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J T Poolman

Publications and source records attributed to J T Poolman.

At least 91 records · Page 5Linked to original sources
PubMed1990

Description of a hybridoma bank towards Bordetella pertussis toxin and surface antigens.

This paper describes the development of a murine bank of monoclonal antibodies against Bordetella pertussis toxin, filamentous hemagglutinin (FHA), pili, lipopolysaccharide (LPS), or outer membrane proteins (OMPs). Subunits S1, S2, S3 of pertussis toxin (PT) bound immunoglobulins and glycoproteins such as fetuin and haptoglobin in an unspecific manner. The specificity of monoclonal antibodies towards subunits S1, S2, S3 or S4 of PT could be demonstrated by using purified immunoglobulins or their Fab2 fragments. A set of FHA-specific monoclonal antibodies could be differentiated on the basis of their binding to the various breakdown products present in FHA preparations. Pili-specific monoclonal antibodies reacted with either native pili or denatured pilin, and both demonstrated serotype specificity. Monoclonal antibodies to Bordetella pertussis OMPs were directed to either the virulent phase-regulated trypsin-sensitive, detergent-extractable OMPs 92 kDa, 32 kDa, and 30 kDa or the non-virulent phase-expressed, not-trypsin sensitive OMPs 38 kDa, 33kDa, and 18 kDa.

Animals↗
PubMed1990

Isolation of Neisseria meningitidis mutants deficient in class 1 (porA) and class 3 (porB) outer membrane proteins.

The class 1 major outer membrane protein of Neisseria meningitidis is a serious candidate for a meningococcal vaccine. To facilitate studies on the function of this protein, mutants were isolated that lacked this protein or the structurally related class 3 protein. These mutants were obtained by using the antibody-dependent bactericidal action of the complement system. The class 1 protein-deficient strain grew normally in vitro, whereas growth of the class 3 protein-deficient strain was slightly retarded. The class 3 protein-deficient strain displayed increased resistance to the antibiotics tetracycline and cefsulodin, which is consistent with the proposed role of the protein as a pore-forming protein. The class 1 protein was purified to homogeneity from the class 3 protein-deficient strain. Lipid bilayer experiments revealed that this protein also formed pores. The class 1 protein pores were cation selective.

Bacterial Outer Membrane Proteins↗
PubMed1990

Monoclonal antibodies against the 70-kilodalton iron-regulated protein of Neisseria meningitidis are bactericidal and strain specific.

When grown under iron limitation, Neisseria meningitidis expresses a number of outer membrane proteins (OMPs), one of which is a 70-kilodalton (kDa) major OMP. After immunization of mice with outer membrane preparations of iron-depleted cells of strain H44/76 (B:15:P1.7,16), hybridoma cell lines producing monoclonal antibodies against the 70-kDa OMP were obtained. Some of these monoclonal antibodies demonstrated strong bactericidal activity against the homologous strain H44/76 in the presence of human complement, suggesting potential application of the 70-kDa OMP as a vaccine component. However, none of the 10 selected monoclonal antibodies was able to recognize the corresponding protein from five heterologous strains of various serosubtyping characteristics. A polyclonal anti-70-kDa OMP serum also did not react with the other strains. This result shows that immunodominant surface-exposed epitopes of the meningococcal 70-kDa iron-limitation-inducible OMP are strain specific.

Animals↗
PubMed1990

Sero-subtyping of group B, C, Y and A meningococci isolated in France in 1988.

During 1988, 291 strains of various serogroups (B, C, Y and A) isolated in France, have been sero and sub-typed. 74.5 p. cent of the strains were isolated from CSF and blood of patients; 25.4 p. cent were isolated from carriers. The "whole cell ELISA" procedure was used with monoclonal antibodies. The distribution of serotypes among serogroup B strains was as follows: serotype 1 (8.1 p. cent); 2a (19 p. cent); 2b (16.2 p. cent); 4 (20.2 p. cent); 14 (13.5 p. cent); 15 (23 p. cent): 40 p. cent remained non typable (NT). Among serogroup C strains serotype 2a represented 74.5 p. cent of the strains. Serogroup A strains were only of the serotype 4. Serogroup Y strains belonged to serotype 14 or were NT. Some associations between serotypes and subtypes were predominant such as B:2a:P1.2 or B:2b:P1.2 or B:4:P1.15 or B:15:P1.7. Among serogroup C strains the antigenic pattern C:2a:P1.2 was most prevalent.

