in 3 skin tests. With egg (41 cases) the hemagglutination test has been positive in 14 cases, and the skin test in 5 cases. Conjunctly in the 113 cases we have obtained positive hemagglutination test in 44 cases, and positive skin test in 14 cases. In 65 cases both tests have been negative. This fact points to the necessity to realize other diagnostic tests, as well the possibility that these children have no allergic disease. Summarizing, these results support the superior value of the hemagglutination Boyden test in comparison with the skin test as diagnosic proof in food allergy.

Adolescent↗
PubMed1995

Hemagglutination properties of Enterococcus.

In total, 86 enterococcal strains including representatives of most of the described species were tested for the ability to agglutinate human, sheep, and rabbit erythrocytes. Five strains did not react with any of the erythrocytes tested, and 81 (94.2%) strains agglutinated only rabbit erythrocytes. The hemagglutination titers ranged from 2 to 64. Loss of the hemagglutination activity was observed when rabbit erythrocytes were treated with trypsin or neuraminidase. Trypsin treatment of the bacterial suspensions also caused loss of the agglutination ability. On the other hand, heat treatment of bacterial suspensions increased the efficiency of the interactions, and higher titers were obtained. Assays for inhibition of hemagglutination were performed with alpha-D-fucose, alpha-D-galactose, beta-D-galactose, D-glucose, N-acetyl-galactosamine, N-acetyl-glucosamine, N-acetylneuraminic acid, N-acetylneuraminic acid-lactose, and fetuin. Only fetuin was able to inhibit the hemagglutination reactions. The results showed that hemagglutination properties are common to the different enterococcal species tested. They also suggest that enterococci possess hemagglutinins of proteic and non-proteic nature that are involved in the attachment to sialic acid-containing receptors on the surface of rabbit erythrocytes.

Adhesins, Bacterial↗