한국식품과학회지 (Korean Journal of Food Science and Technology)

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

Listeria monocytogenes 신속 검출을 위한 면역크로마토그래피법의 개발

Development of Immunochromatography for the Rapid Detection of Listeria monocytogenes

  • 최진길 (경상대학교 대학원 응용생명과학부(BK21)) ;
  • 심원보 (경상대학교 대학원 응용생명과학부(BK21)) ;
  • 제정현 (경상대학교 대학원 응용생명과학부(BK21)) ;
  • 김지영 (경상대학교 대학원 응용생명과학부(BK21)) ;
  • 이규호 (한국외국어대학교 환경학과) ;
  • 김민곤 (한국생명공학연구원) ;
  • 하상도 (중앙대학교 식품공학과) ;
  • 김근성 (중앙대학교 식품공학과) ;
  • 김광엽 (충북대학교 식품공학과) ;
  • 김철호 (성균관대학교 생명과학과) ;
  • 정덕화 (경상대학교 대학원 응용생명과학부(BK21))
  • Choi, Jin-Gil (Division of Applied Life Science(BK21), Graduate School of Gyeongsang National University) ;
  • Shim, Won-Bo (Division of Applied Life Science(BK21), Graduate School of Gyeongsang National University) ;
  • Je, Jung-Hyun (Division of Applied Life Science(BK21), Graduate School of Gyeongsang National University) ;
  • Kim, Ji-Young (Division of Applied Life Science(BK21), Graduate School of Gyeongsang National University) ;
  • Lee, Kyu-Ho (Environmental Science, HanKuk University of Foreign Studies) ;
  • Kim, Min-Gon (Korea Research Institute of Bioscience and Biotechnology) ;
  • Ha, Sang-Do (Department of Food Science & Technology, Chung-Ang University) ;
  • Kim, Keun-Sung (Department of Food Science & Technology, Chung-Ang University) ;
  • Kim, Kwang-Yup (Department of Food Science & Technology, Chung-Ang University) ;
  • Kim, Cheol-Ho (Department of Biological Sciences, Sungkyunkwan University) ;
  • Chung, Duck-Hwa (Division of Applied Life Science(BK21), Graduate School of Gyeongsang National University)
  • 발행 : 2007.06.30
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초록

본 연구에서는 L. monocytogenes를 신속하게 분석할 수 있는 면역크로마토그래피법을 개발하고자 하였다. 먼저 L. monocytogenes에 대한 단크론성 항체(FKLM-3B12-37)를 개발하여 직경 40nm로 제작된 colloidal gold와 합성한 후 conjugate pad에 처리하였고 nitrocellulose membrane 상의 test line과 control line에 단크론성 항체와 anti-mouse IgG를 각각 처리하여 면역크로마토그래피법을 개발하였다. 개발된 면역크로마토그래피법의 검출한계는 $10^{5}$ cell/mL 수준까지 검출이 가능했으며 다른 식중독 세균과는 교차 반응이 없는 것으로 나타났다. 임으로 오염시킨 식품시료의 경우 증균과정 없이 $10^{4}$ cell/25g까지 바로 분석하는데 어려움이 있으며 24시간의 증균과정을 거친 후에는 $10^{4}$ cell/25g까지 검출할 수 있었다. 본 연구에서 개발된 면역크로마토그래피법은 24시간의 증균 만으로 식품에 오염되어있는 L. monocytogenes를 검출할 수 있는 것으로 사료되었고, 기존의 식중독균 진단방법에 비해 신속하고 간편하게 그 결과를 확인할 수 있었다.

The objective of this study was the development of immunochromatography (ICG) for the rapid and accurate detection of Listeria monocytogenes. Here, monoclonal antibodies (MAb) were conjugated with 40 nm colloidal gold particles, where the conjugate was used as the detection reagent in the ICG. The ICG was composed of three pads (sample, conjugate, and absorbance pads) and one nitrocellulose membrane. The colloidal gold-MAb conjugate was applied to the conjugate pad, and the test line and control line on the membrane were treated with MAb (FKLM-3BI2-37) and anti-mouse IgG, respectively. The detection limit of the ICG was $10^{5}$ cell/mL and it showed no cross-reaction to food borne pathogens. We inoculated meat and lettuce samples with various counts of L. monocytogenes, and analyzed them by ICG. All the inoculated meat samples gave positive results after enrichment for 24 h in LEB. These results indicate that ICG was able to serve as a primary screening tool for L. monocytogenes in various foods and agricultural products within 20 min after enrichment.

