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Biomedical subjects

H Snippe

Publications and source records attributed to H Snippe.

At least 73 records · Page 4Linked to original sources
PubMed1991

A protective monoclonal anti-idiotypic vaccine to lethal Semliki Forest virus infection in BALB/c mice.

Two monoclonal anti-idiotypic antibodies (ab2 MAbs), designated 1.13A112 (immunoglobulin G type 2a [IgG2a]) and 1.13A321 (IgG1), were prepared against Semliki Forest virus (SFV)-neutralizing ab1 MAb UM 1.13. They were identified in hybridoma supernatant fluid by their capacity to block UM 1.13-mediated neutralization of SFV. Although the neutralization-blocking capacities of the ab2 MAbs did not differ, only 1.13A321 evoked SFV-neutralizing ab3 antibodies upon intracutaneous and subcutaneous immunization of BALB/c mice with 1.13A321 chemically cross-linked to keyhole limpet hemocyanin and combined with the adjuvant Quil A. SFV-neutralizing ab3 antibodies appeared in serum within 10 days after primary immunization, and neutralizing antibody titers could be as high as 1/1,000 at day 35. All mice who had developed SFV-neutralizing antibodies upon anti-idiotypic immunization survived an otherwise lethal challenge with virulent SFV. However, induction of SFV-neutralizing ab3 antibodies by ab2 MAb 1.13A321 proved to be genetically restricted to BALB/c mice; even haplotype-identical (H-2d) DBA/2 mice did not respond, and consequently those animals died after infection with virulent SFV.

Animals↗
PubMed1990

Measurement of the humoral immune response against Streptococcus pneumoniae type 14-derived antigens by an ELISA and ELISPOT assay based on biotin-avidin technology.

A Streptococcus pneumoniae type 14-specific ELISA and ELISPOT assay have been developed based on the use of biotinylated type 14 capsular polysaccharide (S14PS-biotin). A major advantage of this application over other methods is the use of 10-100-fold less antigen than that reported in the literature for other similar assays. Moreover, the prepared biotinylated polysaccharides are very stable and it is possible to use the same procedures for other pneumococcal polysaccharide antigens (e.g., S6BPS) with no major changes necessary in the ELISA and ELISPOT protocols. Furthermore, a simple thin layer chromatography method has been developed as a method for quality control of the biotinylated polysaccharide. Immunization with the thymus-independent antigen S14PS resulted in the induction of IgM spot-forming cells (SFC) and antibodies while S14PS-protein conjugates induced a thymus-dependent response. The immune response to the conjugates was enhanced by the addition of the adjuvant Quil A resulting in high levels of both IgG SFC and antibodies at day 14 after immunization. The developed assays are reliable and reproducible tools for studying the humoral immune response against Streptococcus pneumoniae type 14 capsular polysaccharide derived antigens.

Animals↗
PubMed1990

Blocking by anti-idiotypic antibodies of monoclonal antibody-mediated protection against lethal Semliki Forest virus in mice.

Semliki Forest virus-(SEV) neutralizing monoclonal antibodies (MoAbs), produced after fusion of spleen cells from BALB/c mice and myeloma cell line P3-X63-AG8. 653 or SP2/0, were used for anti-idiotypic immunization of female BALB/c mice. Two intracutaneous immunizations (2 x 40 micrograms per animal), 3 weeks apart, with keyhole limpet haemocyanin-conjugated MoAbs mixed with the saponin Quil A were sufficient to induce high levels of anti-idiotypic antibodies in the circulation of these mice with the capacity to block specifically in vitro MoAb-mediated virus neutralization. Anti-idiotypic antibodies against SFV-neutralizing MoAbs, either passively transferred or actively acquired by immunization, are also able to abrogate (specifically) passive immunity, mediated by critical protective doses of MoAb, in mice against infection with a lethal strain of SFV. Furthermore we confirmed by intervention with anti-idiotypic serum in vivo that an SFV-neutralizing MoAb exerts its greatest protective effect during the first 2 days of infection.

Adjuvants, Immunologic↗
PubMed1989

Nonionic block polymer surfactants enhance the avidity of antibodies in polyclonal antisera against Streptococcus pneumoniae type 3 in normal and Xid mice.