Enzyme-Linked Immunosorbent Assay↗
PubMed1989

Comparative evaluation of potential components for group B meningococcal vaccine by passive protection in the infant rat and in vitro bactericidal assay.

Seventeen monoclonal antibodies to one of three main cell surface antigens of Neisseria meningitidis group B were tested for protective efficacy in the infant rat using as challenge seven strains of different class 2/3 protein serotypes, class 1 protein (P1) subtypes and LPS immunotypes. Type-specific protection indicated both by a reduction of bacteraemia and meningitis and survival of the animals was regularly obtained with antibodies to the P1 protein and to LPS. By contrast, only one of seven antibodies to the serotype-specific class 2/3 protein was protective, even though four of them were highly bactericidal. The animal protection test and in vitro bactericidal assay were otherwise concordant. These data form important guidelines for the design of vaccines to prevent group B meningococcal infections.

Animals↗
PubMed1989

Clonal and variable properties of Neisseria meningitidis isolated from cases and carriers during and after an epidemic in The Gambia, West Africa.

A representative collection of meningococci was isolated from cases and healthy carriers in The Gambia between 1982 and 1988, during and after an epidemic of meningococcal meningitis. These bacteria were subjected to a clonal analysis. All serogroup A bacteria from both cases and carriers were of one clone (A IV-1). Several unrelated clones were observed among serogroup 29E and serogroup Y carrier strains. The serogroup A strains were uniform for serotype and subtype antigens (serotype 4, subtype P1.7) and antibiotic sensitivity pattern. Occasional strains varied in their lipopolysaccharide (LPS), DNA fingerprint pattern, and/or the quantitative expression of the class 1 protein. A high degree of strain-specific variation was found for the expression of class 5 proteins, pili, and sulfonamide sensitivity. The frequency of strains expressing reduced amounts of the class 1 protein, altered LPS, and/or increased amounts of capsular polysaccharide rose among case strains obtained after the epidemic had ceased. These strains seem to be generally resistant to antibody-mediated bactericidal activity.

Antibodies, Monoclonal↗
PubMed1989

Binding of pertussis toxin to eucaryotic cells and glycoproteins.

The binding of pertussis toxin and its subunits to cell surface receptors and purified glycoproteins was examined. The interaction of pertussis toxin with components of two variant Chinese hamster ovary (CHO) cell lines was studied. These cell lines are deficient in either sialic acid residues (LEC 2) or sialic acid and galactose residues (LEC 8) on cell surface macromolecules. The binding of pertussis toxin to components of these cells differed from the binding of the toxin to wild-type components. Although the toxin bound to a 165,000-dalton glycoprotein found in N-octylglucoside extracts of wild-type cells, it did not bind to components found in extracts of LEC 2 cells. In contrast, the toxin bound to components found in extracts of LEC 8 cells, which are variant cells that contain increased amounts of terminal N-acetylglucosamine residues on cell surface macromolecules. These results suggest that the receptor for pertussis toxin on CHO cells contains terminal acetamido-containing sugars. The cytopathic effect of the toxin on both types of variant cells was much reduced compared with its effects on wild-type cells. Thus, optimal functional binding of pertussis toxin appears to require a complete sialyllactosamine (NeuAc----Gal beta 4GlcNAc) sequence on surface macromolecules. In addition to studying the nature of the eucaryotic receptor for pertussis toxin, we examined corresponding binding sites for glycoproteins on the toxin molecule. Binding of both S2-S4 and S3-S4 dimers of the toxin to cellular components and purified glycoproteins was observed. The two dimers bound to a number of glycoproteins containing N-linked oligosaccharides but not O-linked oligosaccharides, and differences in the binding of the two dimers to some glycoproteins was noted. These data indicate that the holotoxin molecule contains at least two glycoprotein-binding sites which may have slightly different specificities for glycoproteins.

