Korean Journal of Food Science and Technology (한국식품과학회지)

Korean Society of Food Science and Technology (한국식품과학회)
권호 표지
DOI QR코드

DOI QR Code

Development of Sandwich ELISA for the Detection of Mackerel in Processed Foods

가공식품 중 고등어의 검출을 위한 ELISA의 개발

  • Received : 2011.04.26
  • Accepted : 2011.11.16
  • Published : 2012.02.29
Download PDF
⟨ Previous Next ⟩

Abstract

There have been few studies related to ELISA for mackerel. In this study we developed a sandwich ELISA for mackerel in processed foods using rabbit polyclonal antibodies against mackerel parvalbumin, the major allergen of mackerel and heat-stable protein. The parvalbumin was purified by ammonium sulfate precipitation and Sephadex G-50 column chromatography. From the standard ELISA curves, the detection limit of parvalbumin was 3 ng/mL and the detection range of mackerel was 5-5,000 ${mu}g/mL$. We further investigated the cross-reactivity of the antibodies toward mackerel, mackerel pike, salmon, flatfish, armorclad rockfish, cod fish, squid, shrimp, blue crab, and lobster. The antibodies were specific for mackerel only. The mean assay recoveries in cooked cream soup, baby food, sausage, and sauce spiked with 0.01 to 0.3% mackerel were 104, 101, 54, and 0%, respectively. In sample tests of 16 commercial items, the qualitative coincidence ratio of assay result and indication was 75%.

기존에 생선 검출을 위한 ELISA 개발은 여러 편 보고된 바 있으나, 고등어만을 특이적으로 검출하는 방법에 관한 보고는 거의 없었다. 본 연구에서는 고등어 parvalbumin에 대한 토끼 항체를 이용하여 가공식품 중 고등어의 검출을 위한 ELISA를 개발 하였다. 암모늄침전법과 Sephadex G-50 column chromatography를 이용하여 열에 안정한 12 kDa의 고등어 parvalbumin을 분리, 정제하였다. 여기에서 얻어진 parvalbumin을 토끼에게 면역하여 생산한 항 parvalbumin 항체 및 특이항체-HRP 복합물을 이용하여 샌드위치 ELISA(sELISA)의 조건을 확립하였다. 이때 parvalbumin 의 검출한계는 3 ng/mL이고, 고등어(생살)의 검출범위는 5-5,000 ${mu}g/mL$이었다. 이어서. 광어, 우럭, 오징어, 꽃게, 꽁치, 대구, 새우, 연어, 바다가재, 고등어에 대한 교차반응을 조사한 결과 고등어에 대한 반응성은 월등히 높았으나 다른 수산식품에서는 반응성이 거의 없거나 매우 낮게 나타나 고등어에 대한 특이성이 매우 높은 항체임을 알 수 있었다. 또한 분석시 시료의 열 안정성은 $100^{\circ}C$까지 양호하였으며 크림스프, 이유식, 소시지, 소스에 대한 spike test(0.01-0.3%)에서 분석회수율은 각각 104, 101, 54, 0%로 나타났다. sELISA에 의하여 16점의 시판시료 중 고등어의 함유 유무를 조사한 결과, 정성적으로 표시와 일치하는 것은 75%이었다. 그러므로 본 연구에서 개발한 sELISA는 일부 한계가 있기는 하지만, 가공식품 중 고등어를 정성적으로 검출하는데는 효과적으로 활용할 수 있을 것이다.