Avidities of antibody (sub)classes in polyclonal antisera against Streptococcus pneumoniae type 3 (S3) can be (semi) quantitatively determined with a specific inhibition ELISA. A hexasaccharide was isolated from the hydrolyzed S3 capsular polysaccharide and coupled to a protein-carrier. Mixtures containing these conjugates and nonionic block polymer (NBP) surfactants were used for immunization. After various immunizations of these conjugates without NBP the anti-S3 specific antibodies of IgM and IgG2a isotype decreased in both antibody level and avidity. The adjuvants NBP 1501 and L121 not only enhanced the hexasaccharide-protein induced IgM and IgG antibody levels but also clearly increased the avidity of the two antibody (sub)classes IgM and IgG2a. This effect was observed in normal (data not shown) and X-linked immunodefective mice. A maturation of the IgG antibody response was realized by the second immunization with hexasaccharide-protein conjugate whereas the third immunization showed no further increase in antibody level and avidity.

Adjuvants, Immunologic↗
PubMed1989

Identification of a DTH-inducing T-cell epitope on the E2 membrane protein of Semliki Forest virus.

Mapping of T-cell epitopes on the structural proteins of Semliki Forest virus (SFV) was performed by measuring the ability of cloned SFV protein fragments to induce delayed-type hypersensitivity (DTH). The cloned SFV protein fragments were expressed as hybrid proteins with cro-beta-galactosidase in Escherichia coli from constructed recombinant plasmids. DTH reactions were measured, as footpad swelling, in BALB/c mice after immunization with whole, UV-inactivated SFV and challenge with the hybrid proteins, and vice versa, using the adjuvant dimethyl dioctadecyl ammonium bromide to enhance DTH. Only two of the tested hybrid proteins induced DTH, and these DTH reactions were equally strong. The largest DTH-inducing hybrid protein contained the N-terminal 350 amino acids of E2 and part of E3, the smallest contained only the region from amino acid residues 115 to 151 of the E2 membrane protein without any other SFV protein parts. It was concluded that the segment between amino acid residues 115 and 151 of the E2 membrane protein of SFV was responsible for the observed DTH, and thus, contains a T-cell epitope. Sequence homology with known T-cell epitopes on other proteins makes it likely that the DTH-inducing T-cell epitope is located from amino acid residues 120 to 128 of E2.

Adjuvants, Immunologic↗
PubMed1989

Phagocytosis of herpes simplex virus by human granulocytes and monocytes.

Polymorphonuclear leukocytes (PMN) can mediate cytotoxic reactions against virus infected targets cells. We observed very efficient binding of PMN to HSV-infected fibroblasts when loaded with HSV-specific antibodies. Using electron microscopy, infected fibroblasts were found to be totally surrounded by PMN and the phagocytosis of virions and fragments of infected cells was demonstrated. To quantify and study this phenomenon, and to compare PMN with monocytes, we developed radiometric and fluorometric phagocytosis assays. Leukocytes were mixed with [3H]glucosamine- or FITC-labeled virus and incubated at 37 degrees C. PMN associated radioactivity or fluorescence per cell as measured by flow cytometry was determined. PMN phagocytosis was dependent on the presence of specific anti-HSV antibodies and could be enhanced by addition of complement. Monocytes were also able to phagocytize virions; however, the rate of uptake was less than that for PMN. Under optimal conditions the total amount of herpes simplex particles that could be associated with one PMN or monocyte was about 10,000. PMN and monocytes are capable of phagocytosis of HSV. This may be an important factor in preventing the spread of infection in vivo.

Antibodies, Viral↗
PubMed1989

Estimation of the avidity of antibodies in polyclonal antisera against Streptococcus pneumoniae type 3 by inhibition ELISA.

The reliability of the determination of antibody avidity in polyclonal sera by indirect sandwich ELISA was studied. Binding of IgM and IgG (sub)classes in unpurified serum to Streptococcus pneumoniae type 3 capsular polysaccharide, which was coated onto ELISA plates, was inhibited with different inhibitors. The inhibitor concn at which 50% inhibition of antibody binding to the ELISA coat was achieved, was used as a measure for antibody avidity. As this 50% inhibition value is dependent upon the dilution of the serum and thus upon the initial amount of free antibody, it is necessary to define (a narrow range of) final ELISA absorbance values to which the dilutions of non-inhibited sera have to be adjusted. The shapes of the serum dilution curves have a good correlation with the numerical 50% inhibition values of the antibody avidity. The inhibition ELISA is suitable to compare the avidity values of the different antibody isotypes, but two remarks should be made: (1) antibody heterogeneity should be considered to influence the results and prevent the accurate measurement of absolute numerical avidity values. Because in the ELISA system merely antibody "activity" is measured, comparison of the efficacy of vaccines by means of the 50% inhibition (avidity) value of various antibody (sub)classes can still be performed in a reliable way; (2) results of the determination of the 50% inhibition values of the different antibody (sub)classes showed them to be dependent on the molecular ratio between antibody (sub)class levels. More aspects of the determination should be taken into account, like shapes of simple dilution curves, influences of various inhibitor concns in the diluent and whole (extended) inhibition curves.

