Concentrations of the major birch tree allergen Bet v 1 in pollen and respirable fine particles in the atmosphere

doi:10.1016/S0091-6749(97)70170-2

Journal of Allergy and Clinical Immunology

Volume 100, Issue 5, November 1997, Pages 656-661

https://doi.org/10.1016/S0091-6749(97)70170-2 Get rights and content

Abstract

Background: Birch tree pollen allergens are an important cause of early spring hay fever and allergic asthma. Pollen counts provide a guide for individuals with birch pollen allergy. However, birch pollen, because of its size, has a low probability of entering the lower airways to trigger asthma. Yet birch pollen allergens are known to be associated with respirable particles present in the atmosphere. Objective: We sought to determine the concentration of major allergen Bet v 1 in birch pollen and respirable particles in the atmosphere during the birch pollen season. Methods: We used a two-site monoclonal antibody–based assay (ELISA) to quantitate Bet v 1 in pollen extracts and high-volume air sampler filters collecting particles larger and smaller than 7.2 μm. Results: Bet v 1 (0.006 ng) is detectable per birch pollen grain, of which 0.004 ng is present in aqueous extracts (13.9% of soluble proteins). Atmospheric Bet v 1 concentrations are correlated with birch pollen counts. Heavy rainfall tended to wash out pollen and particles, indicated by a mean daily Bet v 1 concentration of 0.12 ng/m³ (20 pollen equivalents), but light rainfall produced a dramatic increase in allergen-loaded respirable particles with Bet v 1 concentrations of 1.2 ng/m³ (200 pollen equivalents). Conclusion: These results highlight the different environmental risk factors for hay fever and allergic asthma in patients sensitized to Bet v 1. Light rainfall causes an increase in respirable particles; hence, this is an important risk factor for asthma. (J Allergy Clin Immunol 1997;100:656-61.)

Section snippets

Quantification of Bet v 1

A monoclonal antibody–based two-site binding assay (ELISA) was used to quantitate the Bet v 1 concentrations, essentially as described previously.⁷ Briefly, each well of a microtiter plate (Sarstedt Inc., Newton, N.C.) was coated with 100 μl of purified monoclonal antibody 3B4F11D6 (Allergopharma Joachim Ganzer KG, Reinbek/Hamburg, Germany) specific to Bet v 1 at 10 μg/ml in sodium carbonate buffer overnight at 4° C. All subsequent incubations were performed at room temperature. After blocking

Concentration of Bet v 1 in birch pollen grains

As determined by the quantitative two-step ELISA, the soluble extract of birch pollen contains reasonably high yields of Bet v 1 (mean, 0.004 ng of Bet v 1 per pollen grain), which represents 13.9% of the soluble protein. These calculations are based on a sample of 20.8 mg of pollen, which contained 2.62 × 10⁶ grains, so that results can be estimated on a per-pollen-grain basis.

To determine whether all of the allergens had diffused from the grains, the extracted pollen was subjected to grinding

Discussion

In this study we have made, for the first time, a direct comparison between the amount of a major birch allergen, Bet v 1, in daily atmospheric samples (detected by a quantitative ELISA of extracts from filters of a high-volume sampler) and the numbers of birch pollen grains in the atmosphere (estimated by using a Burkard volumetric trap). Total daily atmospheric Bet v 1 concentrations are positively correlated with the amount of detected birch pollen. Amounts of Bet v 1 varied up to 1.5 ng/m³

Acknowledgements

We thank Dr. Bernhard Weber (Allergopharma J. Ganzer KG) for providing antibodies and Bet v 1 standards.

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This study sought to extend the allergen-specific antibody approach and demonstrate that a combination of mAbs targeting Bet v 1, the immunodominant and most abundant allergenic protein in birch pollen, can prevent the birch allergic response.
Bet v 1–specific mAbs, REGN5713, REGN5714, and REGN5715, were isolated using the VelocImmune platform. Surface plasmon resonance, x-ray crystallography, and cryo-electron microscopy determined binding kinetics and structural data. Inhibition of IgE-binding, basophil activation, and mast cell degranulation were assessed via blocking ELISA, flow cytometry, and the passive cutaneous anaphylaxis mouse model.
REGN5713, REGN5714, and REGN5715 bind with high affinity and noncompetitively to Bet v 1. A cocktail of all 3 antibodies, REGN5713/14/15, blocks IgE binding to Bet v 1 and inhibits Bet v 1– and birch pollen extract–induced basophil activation ex vivo and mast cell degranulation in vivo. Crystal structures of the complex of Bet v 1 with immunoglobulin antigen-binding fragments of REGN5713 or REGN5715 show distinct interaction sites on Bet v 1. Cryo-electron microscopy reveals a planar and roughly symmetrical complex formed by REGN5713/14/15 bound to Bet v 1.
These data confirm the immunodominance of Bet v 1 in birch allergy and demonstrate blockade of the birch allergic response with REGN5713/14/15. Structural analyses show simultaneous binding of REGN5713, REGN5714, and REGN5715 with substantial areas of Bet v 1 exposed, suggesting that targeting specific epitopes is sufficient to block the allergic response.

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^☆: Professor R. Bruce Knox passed away suddenly on August 30, 1997. The remaining authors wish to dedicate this paper to his memory.

^☆☆: From the Pollen and Allergen Research Group, School of Botany, The University of Melbourne, Parkville, Victoria.

^★: Supported in part by the Swiss National Science Foundation, the Australian Research Council, and the Australian National Health and Medical Research Council.

^★★: Reprint requests: Cenk Suphioglu, PhD, Pollen and Allergen Research Group, School of Botany, The University of Melbourne, Parkville, Victoria 3052, Australia.

^♢: 1/1/84054

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Concentrations of the major birch tree allergen Bet v 1 in pollen and respirable fine particles in the atmosphere☆,☆☆,★,★★,♢

Abstract

Section snippets

Quantification of Bet v 1

Concentration of Bet v 1 in birch pollen grains

Discussion

Acknowledgements

J Allergy Clin Immunol

Lancet

Anal Biochem

IgE and IgG antibodies of patients with allergy to birch pollen as tools to define the allergen profile of Betula verrucosa

Allergy

Immunogold electron microscopy of soluble protein: localization of Bet v 1 major allergen in ultra-thin sections of birch pollen after anhydrous fixation techniques

J Histochem Cytochem

Seasonal distribution of pollen in the atmosphere of Melbourne, an airborne pollen calendar

Aerobiologia

Deposition of inhaled pollen extract in human airways

N Engl J Med