Quantification of Genetically Modified Soy Proteins in Fresh Soybean Curd by Antigen-coated Plate ELISA

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(1)KOREAN J. FOOD SCI. TECHNOL. Vol. 36, No. 5, pp. 828~832 (2004). ©The Korean Society of Food Science and Technology. Î²/;»j Ï L¦ 7~ F*¶ Ò &LWî ªC ;*ÁV;8ÁBãÁË?Áf;ÁVÿ^* ]L wÏ>z. Quantification of Genetically Modified Soy Proteins in Fresh Soybean Curd by Antigen-coated Plate ELISA Mee-Hyun Jung, Hyung-Ki Bae, Kyung-Mi Kim, In-Suk Jang, Eun-Jung Ko, and Dong-Ho Bae* Department of Applied Biology and Chemistry, KonKuk University Enzyme-linked immuno sorbent assay (ELISA) was applied to quantify soy protein in fresh soybean curd (bean curd) produced by combination of genetically modified (GM) and genetically not modified (non-GM) soybeans. Antibodies against 113 and 24 kDa proteins, which appeared only in non-GM bean curd (specific band), and in both non-GM and GM bean curds (non-specific band) based on SDS-PAGE results, were prepared by immunization to rabbit. Through ELISA using either antibody, GM bean curd protein content was determined at dilution rates of 10−1-10−6. Standard curve showing relationship between ELISA optical density and non-GM protein content was produced using antibody against 113 kDa protein at protein dilution between 10−7 to 10−6, highly antigen content-dependent dilution. Bean curd prepared by random combinations of GM and non-GM soybeans were analyzed by ELISA, and standard curve was produced. Results reveal non-GM protein content of bean curd could be quantified with higher than 93% accuracy. Key words: GM soybean curd, ELISA, quantification of GMO. B. . VB ê, ÖÒ¾¢öBê ²j¶ 5 "Ú~ ºö V¢ GM ³Öbö & ²j¶~ r ²Òf F~ ¶F¢ Ë~ º NööB GM Bê¢ ~ ® (3). *Ò Úö B &Ïb FÛ>º &v 7 ôf ·f ¦V >« B ©, >«&v~ &¦ªf F*¶ ÒB ©b ºG> ® . ²j¶ f GM ®~ *W Ã«B : ìb¾ ï ®n6b GM öò~ ÒÏj Vb~ ® . ö V¢ Ú~ v¦ BëÚöBº GM &v¢ Ò Ï~æ p~rj ;~ ®b¾ &® Ú~ b«B GM öòræ ªC > ®º O»f jç ;ã>Ú ®æ pf. ; . *Ò ÒÏ> ®º &' ªC»f PCR(F*¶Ã>w) j Ï~ Ò F*¶¢ ¦Â~º O» . ò¦V ºÂ "; DNA¢ Ã~º O» PCR ;ï»f ò 7 ~ ÚÒW F*¶f Ò F*¶~ &ö V ªN ?. &;~º ãÖöò &Ë O» . ¾ >~ Ò Ú &®j öò ~º ¢>&®öBº ' öò~ ª N ¢ PCR~ ";VËj ~º DNA^& ¢;~æ p V r^ö PCR ;ï»b ò 7~ GM ³Öb~ b«Nj G;~º öº ÚJæ ® (4,5). 6 · «~~ GM ³Öb BB> ®º rö PCR ¦ÂVF~ &Ë ^ B6f ê« F*¶~ ;& ìº ©f ¦Â ®&Ë~ &~ jÏj jº ~ BWj æîº ©b (5). *Ò PCR ;ï»~ &nb Î²G;»(enzyme. F*¶ Ò ®~ n*W ^B& " ² ¢¦ > ®º ©f V~ G«¾ vVöB ¢ÚÆ > ìº « ~ F*¶¢ ã« ³Öb j ç7 S º 6 . 6 , F*¶ Ò VFöB " ÒÏ>º ©f B ÚW F*¶, J¾Æ B ÚW~ {Öö V B ~ò ~ ÚJæ rJ^ B ÚW F*¶~ Ïf B Ú W¢º F*¶ Ò(genetically modified, GM) ®~ 6. n*W^B¢ ¢V~ ® (1). F*¶ Ò ³·b~ B' BB*j 1986j ¦V 1997jö Vræ 45BöB 10«ö º ßW j <º 60«~ ³·b BB>Ú ® . B.B ÚW ³· b 1996jêö öB &v(Monsanto), Fj(ArEvo, Monsanto), K>>(Novartis, Dekalb, AgrEvo )& BB>î 590 « ÒV>Ú *Ú F*¶ Ò ³·b~ 27.7%¢ Næ~ ® (2). 1997j EUöB F*¶ Ò ®ö & ^B& B *Corresponding author: Dong-Ho Bae, Department of Applied Biology and Chemistry, Konkuk University, 1 Hwayang-dong, Gwangjin-gu, Seoul 143-701, Korea Tel: 82-2-450-3756 Fax: 82-2-456-7011 E-mail: [email protected] 828.