Animals↗
PubMed1989

The use of a hybridization assay for the study of host defences against herpes simplex virus.

A rapid and simple hybridization assay was developed as an alternative for virus titration for the investigation of host resistance against HSV-1 infections in vitro. The probe which was constructed for this assay was shown to be HSV-1-specific. When a monolayer of fibroblasts was infected for 24 h before hybridization, 15 PFU were detected reliably. A plateau in hybridization levels was found when the multiplicity of infection reached 1. In order to demonstrate the applicability of the probe for the study of host defences against HSV in vitro, fibroblasts were infected with HSV in the presence of different numbers of adherent cells and different concentrations of serum containing high titres of anti-HSV antibodies and complement. After 20 h of incubation, samples were lysed, spotted on Zetaprobe filter paper and hybridized with a 32P-labelled RNA probe. Spots were counted for radioactivity. The radioactivity was taken as a measure of the success of infection. Results showed that at high (10%) concentrations of serum containing high titres of anti-HSV antibodies and complement neutralization plays an important role. At low (1%) concentrations of serum containing high titres of anti-HSV antibodies and complement the phagocytic role of adherent cells becomes the dominant factor in preventing infection of the fibroblasts. However, when the number of infectious particles is increased, the protection provided by adherent cells is overwhelmed.

Antibodies, Viral↗
PubMed1989

Modulation of the immune response to pneumococcal type 14 capsular polysaccharide-protein conjugates by the adjuvant Quil A depends on the properties of the conjugates.

Streptococcus pneumoniae type 14 capsular polysaccharide-bovine serum albumin (S14PS-BSA) conjugates were prepared by water-soluble-carbodiimide-mediated condensation with or without the use of N-hydroxy-sulfosuccinimide. The immunogenicities of the capsular polysaccharide (S14PS) and of the conjugates were studied in (CBA/N x BALB/c)F1 mice and in female BALB/c mice. The response in these mice indicates that S14PS could be classified as a thymus-independent type 2 antigen. Coupling of S14PS to BSA improved the immunogenicity of this polysaccharide, and an immunoglobulin G memory response was evoked. Conjugation with N-hydroxysulfosuccinimide resulted in a product with a higher polysaccharide/protein ratio. This conjugate induced a greater immune response than did the classical conjugate. Quil A enhanced the immune response to S14PS and to most S14PS-BSA conjugates. The enhancement of the immune response to the conjugates seemed to depend on the coupling procedure. Our results indicate that for the construction of immunostimulating complexes based on polysaccharide or oligosaccharide-protein conjugates, attention should be paid to the degree of cross-linking of the antigens involved.

Adjuvants, Immunologic↗
PubMed1989

Nonionic block polymer surfactants modulate the humoral immune response against Streptococcus pneumoniae-derived hexasaccharide-protein conjugates.

Nonionic block polymer surfactants (NBPs) were tested for the capacity to stimulate the antibody response against hexasaccharide (HS), derived from Streptococcus pneumoniae type 3 capsular polysaccharide (S3PS), which was conjugated to proteins. The immune response was evaluated in the (CBA/N x BALB/c)F1 progeny, in which female mice are phenotypically normal whereas male mice carry an X-chromosome-linked immunodeficiency. NBPs L101, L121, 1101, and 1501 were able to increase anti-HS immunoglobulin M (IgM) and IgG levels in both normal and X-chromosome-linked immunodeficient mice (with up to 74-fold stimulation of antibody titers). Distribution of S3PS-specific antibodies over the various IgG isotypes was restricted after immunization with either HS-bovine serum albumin or HS-keyhole limpet hemocyanin (HS-KLH). Addition of NBPs (in particular 1501) resulted in a more diverse immune response with either antigen as judged by isotype distribution. Isoelectric focusing of individual sera and subsequent detection of S3PS-binding antibodies in these sera by immunochemical staining revealed a restricted number of different spectrotypes in the course of the immune response. Upon immunization of mice with HS-KLH, spectra of secreted antibodies were slightly more complex and more densely stained than after immunization with HS-bovine serum albumin. Furthermore, NBPs 1101 and 1501 appeared to be able to stimulate the secretion of antibodies, which were secreted only in small amounts without the use of NBPs. Different explanations for increased spectrotype diversity after immunization with KLH as the carrier and after administration of NBPs as the adjuvant are discussed.