Adenosine Diphosphate Ribose↗
PubMed1989

Accessibility of gonococcal and meningococcal surface antigens: immunogold labeling for quantitative electron microscopy.

The parallel application of two electron microscopic immunogold labeling procedures was used to assess the surface exposure and accessibility of gonococcal and meningococcal surface antigens. Monoclonal antibodies were used as markers for the surface antigens, i.e., outer membrane proteins and lipooligosaccharides. To evaluate the labeling densities obtained after incubation of whole bacteria in suspension or ultrathin cryosections of bacteria, a method of electron microscopic quantitation was developed. Incubation of whole bacterial suspensions with monoclonal antibodies and protein A-gold resulted in specific labeling of the bacterial surfaces. However, the labeling densities varied largely in each cell. By contrast, cryosections showed uniform heavy labeling densities at the surface of the outer membranes of all cells. Apparently, by sectioning the cells the antigen-masking barrier could be evaded, and steric hindrance was no longer restrictive. Thus, a better estimate of both the presence and the surface exposure, i.e., the accessibility of antigens, could be made. Such information is essential for us to better understand host-bacterial interactions and to develop new vaccines.

Animals↗
PubMed1989

Purification, cyanogen bromide cleavage, and amino terminus sequencing of class 1 and class 3 outer membrane proteins of meningococci.

Meningococcal class 1 and 3 outer membrane proteins (OMPs) were subjected to cyanogen bromide treatment. The class 3 OMP was found to be resistant to cyanogen bromide, while the class 1 OMP was cut into two main fragments of 25 and 17 kilodaltons. The N-terminal sequences were determined for class 1 and class 3 proteins, which exhibit similarities to one another and to OMP I of gonococci. The C-terminal class 1 OMP fragment bound the bactericidal monoclonal antibodies tested.

Amino Acid Sequence↗
PubMed1988

Outer membrane protein serosubtyping of Neisseria meningitidis.

A system for typing Neisseria meningitidis has been developed which uses monoclonal antibodies against two separate classes of outer membrane proteins (class 1 and class 2/3) in addition to capsular polysaccharide serogrouping. It was shown that class 1 outer membrane protein subtypes are common to meningococcal reference strains of different serotypes. Application of the system to 50 group B meningococcal patient isolates revealed that 75% could be categorized as class 1 subtypes. The typing system has potential usefulness in epidemiological surveillance and vaccine development.

Antibodies, Monoclonal↗
PubMed1988

Two monoclonal antibodies specific for serotype 4 antigen of Neisseria meningitidis.

Two new monoclonal antibodies (MN14G21.17 and MN14E9.15) specific for the serotype 4 antigen of meningococci were isolated. The antibodies were raised against a previously non-typable serogroup B strain from the Netherlands and were shown to react with the serotype antigen of the prototype reference strains for serotype 4 and serogroup A as well as with that of the homologous strain. Further screening of 290 serogroup A and B case isolates with the monoclonal antibodies indicated that the serotype 4 epitope was present on all 100 serogroup A strains tested, including representative isolates from 28 epidemics, and on most isolates from a recent serogroup B epidemic (1981-1983) in Cuba. In addition, 68 isolates from recent sporadic cases (1980-1986) and/or clusters of cases of serogroup B disease in many parts of Europe, the USA, the USSR and Australia were also of this serotype.

Antibodies, Monoclonal↗
PubMed1988

Neisseria meningitidis group B serosubtyping using monoclonal antibodies in whole-cell ELISA.