(2) Î²G;»j Ï v¦ 7~ F*¶ Ò &vWî ªC. linked immuno-sorbent assay, ELISA) B> ® (6,7). ELISAº Ú~ ßWj Ï~º O»b ¦Â 6ê& ¸ òªC * b ôf ò¢ ÿö ªC > ® º Ë6 ® . Kwak (7)f B.Bö & &W F*¶ CP4 EPSPSö & ßÚ¢ Ï~ ELISA»b & vò~ F*¶ Ò ¦¢ 6 > ®rj ~& . 6, Seo(8)º ' O»j Ï~ F*¶ · ~ ; W 5 ;ï' ¦Ò& &Ëj ~& . ¾ v¦f ? f &®öB öò~ GMO 6êö & º B : ì . 6, &® Úö GMOf F*¶ Ò Úææ pf öò(non-GMO)& b«>Ú ®j ãÖf CP4 EPSPS& jò ÒF*¶& Ú®j röº ¢ 6 &k îN>Ú ®æ p . V¢B öBº F*¶ Ò "º ³·b &v¢ Ï &® 7ö &' B®¢ > ®º v¦ ¢ ò ~ ELISA¢ Û .Ó¦Ò F*¶ Ò ¦¢ ¦Ò~¶ ~&b, GM &vf non-GM &v& b« B ãÖö b«jNræê G; > ®º &ËWj ¦Æ ~¶ ~& .. Òò 5 O» Òò 5 ¾Ò *Ò v¦&Ïb &Ë ô ÒÏ> ®º non-GM & vf GM &v¢ ·bþË(>ö)öB VÃAj *Ò ÖÁ FÛ>º v¦f ?f B;j Û~ v¦ &~ ÿ ÖÎ ê, ïË &~B ò ÒÏ~& . &Nê f &* öö ~º 'Ëj &V~V*~ &N êº 70-100oC, &*f 10-40ªb ~& . ÚB¢ * þÿbº Æ¢(New Zealand White)¢ Essence Medical Inc.(Seoul, Korea)¦V «~ ÒÏ~& . WîªC '' ò~ Wî~ ïf Micro-kjeldahl»(9)b 3² > G;~ ï8j ÒÏ~& . ''~ òöB Wî ~ ª¶ïj Ò~ ß Wî Z¢ dV * Weber (10)~ O»ö V¢ *V'ÿ~& . r gelf 12% polyacrylamide¢ ÒÏ~& .. 829. ÚNJ wellö If ê, «V R«®~ PBS jÏj. B~ ^¿~& . ^¿j Þ plateö PBS jÏö 1% îæÖF¢ 90 µLO I 37oCöB 1* ÿn V·~ blocking ê, ^¿";j 4² >~& . Primary antibody solution(1 : 10,000)j 85 µLO ' wellö R«~, 37oC V·V öB 1* ÿn V·~, ¢ 4² ^¿~& . HRPgoat anti-rabbit lgG ¯, secondary antibody solution(1 : 10,000) j 80 µLO I 37oC~ V·Vö 1* v 4² ^¿~& . Primary antibodyf secondary antibodyræ If plateö TMB(3,3'-5,5'-tetramethyl