Adjuvants, Immunologic↗
PubMed1988

A neutralization-inhibition enzyme immunoassay for anti-idiotypic antibodies that block monoclonal antibodies neutralizing Semliki Forest virus.

In this paper we compare a solid-phase enzyme immunoassay (EIA) with a neutralization-inhibition enzyme immunoassay (NI-EIA) for the determination of anti-idiotypic antibodies against Semliki Forest virus (SFV)-neutralizing monoclonal antibodies (MAs) UM 5.1 (IgG2a) and UM 1.4 (IgG2a). Against these MAs strong immune sera were induced in female BALB/c mice by two subcutaneous injections, 3 weeks apart, with keyhole limpet hemocyanin coupled MA mixed with the saponin Quil A. Rabbit immune sera were prepared by intracutaneous injections of purified MA mixed with either FCA (first immunization) or IFA (second and third immunization). In the NI-EIA serum is preincubated with neutralizing MA, in wells of 96-well plates, before SFV is added. Binding of anti-idiotypic antibodies to MA results in a diminished capacity of that MA to neutralize SFV. After 1 h incubation with SFV L cells are added and residual infectious virus is allowed to multiply for 6 h at 37 degrees C. Then the monolayers are fixed with glutaraldehyde and subsequently SFV is quantified with a horseradish peroxidase-labelled SFV-specific MA. Low absorbance values indicate that the neutralizing capacity of MA is intact and that blocking antibodies were not present in serum. In contrast high absorbance values indicate that blocking (anti-idiotypic) antibodies had abrogated the neutralizing capacity of MA. With the strongly neutralizing MA UM 5.1 as idiotypic antigen the NI-EIA proved to be at least as sensitive as the solid-phase EIA. Furthermore both normal mouse serum-absorbed rabbit immune sera and mouse immune sera were not cross-reactive in both solid-phase EIA and NI-EIA.

Animals↗
PubMed1988

Measurement of the humoral immune response against Streptococcus pneumoniae type 3 capsular polysaccharide and oligosaccharide containing antigens by ELISA and ELISPOT techniques.

A sensitive ELISA has been developed to study immune responses in mice against Streptococcus pneumoniae type 3 capsular polysaccharide (S3PS) and hexasaccharide (HS)-protein conjugates derived therefrom. An advantage of the described system is that the same microtiter plates can be used for both ELISA and ELISPOT tests with a standardized washing procedure and diluent composition. S3PS induced predominantly IgM antibodies and minute amounts of IgG as measured by ELISA in serum. This was accompanied by large numbers (greater than 14000) of IgM spot-forming cells in the spleen. A shift towards IgG production was achieved by addition of lipid A. HS-protein conjugates induced predominantly IgG antibodies after booster immunization(s). Furthermore these conjugates induced large numbers (greater than 40000) of IgG spot-forming cells (SFC) in the spleen. ELISA and ELISPOT assays on microtiter plates are both reliable and highly reproducible assays for the evaluation of immune responses to S. pneumoniae antigens.

Animals↗
PubMed1988

Discrimination of determinant specificity of two encephalomyocarditis virus neutralizing monoclonal antibodies by competition, mixed neutralization and anti-idiotypic antibodies.

To investigate the epitope(s) on encephalomyocarditis virus (EMCV) involved in neutralization, two neutralizing monoclonal antibodies (MAs) (MA UM 21.1 and MA UM 21.2) were tested in a competition binding assay (CBA), a mixed neutralization test and an enzyme immunoassay (EIA) with specificity for the detection of idiotypes on MAs. With a CBA in cell culture, using EMCV infected L cell monolayers as binding antigen, strong homologous competition was observed between unlabelled MAs and horse radish peroxidase (HRPO-) labelled MAs but considerable heterologous competition did also occur, especially between the unlabelled MA UM 21.1 and HRPO-labelled MA UM 21.2. In the mixed neutralization test (50% plaque reduction) preincubation with slightly neutralizing or nonneutralizing doses of MA UM 21.2 had no diminishing effect on the neutralizing capacity of MA UM 21.1, the PRT50 value remains in all cases -10log titre of 5.8. Furthermore rabbit polyclonal antibodies against the idiotypes of MAs UM 21.1 and UM 21.2 did not cross react in the EIA. In conclusion both MAs recognize different viral determinants as indicated by the results obtained with a CBA, a mixed neutralization test and an EIA for detection of idiotypes on MAs.

Animals↗