Abstract
Allergy diagnostics is being transformed by the advent of in vitro IgE testing using purified allergen molecules, combined with multiplex technology and biosensors, to deliver discriminating, sensitive, and high-throughput molecular diagnostics at the point of care. Essential elements of IgE molecular diagnostics are purified natural or recombinant allergens with defined purity and IgE reactivity, planar or bead-based multiplex systems to enable IgE to multiple allergens to be measured simultaneously, and, most recently, nanotechnology-based biosensors that facilitate rapid reaction rates and delivery of test results via mobile devices. Molecular diagnostics relies on measurement of IgE to purified allergens, the “active ingredients” of allergenic extracts. Typically, this involves measuring IgE to multiple allergens which is facilitated by multiplex technology and biosensors. The technology differentiates between clinically significant cross-reactive allergens (which could not be deduced by conventional IgE assays using allergenic extracts) and provides better diagnostic outcomes. Purified allergens are manufactured under good laboratory practice and validated using protein chemistry, mass spectrometry, and IgE antibody binding. Recently, multiple allergens (from dog) were expressed as a single molecule with high diagnostic efficacy. Challenges faced by molecular allergy diagnostic companies include generation of large panels of purified allergens with known diagnostic efficacy, access to flexible and robust array or sensor technology, and, importantly, access to well-defined serum panels form allergic patients for product development and validation. Innovations in IgE molecular diagnostics are rapidly being brought to market and will strengthen allergy testing at the point of care.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Papers of particular interest, published recently, have been highlighted as: • Of importance •• Of major importance
Hamilton RG. Clinical laboratory assessment of immediate-type hypersensitivity. J Allergy Clin Immunol. 2010;125:S284–96.
Hamilton RG, Williams PB. Human IgE antibody serology: a primer for the practicing North American allergist/immunologist. J Allergy Clin Immunol. 2010;126:33–8.
Asarnoj A et al. Peanut component Ara h 8 sensitization and tolerance to peanut. J Allergy Clin Immunol. 2012;130:468–72.
Beyer K et al. Predictive values of component-specific IgE for the outcome of peanut and hazelnut food challenges in children. Allergy. 2015;70:90–8.
Chapman MD, Pomés A, Breiteneder H, Ferreira F. Nomenclature and structural biology of allergens. J Allergy Clin Immunol. 2007;119:414–20.
Thomas WR. The advent of recombinant allergens and allergen cloning. J Allergy Clin Immunol. 2011;127:855–9.
Erwin EA et al. Quantitative measurement of IgE antibodies to purified allergens using streptavidin linked to a high-capacity solid phase. J Allergy Clin Immunol. 2005;115:1029–35.
Hiller R et al. Microarrayed allergen molecules: diagnostic gatekeepers for allergy treatment. FASEB J. 2002;16:414–6.
Wohrl S et al. The performance of a component-based allergen-microarray in clinical practice. Allergy. 2006;61:633–9.
Canonica GW et al. A WAO-ARIA-GA2LEN consensus document on molecular-based allergy diagnostics. World Allergy Organ J. 2013;6:17. This paper is a thorough review by leaders of the allergy field on molecular-based allergy diagnostics, focusing on ISAC for specific IgE testing in vitro. The review describes many clinical studies that have used this technology and provides recommendations for future studies on molecular diagnostics.
Lupinek C et al. Advances in allergen-microarray technology for diagnosis and monitoring of allergy: the MeDALL allergen-chip. Methods. 2014;66:106–19.
Pokoj S et al. Pichia pastoris is superior to E. coli for the production of recombinant allergenic non-specific lipid-transfer proteins. Protein Expr Purif. 2010;69:68–75.
Fenaille F et al. Mass spectrometric investigation of molecular variability of grass pollen group 1 allergens. J Proteome Res. 2009;8:4014–27.
Seppala U et al. Absolute quantification of allergens from complex mixtures: a new sensitive tool for standardization of allergen extracts for specific immunotherapy. J Proteome Res. 2011;10:2113–22.
Chapman MD, Briza P. Molecular approaches to allergen standardization. Curr Allergy Asthma Rep. 2012;12:478–84.
Zybailov B et al. Statistical analysis of membrane proteome expression changes in Saccharomyces cerevisiae. J Proteome Res. 2006;5:2339–47.
Nilsson OB et al. Designing a multimer allergen for diagnosis and immunotherapy of dog allergic patients. PLoS ONE. 2014;9:e111041. The study is an excellent paper on how to use recombinant DNA technology to fuse multiple allergens into one for diagnostic purposes. This is an innovative strategy which could simplify molecular diagnostics. The paper lays out a comprehensive approach to ensuring correct folding of dog allergens in the multimer and to assessing IgE reactivity.