benzidine)Ïj 100 µLO I 37oC öB 20ª* Bï>wÎ r, ELISA reader(Sunrise, Tecan, Austria) 405 nmöB 7ê¢ G;~& .. Ö 5 V WîªC þ~ non-GM v¦f GM v¦öB Wî ïf '' 10.0% 5 8.8% ©b ªC>Ú GM v¦ non-GM v¦öB Wî ï ² ¸f ©b ¾æÒ . öò &vf &B v¦~ *V'ÿ NZ æzº Fig. 1" ? . *>'b &vöB ¾æ¾º Wî Z& v¦. F«~ z ô~b, ö j v¦~ Z >& 'f ãË j & . º vF& ¾Òö ~ WB ®ÏW Wî jæf þ B>, *>¢ Î&~ v¦¢ w Ò r >ÏW Wî ¢¦ B>îV r^ ©b 'B . v¦f &v, ÎvöB non-GM ò~ Z& GMò. F«~ ôf ©j " > ® . non-GM &v~ ãÖö GM &vöBº æ pº 6 kDa~ Wî Z& ¾æÒb, non-GM v¦~ ãÖöº GM v¦öB ¾æ¾æ pº 113 kDa ~ Z& ¾æÒ 24 kDa~ Zê F«ö ®ÚB N¢ & . *F :f ? Kwak (7)f GM &v~ CP4 EPSPSö & ßÚ¢ Ï~ F*¶ Ò ¦¢ 6 ~&b¾ CP4 EPSPS& jò Ò F*¶& Ú® j röº ¢ 6 > ìº ^B6 ® . V¢B öBº GM v¦öBº ¾æ¾æ pº non-GM v¦~. ÚB *V'ÿB gelj 0.1 M KCl Ïö 5-15ª* æ~ Z ö ï * V öZ(specific band)òj .~ PBS(phosphate-buffered saline) jÏj IÚ 1 mL~ ¦b ; ï~ îz~& . îz öj Æ¢~ 3-5 ¦*ö 2" *Ïb 3² b~"Ò~& . îæï "Ò 1"¢ ê, Æ ¢~ æ ; b¦V 200-300 µL ;ê~ .j j~, ¢ 37oC V·VöB 1* ÿn O~~ .w¢ B Î ê, öªÒ(5,000 rpm, 10 min)~ ç[j ~ sodium azide¢ 0.02% Î&~ −70oCöB ïÿ &~& . ELISA Microplate(96-well plastic plate)~ ' wellö « ³ê& 1 mg/mL >ê PBS jÏb C öj 90 µLO I Ú 37oCöB 2* ÿn V· ê, antigen Ïj B~& . Tween-20j 0.05% ~º PBSjÏ 200 µL¢ OÞO Î. Fig. 1. Gel electrophoretic patterns of soybeans and soybean curds. M: protein marker, 1: non-GM soybean, 2: GM soybean, 3: soybean curd produced from non-GM soybean, 4: soybean curd produced from GM soybean..

