OWLS detektáláson alapuló immunszenzorok fejlesztése penészgomba eredetű toxinok kimutatására
Title in English
Development of OWLS based immunosensors for detecting toxins produced by moduls
Panel
Chemistry 2
Department or equivalent
Élelmiszer-analitikai Osztály (Central Environmental and Food Science Research Institute)
Participants
Adányiné dr. Kisbocskói, Nóra Trummer, Nikoletta
Starting date
2004-01-01
Closing date
2007-12-31
Funding (in million HUF)
3.926
FTE (full time equivalent)
0.00
state
closed project
Final report
Results in Hungarian
A mikotoxinok gombák másodlagos metabolitjai, amelyek az élelmiszereket és takarmányokat szennyezhetik. Az optikai hullámvezető fénymódus spektroszkópia (OWLS) alkalmazásával direkt és indirekt immunanalitikai eljárást dolgoztunk ki aflatoxin B1 és ochratoxin A meghatározására.
A biológiai molekulákat a szenzor felületén kialakított karboxil-csoportokhoz EDC/NHS technika alkalmazásával rögzítettük. A kialakított rétegeket ill. a rögzítési eljárásokat BSA - anti-BSA molekula párral vizsgáltuk.
A nem-kompetitív (direkt) eljárás esetében a toxin detektálásakor csak 0,1-10 μg/ml méréstartományban kaptunk jeleket. A jelek mindkét esetben nagyon instabilak voltak.
A kompetitív eljárás kidolgozása során az antigén-konjugátummal érzékenyített szenzorral az antitestet tartalmazó szérum optimális hígítását határoztuk meg, ez az aflatoxin esetében 1000x, míg az ochratoxin mérésekor 1.0 μg/ml volt. A kompetitív mérésnél a standardokat az antitest megfelelően hígított oldatával kevertük össze és 1 perces inkubálás után injektáltuk az OWLS rendszerbe. Az optimalizált paramétereket alkalmazva 0,01-10 ng/ml tartományban kapunk lineáris válaszjeleket mindkét esetben. Az indirekt mérési eljárás kidolgozása után különböző élelmiszer és takarmány mintákat vizsgáltunk, az eredmények összhangban voltak a referencia módszerrel mért értékekkel.
Results in English
Mycotoxins are toxic secondary metabolites produced by fungi, and can be present in a wide range of food and feed commodities. The OWLS technique has been applied for developing new method to the detection of Aflatoxin B1 and Ochratoxin A in both competitive and in direct immunoassays.
In order to immobilize the biological molecules on the surface of the sensor, the coupling process by EDC/NHS technique to the carboxyl groups provide on the sensor surface was investigated. To test the layers formed, and the process of immobilization, experiments were undertaken using BSA - anti-BSA model molecule pair.
When using non-competitive method, sensor responses were obtained only at analyte concentrations of 5-10 ng/ml. In both cases, the responses were very unstable. For competitive sensor investigation with the sensitized chip first the optimal dilution rate of antibodies was determined, for the measurement of Aflatoxin B1 and Ochratoxin A the antibody stock solution was diluted to 1000x and to 1.0 μg/ml, respectively. During the competitive measurement standard solutions were mixed with antibodies at the appropriate concentration, the mixture was incubated for 1 min and injected into the OWLS system. The sensitive detection range of competitive detection was between 0,5-10 ng/ml in both cases. After the establishment of the indirect method, different grain and food samples were measured, and the results were in good correlation by those measured with reference method.
Adányi N., Levkovets I.A., Rodriguez-Gil S., Ronald A., Váradi M., Szendrő I.: Development of immunosensor based on OWLS technique for determining Aflatoxin B1 and Ochratoxin A, Biosensors and Bioelectronics 22. 797-802., 2007
Adányi N., Váradi M., Kim N., Szendrő I.: Development of new immunosensors for determination of contaminants in food, Current Applied Physics 6(2) 279-286, 2006
Váradi M.: Biosensor as a new tool in food analysis., Acta Alimentaria 34(3), 209-210, 2005
Váradi M.: Az élelmiszerbiztonság és -minőség kihívásai az élelmiszeranalitikában, Magyar Kémiai Folyóirat 111 (2) 118-123., 2005
Adányi N., Németh E., Halász A., Szendrő I., Váradi M.: Application of electrochemical optical waveguide lightmode spectroscopy (EC-OWLS) for studying the effect of different stress factors on lactic acid bacteria, Anal. Chim. Acta 573-574. 41-47., 2006
Adányi N., Németh E., Halász A., Váradi M., Szendrő I.: Application of electrochemical optical waveguide lightmode spectroscopy (EC-OWLS) for studying the effect of different stress factors on lactic acid bacteria, Int. Conf. on Instrumental Methods of Analysis (Modern Trends and Applications) 2-6 Oct, 2005 Iraklion, Crete Greece, P-I-35., 2005
Váradi M. Adányi, Levkovets I., Székács A., Szendrő I.: Application of optical waveguide lightmode spectroscopy (OWLS) technique for the immunosensing of mycotoxins, Int. Conf. on Instrumental Methods of Analysis (Modern Trends and Applications) 2-6 Oct, 2005 Iraklion, Crete Greece, O-121., 2005
Váradi M., Adányi N., Levkovets I., Székács A., Szendrő I.: Immunosensors based on OWLS technique for the determination of mycotoxins in food., The SAFE Consortium Int. Cong. on Food Safety, 11-14. June 2006. Budapest, Abstr. 2-33. p 56., 2006
Váradi M. (2007): The international regulation of chemical toxicants in food. Codex Alimentarius., in: Food toxicant analysis, techniques, strategies and developments. (Ed. by Pico J.) Elsevier, 1.ed. 1-10., 2007
Németh E., Adányi, N., Halász A.; Váradi M., Szendrő I.: Real-time study of the effect of different stress factors on lactic acid bacteria by electrochemical optical waveguide lightmode spectroscopy, Biomelecular Engineering 24. 631-637., 2007
Váradi M., Adányi N.: Determination of polyphenolic level in wine samples by using biosensors, 5th Int. Conf. on Instrumental Methods of Analysis Modern Trends and Applications Patras, Greece 30Sept. -4.Oct. OR193., 2007
Adányi N., Bukovskaya O., Székács A., Szendrő I., Váradi M.: Determination of vitellogenin content in carp by using direct and competitive immunosensors, 5th Int. Conf. on Instrumental Methods of Analysis Modern Trends and Applications Patras, Greece 30Sept. -4.Oct. OR194., 2007
Adányi N., Majer-Baranyi K., Székács A., Szendrő I., Váradi M.: Development of immunosensors based on optical waveguide lightmode spectroscopy (OWLS) technique for determining mycotoxins in food, 3rd Int. Symp. on Recent advances in food analysis 7-9. Nov. Prague, Czech Republic. L39. p. 80., 2007