Keywords

References

  1. Lee HK, Song WJ, Ha TU. A single or combined effects of IL-4 and other various cytokines on IgE production of human tonsillar mononuclear cells. Korean J. Immunol. 17: 193-201 (1995)
  2. Okunuki H, Teshima R, Sakushima J, Akiyama H, Goda Y, Toyoda M, Sawada JI. Induction of active systemic anaphylaxis by oral sensitization with ovalbumin in mast-cell-deficient mice. Immunol. Lett. 74: 233-237 (2000) https://doi.org/10.1016/S0165-2478(00)00264-9
  3. Broide DH. Molecular and cellular mechanisms of allergic disease. J. Allergy Clin. Immun. 108: S65-S71 (2001) https://doi.org/10.1067/mai.2001.116436
  4. Eggesbø M, Halvorsen R, Tambs K, Botten G. Prevalence of parentally perceived adverse reactions to food in young children. Pediatr. Allergy Immu. 10: 122-132 (1999) https://doi.org/10.1034/j.1399-3038.1999.00022.x
  5. Ostblom E, Lilja G, Pershagen G, van Hage M, Wickman M. Phenotypes of food hypersensitivity and development of allergic diseases during the first 8 years of life. Clin. Exp. Allergy 38: 1325-1332 (2009)
  6. Eckman J, Saini SS, Hamilton RG. Diagnostic evaluation of foodrelated allergic diseases. Allergy Asthma Clin. Immunol. 5: 2 (2009) https://doi.org/10.1186/1710-1492-5-2
  7. Bannon GA. What makes a food protein an allergen? Curr. Allergy Asthm. Rep. 4: 43-46 (2004) https://doi.org/10.1007/s11882-004-0042-0
  8. Pele M, Brohe M, Anklam E, Van Hengel AJ. Peanut and hazelnut traces in cookies and chocolates: Relationship between analytical results and declaration of food allergens on product labels. Food Addit. Contam. 24: 1334-1344 (2007) https://doi.org/10.1080/02652030701458113
  9. Taylor SL, Hefle SL. Food allergen labeling in the USA and Europe - Review. Curr. Opin. Allergy Cl. Immunol. 6: 186-190 (2006) https://doi.org/10.1097/01.all.0000225158.75521.ad
  10. Korea Food and Drug Administration. Labeling Standards for Foods etc. KFDA Notification No. 2010-97 (Amendment on Dec. 30, 2010) Available from: http://www.kfda.go.kr/index.kfda?x= 7&searchkey=title:contents&mid=92&searchword=식품등의 표시 기준&y=4&division=&pageNo=1&seq=3362&cmd=v. Accessed April 25, 2011.
  11. Kirsch S, Fourdrilis S, Dobson R, Scippo ML, Maghuin-Rogister G, De Pauw E. Quantitative methods for food allergens: A review. Anal. Bioanal. Chem. 395: 57-67 (2009) https://doi.org/10.1007/s00216-009-2869-7
  12. Schubert-Ullrich P, Rudolf J, Ansari P, Galler B, Fhrer M, Molinelli A, Baumgartner S. Commercialized rapid immunoanalytical tests for determination of allergenic food proteins: An overview. Anal. Bioanal. Chem. 395: 69-81 (2009) https://doi.org/10.1007/s00216-009-2715-y
  13. Niemann L, Taylor SL, Hefle SL. Detection of walnut residues in foods using an enzyme-linked immunosorbent assay. J. Food Sci. 74: T51-T57 (2009) https://doi.org/10.1111/j.1750-3841.2009.01214.x
  14. Scaravelli E, Brohe M, Marchelli R, van Hengel AJ. The effect of heat treatment on the detection of peanut allergens as determined by ELISA and real-time PCR. Anal. Bioanal. Chem. 395: 127-137 (2009) https://doi.org/10.1007/s00216-009-2849-y
  15. Shinobu S, Rieko M, Reiko A, Hiroshi A, Tamio M, Yasuo O. Interlaboratory evaluation of two enzyme-linked immunosorbent assay kits for the determiniation of crustacean protein in processed foods. J. AOAC Int. 91: 123-129 (2008)
  16. Faeste CK, Plassen C. Quantitative sandwich ELISA for the determination of fish in foods. J. Immunol. Methods 329: 45-55 (2008) https://doi.org/10.1016/j.jim.2007.09.007
  17. Chen L, Hefle SL, Taylor SL, Swoboda I, Goodman RE. Detecting fish parvalbumin with commercial mouse monoclonal antifrog parvalbumin IgG. J. Agr. Food Chem. 54: 5577-5582 (2006) https://doi.org/10.1021/jf060291g
  18. Patrick W, Hans S, Angelika P. Competitive indirect ELISA for the determination of parvalbumins from various fish species in food grade fish gelatins and isinglass with parv-19 anti-parvalbumin antibodies. J. Agr. Food Chem. 57: 11328-11334(2009) https://doi.org/10.1021/jf902470e
  19. Gajewski KG, Hsieh YH. Monoclonal antibody specific to a major fish allergen: Parvalbumin. J. Food Prot. 72: 818-825 (2009)
  20. Valenta R, Hayek B, Seiberler S, Bugajska-Schretter A, Niederberger V, Twardosz A, Natter S, Vangelista L, Pastore A, Spitzauer S, Kraft D. Calcium-binding allergens: From plants to man. Int. Arch. Allergy Imm. 117: 160-166 (1998) https://doi.org/10.1159/000024005
  21. Van Do T, Elsayed S, Florvaag E, Hordvik I, Endresen C. Allergy to fish parvalbumins: Studies on the cross-reactivity of allergens from 9 commonly consumed fish. J. Allergy Clin. Immun. 116: 1314-1320 (2005) https://doi.org/10.1016/j.jaci.2005.07.033
  22. Lim DL, Noe KH, Yi FC, Chua KY, Goh DL, Shek LP, Giam YC, Van Bever HPS, Lee BW. Parvalbumin - the major tropical fish allergen. Pediatr. Allergy Immu. 19: 399-407 (2008) https://doi.org/10.1111/j.1399-3038.2007.00674.x
  23. Sletten G, Van Do T, Lindvik H, Egaas E, Florvaag E. Effects of industrial processing on the immunogenicity of commonly ingested fish species. Int. Arch. Allergy Imm. 151: 223-236 (2010) https://doi.org/10.1159/000242360
  24. Kuehn A, Scheuermann T, Hilger C, Hentges F. Important variations in parvalbumin content in common fish species: A factor possibly contributing to variable allergenicity. Int. Arch. Allergy Imm. 153: 359-366 (2010) https://doi.org/10.1159/000316346
  25. Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680-685 (1970) https://doi.org/10.1038/227680a0
  26. Kim HJ, Shon DH, An enzyme-linked immunosorbent assay for detection of cooked goat meat. Korean J. Food Sci. Technol. 32: 538-543 (2000)
  27. Herrmann JE, Hendry RM, Collins MF. Factors involved in enzyme-linked immunoassay of viruses and evaluation of the method for identification of enteroviruses. J. Clin. Microbiol. 10: 210-217 (1979)
  28. Taylor SL, Nordlee JA, Niemann LM, Lambrecht DM. Allergen immunoassays-considerations for use of naturally incurred standards.Anal. Bioanal. Chem. 395: 83-92 (2009) https://doi.org/10.1007/s00216-009-2944-0

Cited by

  1. Development of Sandwich ELISA for the Detection of Pork in Processed Foods vol.47, pp.3, 2015, https://doi.org/10.9721/KJFST.2015.47.3.401