(3) 830. ®"²æ B 36 ² B 5 ^ (2004). Fig. 2. Effect of heating time on the gel electrophoretic pattern of non-GM soybean curds. M: protein marker, 1: soybean curd produced from non-GM soybean heated at 80oC for 10 min, 2: soybean curd produced from non-GM soybean heated at 80oC for 20 min, 3: soybean curd produced from non-GM soybean heated at 80oC for 30 min, 4: soybean curd produced from non-GM soybean heated at 80oC for 40 min.. 113 kDa Wî" F«~ Ôf ª¶ïj æò 24 kDa W îj Ò*W ®º ß Wî öb F~ CP4 EPSPS & jò Ò F*¶& b«B ãÖöê v¦ Ú~ nonGM öò~ ÒÏïj 6 > ®ê ~& . &* 5 Nêö 8 *8'ÿNZ æz WîöB æW Nêº &B 70-80oC Nê& ¸jî > æW³ê& ¢ê . &¦ª~ Wî ª¶º · ª ¶ Ú FÖ" jF Ö~ ç^·Ïö ~ ;b Ò~ Wî ª¶ f ª¶ ç~ ßö Ö; ¦* epitope j æò . epitopef ³'b ÖB continuous amino acid Bj æò Wî _f non sequential amino acid B j æò Wî" Îv Ö . ¾ æWB Wîf amino acid~ B ³'b ÖB crytotopes¢ W . ¯, Wî æW> native epitopes~ ¦ª' Ò¢ö" ³' cryptotopes~ Wj rÚ ¸f NêöB Wî æW> native epitopesº Ò¢æ, ³' cryptotopes& ¾Ú¾² B. (11). WËr^ö 80oC ç~ &f ö" ÖF native epitopes& Ò¢öb Úf~ >w &ËWê *Ú. > ®j ©b 'B . ¾ Â Wî æWB Wî Úf z ;~² &w~, J®J *çj Ï~ Wî~ Î&ï" ¾Ò Nê¢ 6; (12,13)ê Ú^ ® . f ? ¾Ò ®ö & öÁÚ> w «{® «>æ p~, 6v¦º ' B²Òö V¢ vF~ &* 5 Nêö ²*~ N& B > ®b æ, ¾Ò Nê 5 * *V'ÿNZ ¾j&Bº ELISA»ö ÚÆ 'Ëj ©æ¢ V~V *~, & * 5 Nêö V *V'ÿNZ æz¢ Fig. 2f 3ö ¾ æÚî . ¢>'b v¦& ö vFº 80oCöB 20ª Ú ¾Ò>æ vF¢ 80oCöB 10-40ª* ¾Ò~ &B v ¦~ *V'ÿNZ(Fig. 2)" 70-100oC~ NêöB 20ª* ¾ ÒB v¦~ *V'ÿNZ(Fig. 3)j V~& .. Fig. 3. Effect of heating temperature on the gel electrophoretic pattern of soybean curds produced from non-GM soybean heated for 20 min. M: protein marker, 1: soybean curd produced from non-GM soybean heated at 70oC for 20 min, 2: soybean curd produced from non-GM soybean heated at 80oC for 20 min, 3: soybean curd produced from non-GM soybean heated at 90oC for 20 min, 4: soybean curd produced from non-GM soybean heated at 100oC for 20 min.. Wî æWö V Z æz& ©b .ç>îb ¾, & Nêö V Z~ æzº Â]~² ¾æ¾æ p~. . ¾ 30ª ç~ & öB 113 kDa Wî Z& ;æ *Ú' Z »Z>º *çj " > ®î . ¾ ¢>' v¦B ~ ¾Ò *f 20ª Úæ non-GMv¦ 6êj * ELISAöº 'Ëj ~æ p j ©b 6>Ú 80oCöB 20ª* ¾ÒB non-GM v¦ ~ 113 kDa" 24 kDa Wî Ú¢ B~& . Non-GM 113 kDa 24 kDa Wî~ Ú ;W F«~ >W { ~ Ôf ª¶ïj ¾æÚº nonGM v¦~ 24 kDa Zf GM v¦öBº *& ¾æ¾æ p j non-GM ß Wî Îæº 113 kDa Z¢ öb ~ Æ¢ö '' 2" *Ïb 3² ?f ·~ öj b~" Ò~ Ê, îæï 1"¢ ê 113 kDaj Î Æ¢f 24 kDaj Î Æ¢~ æ ; öB '' j. . b .