Meningococci can be classified by the presence of serotype and subtype antigens on the class 2/3 and class 1 outer membrane proteins, respectively. A typing system employing monoclonal antibodies and a WCE for detection of such antigens enabled us to sero- and/or subtype 255 out of 268 group B meningococci from 19 countries collected from 1959 till 1987. The combination of a comprehensive collection of antibodies and a sensitive, specific, large-scale screening technique greatly facilitates the assignment of bacterial isolates to their sero- or subtype. This will then allow definitive, bass-line information on the epidemiology of meningococci and on the possible construction of outer membrane protein vaccines.

Antibodies, Monoclonal↗
PubMed1988

Definition of meningococcal class 1 OMP subtyping antigens by monoclonal antibodies.

The subtypes of meningococci are defined by antigenic determinants on the class 1 outer membrane proteins. The established subtypes, designated by P1 and a number according to the prototype reference strain on which they were first recognized by monoclonal antibodies, includes P1.2, P1.9, P1.15 and P1.16. We have investigated more prototype reference strains, using new monoclonal antibodies, and identified the new subtypes P1.1, P1.6 and P1.1,16. The P1.1,16 epitope is found on both the P1.1 and the P1.16 reference strains, but not on all P1.1 and P1.16 strains and can occur independently from the P1.1 and the P1.16 epitopes. It appears that class 1 outer membrane proteins contain at least two independent subtype-specific epitopes. For clarity, we now redefine P1.1,16 as P1.7, permitting thus the identification of strains of P1.1, P1.1,7, P1.7, P1.7,16 and P1.16 subtypes. It can clearly be expected that more class 1 outer membrane protein determinants will be recognized as more monoclonal typing antibodies are produced. The monoclonal antibodies now available to us can subtype 80-90% of group B and C meningococci; they also react with group A meningococci, but not with other Neisseriae. The immunological dissection of these subtyping antigens will improve our understanding of the relationship between components of the bacteria and the induction or prevention of disease.

Animals↗
PubMed1988

Electromorphic characterization and description of conserved epitopes of the lipooligosaccharides of group A Neisseria meningitidis.

We studied the lipooligosaccharides (LOS) of 28 group A Neisseria meningitidis of epidemiologically diverse origins to investigate whether each of the LOS serotypes found in serogroup A could be identified physically as well as antigenically. Using a dot blot assay with LOS-specific monoclonal antibodies (MAbs), we identified four epitopes that were serotype specific. The LOS from strains of each serotype were electromorphically and antigenically distinct when analyzed by silver-stained sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. The LOS of L8 strains contained a 3,600-Mr component that bound the L8 MAb. The LOS of L9 strains contained two major components of 4,500 and 4,200 Mr. They bound the L9 MAb to the larger component. The LOS of L10 strains had a single major component of 4,000 Mr that bound the L10 MAb. The LOS of L11 strains contained a major 3,600-Mr component that could not be distinguished from the 3,600-Mr LOS of L8 strains by SDS-PAGE but that bound the L11 MAb. LOS of group A strains contained a highly conserved epitope in addition to a serotype-specific epitope. This was identified by a MAb that bound to all the strains on dot-blots and to multiple LOS components of various Mrs on immunoblots. We conclude that the LOS which bear the L9, L10, and L11 determinants are physically distinct and can be identified by SDS-PAGE or MAb binding or both. L8 and L11 are both borne on a 3.6-kilodalton LOS and can only be distinguished serologically.

Antibodies, Bacterial↗
PubMed1988

Biochemical properties of pertussis toxin.

Pertussis toxin is an exotoxin produced by the organism Bordetella pertussis. The toxin binds to receptors on the eukaryotic cell surface. After introduction into the eukaryotic cell, the toxin is activated by ATP and subsequently ADP-ribosylates a family of GTP-binding regulatory proteins interrupting signal transduction within the cell. We have examined the location of several critical sites on the toxin molecule. These sites include the receptor binding site and the ATP binding site. The B oligomer of the toxin was found to contain at least two sites capable of binding glycoproteins suggesting that the B oligomer may have more than one eukaryotic cell receptor binding site. ATP was also shown to bind to a site on the B oligomer. These results indicate that the B oligomer contains several sites necessary for toxin action.

Adenosine Diphosphate Ribose↗