Analytical and Bioanalytical Chemistry

, Volume 410, Issue 12, pp 3017–3023 | Cite as

Development and in-house validation of a rapid and simple to use ELISA for the detection and measurement of the mycotoxin sterigmatocystin

  • Michalina Oplatowska-StachowiakEmail author
  • Claudine Reiring
  • Nermin Sajic
  • Willem Haasnoot
  • Catherine Brabet
  • Katrina Campbell
  • Christopher T. Elliott
  • Martin Salden
Research Paper


Sterigmatocystin (STG) is a highly toxic secondary fungal metabolite structurally closely related to the well-known carcinogenic aflatoxins. Its presence has been reported in grains and grain-based products as well as in other foodstuffs like nuts, green coffee beans, spices, beer and cheese. Due to the lack of suitable data on the occurrence of STG, in 2013, the European Food Safety Authority (EFSA) could not characterise its risk for human health and recommended that more data on STG in food and feed needed to be collected. In order to provide a new tool for the specific detection of STG, a competitive enzyme-linked immunosorbent assay (ELISA) was developed, optimised and validated in this study based on a sensitive monoclonal antibody specific to STG with no cross-reactivity with aflatoxins. The sample preparation method for rice, wheat and maize was based on a modified QuEChERS (quick, easy, cheap, effective, rugged and safe) approach. The assay was validated for the detection of STG in rice, wheat and maize in accordance with the guidelines for validation of semi-quantitative screening methods included in Commission Regulation (EU) 519/2014. The screening target concentration (STC) was set at 1.5 μg/kg. The cutoffs for rice, wheat and maize were 1.2, 1.2 and 1.3 μg/kg and the false suspected rates were 0.34, 1.15 and 0.78%, respectively. Good correlation was found between the results obtained by the STG ELISA and LC-MS/MS method for naturally contaminated rice samples. This validated method can be applied as a sensitive and high-throughput screening for the presence of STG in a range of agricultural commodities.

Graphical abstract

A new enzyme-linked immunosorbent assay based on an antibody specific to sterigmatocystin for the detection of this mycotoxin in corn, wheat and rice.


Enzyme-linked immunosorbent assay Food safety Immunoassay Mycotoxin 



The authors would like to thank Lucia Streppel and Piet van Wichen from EuroProxima for their support of this research.

Funding information

This project has received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No 655119.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Research involving animals

Animal experiments were performed in accordance with the UK Animals Scientific Procedures Act 1986 under the license PPL2756 issued on the 12/02/2014 by the Department of Health, Social Services and Public Safety for Northern Ireland. The study received approval from the Queens University Belfast Animal Welfare and Ethical Review Body on 09/01/2014.