Ój ªÒ~& . .Ój Ï~ non-GM v¦f GM v¦¢ 405 nmöB ELISAö ~ G;B 7ê ¢ Fig. 4ö ¾æÚî . b& solid phaseö ÖÊº ö ¯, non-GM v¦ ºÂbj 10−1-10−8V C~ Úf Ö V . Ö" 113 kDa" 24 kDa Wî ÎvöB Úf >w~º ©b ¾æÒ . ¾ non-GM 113 kDa Wî non-GM 24 kDa Wî ¸f 7ê¢ Ú W ~ N¢ ¾æÚî . GM v¦f ª«~² N¢ º non-GM v¦~ 113 kDa Wî Ôf ª¶ïj æò non-GM v¦~ 24 kDa Wî ¸f 7ê¢ &, non-GM v ¦f GM v¦ *~ N 6 113 kDa Wî 24 kDa W î ôf N¢ Ú ~ Wö N¢ ¾æ Úî . Nº Úf~ ßW" zK ¯, non-GM v¦ ~ 113 kDa Wî non-GM v¦~ 24 kDa Wî Ú.

(4) Î²G;»j Ï v¦ 7~ F*¶ Ò &vWî ªC. Fig. 4. ELISA of various dilutes of soybean curds produced from GM and non-GM soybeans against 24 kDa and 113 kDa proteins of non-GM soybean curd.. f~ ÖK Ö>~V r^ ©b 'B . V¢B Ú W ö F öbB GM v¦f ª«~² N¢ º non-GM v¦~ 113 kDa Z¢ F~º © Ú~ ßW" zKj ¸, ÚW~ Wj ¸º Ï ~ 'B . 6 24 kDa Wîê 113 kDa Wî. Ôf 7ê¢ Vº ~¾ GM v¦~ 24 kDa Wî. º ¸f ÖKj "Ú non-GM v¦~ 24 kDa Wîö &Bê Ú& ;W>îrj r > ®² ~& . f ?, GM WîöBê ¾æÒbæ ß Wî jò ©b 6 >î~ non-GM v¦~ 24 kDa Wîê B ©f F« non-GM v¦~ 24 kDa Z& large peptide ö >Ú Úf~ zKj ¸ ©b 'B . Large peptide f ß öÖ;¦*¢ W~º Wî j æò BÚ~ Ú W·Ïj W'b Fê~ peptide f ®j r ±f ö >º © . 6 ö > º peptide~ ;º Úf~ Öj Ö;~º 7º º²& >º, sequence 56-107j æîº lysozyme peptides~ ãÖö º S-S Öö ~ peptidef Ö~ Úf~ zK ¸jê (6). æ non-GM v¦~ 24 kDa Wî f protein marker lysozyme *~öB ¾æÂ Z Úf ~ zK ¸f lysozyme~ Wîj æò Wî ºGB . 24 kDa Wî~ Wîr^ö GM v¦f~ ßW ìº Wîªöê &Ëê ©b 'B . Non-GM v¦~ 113 kDa Wî Úº CV>& 10−1-10−6 ¢ röº ·f öï~ Nö V 7ê~ N& ~ ¾æ¾æ p~ 10−7 ~~ CV>öBº 6ê& Ôj Ú W ¦¢ G;~V ÚJ . æ 10−1-10−6~ C V>öB 6v¦~ öò& non-GMO ©j 6ê~º © Ï ©b ¾, 10−7 ~~ CV>öBº 6ê Ú JÞ ©b 'B .. 831. Fig. 5. Standard curve of ELISA against 113 kDa protein of nonGM soybean curds at the dilution range from 10−7 to 10−6.. Non-GM &Lf GM &L~ b«jN /; 10−6-10−7~ CV>öBº £*~ non-GM v¦~ 113 kDa Wî ï Nöê Â] 7ê~ Ã6j æ CV>öB non-GMOf GMO~ b«jNj 6;~V *~ *~ Cj ^ªz~ Fig. 5ö ELISAö ~ 7êf non-GM Wî ï~ &ê¢ ¾æÚº &Fb ·W~& . Non-GM &v:GM &v~ jNj 100:0-0:100 b . ò non-GMOf GMO& b«B v¦¢ & ~ 10−6-10−7~ CV>öB ELISAö ~ 7ê¢ G;~ 8j &F(Fig. 5)ö 'Ï~ êÖB non-GMO~ ·" B v¦ Úö Ò~º non-GMO~ ·j Table 1ö ¾æÚî . 