Allergenicity and cross-reactivity of rye grass pollen extracts revealed by monoclonal antibodies

Mourad, Walid; Pelletier, G.; Boulet, A. P.; Islam, Nazrul; Valet, Jean-Paul; Hébert, Jacques

doi:10.1016/0022-1759(86)90031-1

Cited by 34 publications

(26 citation statements)

References 10 publications

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“…To date, grass pollen allergens are divided into 11 groups [1, 2, 3, 4, 5], of which group 1 and group 5 have been identified as the most important. Several group 5 molecules have already been purified and/or cloned [6, 7, 8, 9, 10, 11, 12, 13, 14]and are described as a group of nonglycosylated, highly homologous proteins with an apparent molecular mass (M r ) on SDS-PAGE of 25–33 kDa [15, 16, 17].…”

Section: Introductionmentioning

confidence: 99%

The Major Grass Pollen Group 5 Allergen from Dactylis glomerata and Its C-Terminal Split Product Both Behave as Dimers: Implications for Allergen Standardization

Oort

Dieker

Heer

et al. 2005

Int Arch Allergy Immunol

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Background: On SDS-PAGE grass pollen group-5 allergens migrate as a doublet with an apparent molecular mass (M_r) of 25 kDa. Immunoblot analysis revealed additional group 5 reactivity at double and half this M_r. The aim of this study was to investigate these group 5 molecular entities and to compare their allergenicity and behavior in quantitative immunoassays. Methods: Group-5-specific monoclonal antibodies were produced and used for the development of a group-5-specific sandwich ELISA. Affinity-purified Dac g 5 was separated by SDS-PAGE/Western blotting; individual bands were analyzed by N-terminal sequencing. Size exclusion chromatography (SEC) in conjunction with group-5-specific ELISA, competitive RIA and RAST inhibition were used to analyze the size distribution of Dac g 5. Basophil histamine release assays were used to assess biological activity. Results: The lower band of the typical group 5 doublet was identified as a truncated form lacking the typical group 5 N-terminus AD^L/_AGY, observed in the upper band. The 12-kDa peptide was shown to be the C-terminal half of Dac g 5 (amino acid 127 onwards). SEC in conjunction with competitive RIA revealed that around 45% of Dac g 5 is represented by the 12-kDa peptide. Both the C-terminal half and the whole allergen dimerize under nondenaturing conditions. In competitive RIA and RAST inhibition both forms are equally well detected. In contrast, the half molecule is poorly recognized in sandwich ELISA and displays negligible biological activity in basophil histamine release tests with purified IgE. Conclusions: These observations stress the need to evaluate the performance of allergen standardization protocols in detail, with special attention to allergen size distribution.

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Section: Introductionmentioning

confidence: 99%

The Major Grass Pollen Group 5 Allergen from Dactylis glomerata and Its C-Terminal Split Product Both Behave as Dimers: Implications for Allergen Standardization

Oort

Dieker

Heer

et al. 2005

Int Arch Allergy Immunol

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“…To date grass pollen allergens are divided into 11 groups [1, 2], of which group 1 and group 5 are identified as the most important allergens [3, 4, 5, 6]. …”

Section: Introductionmentioning

confidence: 99%

Immunochemical Characterization of Two Pichia pastoris-Derived Recombinant Group 5 Dactylis glomerata Isoallergens