  1. 1.
    Bhat R, Rai RV, Karim AA. Mycotoxins in food and feed: present status and future concerns. Compr Rev Food Sci F. 2010;9:57–81.CrossRefGoogle Scholar
  2. 2.
    European Commission. Commission regulation (EC) no 1881/2006 of 19 December 2006 setting maximum levels for certain contaminants in foodstuffs. Off J Eur Union. 2006;L364:5–23.Google Scholar
  3. 3.
    European Commission. Commission recommendation of 17 August 2006 on the presence of deoxynivalenol, zearalenone, ochratoxin A, T-2 and HT-2 and fumonisins in products intended for animal feeding (2006/576/EC). Off J Eur Union. 2006;L229:7–9.Google Scholar
  4. 4.
    European Commission. Commission Recommendation of 27 March 2013 on the presence of T-2 and HT-2 toxin in cereals and cereal products (2013/165/EU). Off J Eur Union. 2013;L91:12–5.Google Scholar
  5. 5.
    International Agency for Research on Cancer (IARC). Chemical agents and related occupations. A review of human carciongens. In: IARC Monographs on the Evaluation of Carcinogenic Risks to Humans. 2012;100F:225–244.Google Scholar
  6. 6.
    European Food Safety Authority (EFSA). Scientific opinion on the risk for public and animal health related to the presence of sterigmatocystin in food and feed. EFSA J. 2013;11(6):3254.CrossRefGoogle Scholar
  7. 7.
    Veršilovskis A, De Saeger S. Sterigmatocysin: occurrence in foodstuffs and analytical methods—an overview. Mol Nutr Food Res. 2010;54:136–47.CrossRefGoogle Scholar
  8. 8.
    International Agency for Research on Cancer (IARC). Some naturally occurring substances. In: IARC Monographs on the Evaluation of Carcinogenic Risks to Humans. Summaries and Evaluations. 1987;10:72.Google Scholar
  9. 9.
    Stroka J, Dasko L, Spangenberg B, Anklam E. Determination of the mycotoxin, sterigmatocystin, by thin-layer chromatography and reagent-free derivatisation. J Liq Chromatogr Relat Techno. 2004;27:2101–11.CrossRefGoogle Scholar
  10. 10.
    Mol HGJ, Pietri A, MacDonald SJ, Anagnostopoulos C, Spanjer M. Survey on sterigmatocystin in food. EFSA Supporting Publication. 2015;EN-774:56.Google Scholar
  11. 11.
    Veršilovskis A, Bartkevičs V, Miķelsone V. Sterigmatocystin presence in typical Latvian grains. Food Chem. 2008;109:243–8.CrossRefGoogle Scholar
  12. 12.
    Marley E, Brown P, Mackie J, Donnelly C, Wilcox J, Pietri A, et al. Analysis of sterigmatocystin in cereals, animal feed, seeds, beer and cheese by immunoaffinity column clean-up and HPLC and LC-MS/MS quantification. Food Addit Contam A. 2015;32:2131–7.Google Scholar
  13. 13.
    Zhao Y, Huang J, Ma L, Wang F. Development and validation of a simple and fast method for simultaneous determination of aflatoxin B1 and sterigmatocystin in grains. Food Chem. 2017;221:11–7.CrossRefGoogle Scholar
  14. 14.
    Ok HE, Tian F, Hong EY, Paek O, Kim S-H, Kim D, et al. Harmonized collaborative validation of aflatoxins and sterigmatocystin in white rice and sorghum by liquid chromatography coupled to tandem mass spectrometry. Toxins. 2016;8:371.CrossRefGoogle Scholar
  15. 15.
    Biancardi A, Dall’Asta C. Determination of sterigmatocystin in feed by LC-MS/MS. Food Addit Contam. 2015;32:2093–100.Google Scholar
  16. 16.
    Sulyok M, Krska R, Schuhmacher RA. A liquid chromatography/tandem mass spectrometric multi-mycotoxin method for the quantification of 87 analytes and its application to semi-quantitative screening of moldy food samples. Anal Bioanal Chem. 2007;389:1505–23.CrossRefGoogle Scholar
  17. 17.
    Monbaliu S, Van Poucke C, Detavernier C, Dumoultn F, Van Velde MDE, Schoeters E, et al. Occurrence of mycotoxins in feed as analysed by a multi-mycotoxin LC-MS/MS method. J Agr Food Chem. 2010;58:66–71.CrossRefGoogle Scholar
  18. 18.
    Jackson LC, Kudupoje MB, Yiannikouris A. Simultaneous multiple mycotoxin quantification in feed samples using three isotopically labeled internal standards applied for isotopic dilution and data normalization through ultra-performance liquid chromatography/electrospray ionization tandem mass spectrometry. Rapid Commun Mass Sp. 2012;26:2697–713.CrossRefGoogle Scholar
  19. 19.
    Malachová A, Sulyok M, Beltrán E, Berthiller F, Krska R. Optimization and validation of a quantitative liquid chromatography-tandem mass spectrometric method covering 295 bacterial and fungal metabolites including all regulated mycotoxins in four model food matrices. J Chromatogr A. 2014;1362:145–56.CrossRefGoogle Scholar
  20. 20.
    Oplatowska-Stachowiak M, Haughey SA, Chevallier OP, Galvin-King P, Campbell K, Magowan E, et al. Determination of the mycotoxin content in distiller’s dried grain with solubles using a multianalyte UHPLC–MS/MS method. J Agr Food Chem. 2015;63:9441–51.CrossRefGoogle Scholar
  21. 21.
    Kong D, Xie Z, Liu L, Song S, Kuang H, Cui G, et al. Development of indirect competitive ELISA and lateral-flow immunochromatographic assay strip for the detection of sterigmatocystin in cereal products. Food Agr Immunol. 2017;28:260–73.CrossRefGoogle Scholar
  22. 22.
    Li M, Li P, Wu H, Zhang Q, Ma F, Zhang Z, et al. An ultra-sensitive monoclonal antibody-based competitive enzyme immunoassay for sterigmatocystin in cereal and oil products. PLoS One. 2014;9:e106415.CrossRefGoogle Scholar
  23. 23.
    Li S, Chen PY, Marquardt RR, Han Z, Clarke JR. Production of a sensitive monoclonal antibody to sterigmatocystin and its application to ELISA of wheat. J Agr Food Chem. 1996;44:372–5.CrossRefGoogle Scholar
  24. 24.
    European Commission. Commission regulation (EU) 519/2014 of 16 May 2014 amending regulation (EC) no 401/2006 as regards methods of sampling of large lots, spices and food supplements, performance criteria for T-2, HT-2 toxin and citrinin and screening methods of analysis. Off J Eur Union. 2014;L147:29–43.Google Scholar
  25. 25.
    Kononenko GP, Burkin AA, Soboleva NA. Comparative characterization of immune reagents based on hemiacetals of aflatoxin B1 and sterigmatocystine. Appl Biochem Micro. 2002;38:487–92.CrossRefGoogle Scholar
  26. 26.
    Oplatowska-Stachowiak M, Sajic N, Xu Y, Haughey SA, Mooney MH, Gong YY, et al. Fast and sensitive aflatoxin B1 and total aflatoxins ELISAs for analysis of peanuts, maize and feed ingredients. Food Control. 2016;63:239–45.CrossRefGoogle Scholar
  27. 27.
    Köhler G, Milstein C. Continuous cultures of fused cells secreting antibody of predefined specificity. Nature. 1975;256:495–7.CrossRefGoogle Scholar
  28. 28.
    Oplatowska-Stachowiak M, Kleintjens T, Sajic N, Haasnoot W, Campbell K, Elliott CT, et al. T-2 toxin/HT-2 toxin and ochratoxin A ELISAs development and in-house validation in food in accordance with commission regulation (EU) no 519/2014. Toxins. 2017;9:388.CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Michalina Oplatowska-Stachowiak
    • 1
    Email author
  • Claudine Reiring
    • 1
  • Nermin Sajic
    • 1
  • Willem Haasnoot
    • 2
  • Catherine Brabet
    • 3
    • 4
  • Katrina Campbell
    • 5
  • Christopher T. Elliott
    • 5
  • Martin Salden
    • 1
  1. 1.EuroProxima B.V.ArnhemThe Netherlands
  2. 2.RIKILT Wageningen URWageningenThe Netherlands
  3. 3.Centre de Coopération Internationale en Recherche Agronomique pour le Développement (CIRAD), UMR QualiSudMontpellier Cedex 5France
  4. 4.QualiSud, Université Montpellier, CIRAD, Montpellier SupAgroUniversité d’AvignonMontpellierFrance
  5. 5.Institute for Global Food Security, School of Biological SciencesQueen’s University BelfastBelfastUK

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