100% non-GM &v B v¦~ ãÖö Bº 8.40 mg/ mL~ non-GM &vWî Ò~º 7ê¢ ¾æÚÚ¢ ~ ¾ 8.50 mg/mL Ò~º 7ê¢ ¾æÚî, 60% non-GM &vf 40% GM &v& b«B öò B v¦~ ãÖö º Bº 5.0 mg/mL~ non-GM &vWî Ò~º Table 1. Comparisons of actual and estimated amounts of nonGM proteins in soybean curds Ratio of non-GM soybean:GM soybean in soybean curd 100:0 60:40 40:60 20:80 10:90 0:100. Amount of non-GM Actual amount of protein in soybean curd non-GM protein estimated with ELISA in soybean curd and standard curve (mg/mL) (mg/mL) 8.40 5.00 3.20 1.66 0.80 0.00. 8.50 5.10 3.40 1.70 0.85 0.00.

(5) ®"²æ B 36 ² B 5 ^ (2004). 832. 7ê¢ ¾æÚÚ¢ ~¾ 5.1 mg/mL Ò~º 7ê¢ ¾æ Úî (Table 1). 7% Ú~ JN¢ ¾æêb, non-GM & vf GM &v& b«>Ú ÒÏB ãÖöê ELISA¢ Ï~ &Û' b«jNj G; > ®rj Ò~ ® . æ ELISA»" &Fj Ï~ B FÛ>º v¦ ~ GMO 5 non-GMO b«jNj G;~º ©ê &Ë~Ò¢ ' . Table 1öBf ? B 7êf ÖÂB 7êö £*~ N¢ º ©f öÖ;V¢ F~º Wî ~ ·Ï r^ ©b 'B . ÖÒ& ö~º non-GM v¦~ 113 kDa Wîöò jî¢ non-GM Wî 7~ ¢¦ ª non-GM 113 kDa Wî" öÖ;V¢ F~² >Ú .ç~. ² ¸f 7ê& ¦ÂB ©b 'B. (6). ¾ ^Bº ¦ z ;& ö~ ;Bòbê Ïª® BFF > ®j ©b 'B . öB GM &®" non-GM &®~ Wî ïf N¢ æº p~b¾, SDS-PAGE¢ Ï ªC öBº ª« ß Z¢ &bæ SDS-PAGE~ ¯òb GMOf non-GMO¢ 6ê >ê ®j © . ¾ SDS-PAGE~ ¯òbº &®ö ®ÚB GMOf non-GMO b«jN~ 6; ®&Ë~ GMOªCj «{® ~Vú B ® Îê . *~ Ö"¢ Æ& ~ *Ò 6 > ®º 6 v¦~ F*¶ Ò ¦¢ G; Ö", ¢> Ò¾ËöB 6. >º 6v¦~ ãÖ non-GM &vWîj 28% ;ê F ©b G;>î, 100% Ö&vòj ÒÏ~ ® F* ~ ®º VëÚ v¦~ ãÖöº 80% ;ê non-GM &v Wîj F~ ®º ©b ¾æ¾ non-GM &vWî~ ï jv' ¸~ . ¾ ?f VëÚ~ v¦¢ æ¢ê non-GM &v~ ïö N¢ ³ËöB öò&v¢ > ÷~º êöB¦V Æ& &Ò& jº~ v¦6 ö º F*¶Ò ¦¢ ª«~² ¢ jº& ®º © b 'B .. º. £. öBº F*¶ Ò >æ pf(non-GM) &vf F *¶ ÒB(GM) &v& b«>Ú BB v¦öB Î² G;»j Ï~ non-GM &v~ b«ïj º;~¶ ~ & . v¦~ SDS-PAGE ¯ Ö" non-GM v¦öBò ¾æ ¾º ß Wî non-GM 113 kDa Zf non-GM" GM v ¦öB Îv ¾æ¾º non-GM 24 kDa Z¢ Fê~ . j Æ¢ö ~ ÚW ¦¢ ELISA Ö" non-GM 113 kDa" non-GM 24 kDa Wî Îv Ú& ;WNj {. ~& 10−1-10−6~ Wî CV>öB v¦¢ Úö &~ ELISAb öò&v~ GM¦¢ { > ®î. . 7, 6ê& ¸~~ non-GM 113 kDa Wîj 10−7-10−6~ V> C~ ELISA 7êf non-GM Wî ~ &ê¢ ¾æÚº &Fj ·W~&, ª~ non-GM &vf GM &v¢ b~ B v¦~ ELISA 7ê¢ &F" jv~ non-GM öòf GM öò ·b~ b «Nj G; Ö", ¸f ;{ê¢ & .. ^. ò. 1. Park SH. Genetically modified food and its safety assessment. Korea Soybean Digest. 