Oort

Heer

Lerouge

et al. 2001

Int Arch Allergy Immunol

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Background: Grass pollen of the Poaceae grasses are known to be highly allergenic. Major allergens from the species Lolium, Phleum, Poa and Holcus have been cloned and expressed as recombinant proteins, but of the important species Dactylis glomerata no recombinants are available. Methods: Dac g 5 was cloned by PCR on the basis of homology with Lol p 5 and expressed in Pichia pastoris. Recombinant Dac g 5 (rDac g 5) was affinity purified and compared to natural Dac g 5 (nDac g 5) by immunoblot, radioallergosorbent test (RAST), RAST inhibition, basophil histamine release assay (HRA), competitive radioimmunoassay (RIA) and sandwich enzyme-linked immunosorbent assay (ELISA). In addition, N-terminal sequencing, concanavalin A (Con A) binding, circular dichroism spectrum measurements and matrix-assisted laser desorption ionization-time of flight mass-spectrometric analysis were performed. Results: Clones were obtained that coded for pro-Dac g 5 and two mature isoforms of Dac g 5; the deduced amino acid sequences of both isoforms differed by 4 amino acids. Both mature isoforms were expressed in Pichia at a concentration of ∼15 mg/l. SDS-PAGE analysis showed that rDac g 5 had an apparent M_r approximately 10 kD above nDac g 5. By mass spectrometry this difference was shown to be around 2.5 kD. Positive Con A staining suggested (O-linked) glycosylation as an explanation for this increase in M_r. Whereas both purified recombinants showed a tendency to dimerize, purified nDac g 5 contained a 12-kD peptide not observed for rDac g 5. RAST, RAST inhibition and HRA showed that the IgE reactivity of rDac g 5 was similar to that of nDac g 5. A small subgroup, however, clearly demonstrated decreased IgE reactivity to rDac g 5.02. Differences in immune reactivity of both isoforms were confirmed by monoclonal antibody (mAb)-based sandwich ELISA. Conclusions: Dac g 5 was successfully cloned and expressed in P. pastoris. Minor differences in primary structure between isoforms influence their immune reactivity.

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“…Using direet binding and binding inhibition of Id to Loip I (Fig. 5), we also observed, after injection o( Loip I (week 14), that the levels of Id 29()A-I67 were significantly higher in mice pretreated wilh aid MoAb than in control animals. Interestingly, as shown by direet binding assay, mice pretrealed with anli-290A-167 MoAb produced higher levels of other Id (348A-6 and 539A-6) than mice that received control MoAb.…”

Section: Effects Of Pretreatment With Aid Moab On Idiotypic Responses Tomentioning

confidence: 83%

“…Montreal, Canada) by immunosorbent technique using anti-Z,o/;? I MoAb (290A-I67) fixed on solid phase as previously described [14].…”

Section: Antihodies and Antigensmentioning

confidence: 99%

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Modulation of immune response toLol pI by pretreatment with anti-idiotypic antibody is not restricted to the idiotypic expression

Boutin¹,

Hébert²

1994

Clinical and Experimental Immunology

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SUMMARYTo study the role of anti-idiotypic antibodies in the regulation oflhe immune response to Lolp 1 (the major allergenic component of rye grass pollen), we have recently generated a panel of three MoAbs directed against distinct epitopes of L«//71 and an anti-idiotypic MoAh directed against Ihc idiotype borne by one of the anti-Lol p I MoAbs (290A-I67). The effects of pretreatment with this aniiidiotypic MoAb in BALB/c mice before immunization wilh the antigen have been examined. The anti-idiotypic MoAb or unrelated MoAb were given weekly for 8 weeks intraperitoneally. Mice then received the antigen (2 /jg) adsorbed with alum (2 mg) at weeks 9. I! and 1.1. Scrum anti-Lf)/ p I antibodies (IgG or IgE) and specihc idiotypic responses were measured. \n\\-Lolp I IgG antibodies could be detected before immunization with Lolp I only in mice pretreated with anli-idiotypic MoAb. Immunization with Lo//? I induced an anti-L(>//)ngG response in both groups, but this response was higher in mice that received anti-idiotypic MoAb. Similar prolilcs were seen for specific IgE antibodies and idiotypic responses. Surprisingly, idiotypes borne by other anti-Ac//) I MoAbs (539A-6 and M8A-(i) had also been enhanced after pretreatment with the anti-290A-l67 MoAb. These observations suggested thai the pretreatment with this anti-idiolypic MoAb modulates not only Ihc expression of the respective idiotype. but also affects other idiotype responses.

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Allergenicity and cross-reactivity of rye grass pollen extracts revealed by monoclonal antibodies

Cited by 34 publications

References 10 publications

The Major Grass Pollen Group 5 Allergen from <i>Dactylis glomerata </i>and Its C-Terminal Split Product Both Behave as Dimers: Implications for Allergen Standardization

The Major Grass Pollen Group 5 Allergen from <i>Dactylis glomerata </i>and Its C-Terminal Split Product Both Behave as Dimers: Implications for Allergen Standardization

Immunochemical Characterization of Two <i>Pichia pastoris-</i>Derived Recombinant Group 5 <i>Dactylis glomerata</i> Isoallergens

Modulation of immune response toLol pI by pretreatment with anti-idiotypic antibody is not restricted to the idiotypic expression

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