Vieths S et al. Establishment of recombinant major allergens Bet v 1 and Phl p 5a as Ph. Eur. reference standards and validation of ELISA methods for their measurement. Results from feasibility studies. Pharmeur Bio Sci Notes. 2012;2012:118–34.
Incorvaia C et al. A pitfall to avoid when using an allergen microarray: the incidental detection of IgE to unexpected allergens. J Allergy Clin Immunol Pract. 2014. doi:10.1016/j.jaip.2014.09.020. This rostrum article cautions against interpreting molecular diagnostic data out of context of the clinical history and brings up important questions about how false-positive IgE results should be interpreted.
King EM, Vailes LD, Tsay A, Satinover SM, Chapman MD. Simultaneous detection of total and allergen-specific IgE by using purified allergens in a fluorescent multiplex array. J Allergy Clin Immunol. 2007;120:1126–31.
Pomponi D et al. Allergen micro-bead array for IgE detection: a feasibility study using allergenic molecules tested on a flexible multiplex flow cytometric immunoassay. PLoS ONE. 2012;7, e35697.
Feyzkhanova GU et al. Development of hydrogel biochip for in vitro allergy diagnostics. J Immunol Methods. 2014;406:51–7. This paper presents initial data using three-dimensional hydrogel chips with 1-nl volumes to measure allergen-specific IgE. The chip results correlate with an IgE ELISA for allergen extracts, though the data on purified allergens was somewhat limited. Nonetheless, the paper is a useful application of this promising nanotechnology.
Joshi AA, Peczuh MW, Kumar CV, Rusling JF. Ultrasensitive carbohydrate-peptide SPR imaging microarray for diagnosing IgE mediated peanut allergy. Analyst. 2014;139:5728–33. This technology may have applications for measuring IgE to carbohydrate epitopes on allergens. The SPR technology is highly sensitive and fast, but it is unclear how SPR can be used in the clinic.
Mungroo NA, Neethirajan S. Biosensors for the detection of antibiotics in poultry industry—a review. Biosensors (Basel). 2014;4:472–93. The paper is a comprehensive review of biosensors which covers different biosensing techniques and applications of SPR technology in industry.
Platt GW et al. Allergen immobilisation and signal amplification by quantum dots for use in a biosensor assay of IgE in serum. Biosens Bioelectron. 2014;52:82–8. This paper provides a detailed discussion of the merits of copolymer immobilized allergen (casein) for specific IgE detection. Streptavidin-conjugated quantum dots provide optimal signal enhancement and allow dry-form storage which is particularly useful for assay automation.
Chapman MD. Lateral flow tests for allergy diagnosis: point-of-care or point of contention? Int Arch Allergy Immunol. 2010;152:301–2.
van Ree R et al. The CREATE project: development of certified reference materials for allergenic products and validation of methods for their quantification. Allergy. 2008;63:310–26.
Chapman MD et al. The European Union CREATE Project: a model for international standardization of allergy diagnostics and vaccines. J Allergy Clin Immunol. 2008;122:882–9.
Gadermaier G, Hauser M, Ferreira F. Allergens of weed pollen: an overview on recombinant and natural molecules. Methods. 2014;66:55–66. The paper is an excellent and up-to-date review of allergens of weed pollen from around the world, both natural and recombinant, from the leading experts in the molecular biology of weed allergens. The review covers the molecular biology of pectate lyases, defensins, lipid transfer proteins, profilins and polcalcins: important molecules for diagnostics.
Thorpe SJ et al. The 3rd International Standard for serum IgE: international collaborative study to evaluate a candidate preparation. Clin Chem Lab Med. 2014;52:1283–9.
Acknowledgments
This work is supported in part by the National Institute of Allergy and Infectious Diseases of the National Institutes of Health under Award Number R01AI077653. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. We are grateful to Drs. Nicolas Durand and Iwan Merki (Abionic) for the informative discussions about nanotechnology for IgE detection and for providing the images used in Fig. 2.
Compliance with Ethics Guidelines
ᅟ
Conflict of Interest
Martin D. Chapman declares that he is a founder and co-owner of Indoor Biotechnologies and was recently appointed to the Board of Directors of Abionic SA, Lausanne, Switzerland. Sabina Wuenschmann, Eva King, and Anna Pomés declare that they are employees of Indoor Biotechnologies.
Human and Animal Rights and Informed Consent
This article does not contain any studies with human or animal subjects performed by any of the authors.
Author information
Authors and Affiliations
Corresponding author
Additional information
This article is part of the Topical Collection on Immunologic/Diagnostic Tests in Allergy
Rights and permissions
About this article
Cite this article
Chapman, M.D., Wuenschmann, S., King, E. et al. Technological Innovations for High-Throughput Approaches to In Vitro Allergy Diagnosis. Curr Allergy Asthma Rep 15, 36 (2015). https://doi.org/10.1007/s11882-015-0539-8
Published:
DOI: https://doi.org/10.1007/s11882-015-0539-8