16: 21-30 (1999) 2. Park SH. Food derived from new plants varieties derived through recombinant DNA technology. Korea Soybean Digest. 16: 20-30 (1999) 3. Nordic Council of Ministry. Health Effects of Marker Genes in Genetically Engineered Food Plants. Tema Nord Copenhagen Press, Copenhagen, Denmark (1996) 4. Wurz A, Bluth A, Zeltz P, Pfeifer C, Willmund R. Quantitative analysis of genetically modified organisms in processed food by PCR-based methods. Food Cont. 10: 385-389 (1999) 5. Meyer R. Development and application of DNA analytical methods for the detection of GMOs in foods. Food Cont. 10: 391-399 (1999) 6. Kwak BY, Ko SH, Park CW, Son DY, Shon DH. Development of enzyme-linked immunosorbent assay for glyphosate-tolerant soybeans. Korean J. Food Sci. Technol. 35: 366-372 (2003) 7. Kwak BY, Ko SH, Shin WS, Shon DH. Analysis of genetically modified soybean and soybean sprout by enzyme-linked immunosorbent assay. Korean J. Food Sci. Technol. 35: 556-560 (2003) 8. Seo HS. The qualitative and quantitative detection of RoundupReady soybeans using immnological methods. PhD thesis, Seoul National University, Seoul, Korea (2002) 9. AACC. Approved Method of the AACC. 10th ed. American Association of Cereal Chemists, St. Paul, MN, USA (2000) 10. Weber K, Pringle JR, Osborn M. Measurement of molecular weight by electrophoresis on SDS-acrylamide gel. Meth. Enzymol. 26: 3-27 (1971) 11. Paraf A. Application of immunochemistry for protein structure control. pp. 613-620. In: Food Proteins and Their Applications. Damodaran S, Paraf A (eds). Marcel Dekker, Inc., New York, NY, USA (1997) 12. Varshney G, Paraf A. Use of specific polyclonal antibodies to detect heat treatment of ovalbumin in mushroom. J. Sci. Food Agric. 52: 261-267 (1990) 13. Wang CH, Booren AM, Abouzied MM, Pestka JJ, Smith DM. ELISA determination of turkey roll end-point temperature: effects of formulation, storage and processing. J. Food Sci. 58: 12581263 (1993) (2004j 7ú 5¢ %>; 2004j 10ú 13¢ j).

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