키워드

참고문헌

  1. Bahk JR, Marth EH. Listeriosis and Listeria monocytogenes. Korean J. Appl. Microbiol. Bioeng. 17: 634-644 (1989)
  2. Farber JM, Johnston MA, Purvis D, Loit A. Surveillace of soil and semi-soft cheese or the presence of Listeria spp. Int. J. Food Microbiol. 5: 157-163 (1987) https://doi.org/10.1016/0168-1605(87)90033-X
  3. Fleming DW, Cochi SL, MacDonald KL, Brondum J, Hayes PS, Plikaytis BD, Holmes MB, Audurier A, Broome CV, Reingold AL. Pasteurized milks a vehicle of infection in an outbreak of listeriosis. New Engl. J. Med. 312: 404-407 (1985) https://doi.org/10.1056/NEJM198502143120704
  4. Farber JM, Peterkin PI. Listeria monocytogenes, a food-borne pathogen. Microbiol. Rev. 55: 476-511 (1991)
  5. Sewell AM, Warburton DW, Boville A, Daley EF, Mullen K. The development of an efficient and rapid enzyme linked fluorescent assay method for the detection of Listeria spp. from foods. Int. J. Food Microbiol. 81: 123-129 (2003) https://doi.org/10.1016/S0168-1605(02)00221-0
  6. Kerdahi KF, Istafanos PF. Rapid determination of Listeria monocytogenes by automated enzyme-linked immunoassay and nonradioactive DNA probe. J. Assoc. Off. Ana. Chem. Int. 83: 86-88 (2000)
  7. Hahm BK, Bhunia AK. Effect of environmental stresses on antibody-based detection of Escherichia coli O157:H7, Salmonella enterica serotype Enteritidis and Listeria monocytogenes. J. Appl. Microbiol. 100: 1017-1027 (2006) https://doi.org/10.1111/j.1365-2672.2006.02814.x
  8. Chiao DJ, Shyu RH, Hu CS, Chiang HY, Tang SS. Colloidal gold-based immunochromatographic assay for detection of botulinum neurotoxin type B. J. Chromatogr. B 809: 37-41 (2004) https://doi.org/10.1016/j.jchromb.2004.05.033
  9. Paek SH, Lee SH, Cho JH, Kim YS. Development of rapid one-step immunochromatographic assay. Methods 22: 53-60 (2000) https://doi.org/10.1006/meth.2000.1036
  10. Jung BY, Jung SC, Kim JM. Development of the immunochromatographic strip for the rapid detection of Listeria spp. Korean J. Vet. Res. 45: 169-177 (2005)
  11. Jung BY, Jung SC. Development of the rapid detection kit for Salmonella spp. using immunochromatographic assay. Korean J. Vet. Res. 45: 191-197 (2005)
  12. Kohler G, Milstein C. Continuous cultures of fused cells producing antibody of predefined specificity. Nature 256: 495-501 (1975) https://doi.org/10.1038/256495a0
  13. Mckearn TJ, Weiss A, Stuart FP, Fitch FW. Selective suppression of humoral and cell-mediated immune responses to rat alloantigens by monoclonal antibodies produced by hybridoma cell lines. Transplant. P. 11: 932-935 (1979)
  14. Geoghegan WD, Ackerman GA. Adsorption of horseradish peroxidase, ovomucoid and anti-immunoglobulin to colloidal gold for the indirect detection of concanavalin A, wheat germ agglutinin and goat anti-human immunoglobulin G on cell surfaces at the electron microscopic level: a new method, theory and application. J. Histochem. Cytochem. 25: 1187-1200 (1977) https://doi.org/10.1177/25.11.21217
  15. Frens G. Preparation of gold dispersions of varying particle size: controlled nucleation for the regulation of the particle size in monodisperse gold suspensions. Nature-Phys. Sci. 241: 20-22 (1973) https://doi.org/10.1038/physci241020a0
  16. Siragusa GR and Johnson MG. Monoclonal antibody specific for listeria monocytogenes, Listeria innocua, Listeria welshimeri. Appl. Environ. Microb. 56: 1897-1904 (1990)
  17. Kim SH, Park MK, Kim JY, Chuong PD, Lee YS, Yoon BS, Hwang KK, Lim YK Development of a sandwich ELISA for the detection of Listeria spp. using specific flagella antibodies. J. Vet. Sci. 6: 41-46 (2005)