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Abstrak :
[ABSTRAK
Antibodi poliklonal anti aflatoksin B1 telah berhasil diproduksi pada hewan uji kelinci betina New Zealand White setelah diimunisasikan hapten aflatoksin B1-CMO yang dikonjugasikan dengan Bovine Serum Albumin (BSA) sebagai antigen. Hapten aflatoksin B1-CMO disintesis menggunakan metode karbodiimida dengan substrat aflatoksin B1 dan carboxymethyl hydroxylamine hemihydrochloride (CMO) sebagai linkernya. Hasil karakterisasi kromatografi lapis tipis dengan nilai Rf rata-rata sebesar 0.395, spektrum UV-Visibel dengan puncak λ maks pada 362, 264, 218 nm, spektrum IR dengan puncak 3448.126 cm-1 (3000-3600 cm-1) : OH, pada 1632.249 cm-1(1540-1725 cm-1) : C=O, dan 1642.451 cm-1 (1640-1690 cm-1) :C=N (Oksim), dan hasil fragmentasi spektrometri massa (MS/MS) pada m/z 386, 368.2, 310 membuktikan hapten aflatoksin B1-CMO berhasil disintesis. Hapten ini kemudian dikonjugasikan dengan BSA membentuk antigen aflatoksin B1-BSA (AFB1-BSA) sebelum diimunisasikan ke kelinci. Spesifitas antigen AFB1-BSA terhadap antibodinya dan uji konjugasi hapten ke BSA menunjukkan hasil positif menggunakan uji Dot Blot Immunoassay dengan konsentrasi BSA di dalam antigennya sebesar 1.74 mg/mL. Serum darah kelinci berdasarkan uji Agar Gel Precipitation Test (AGPT) positif mengandung antibodi poliklonal anti aflatoksin B1 setelah dua pekan (hari ke-11) sejak imunisasi primer antigen AFB1-BSA dilakukan. Dari serum darah bleeding panen, diperoleh konsentrasi antibodinya sebesar sebesar 2.19 mg/mL. Immunokromatogafi strip tes berhasil dibuat dengan nanopartikel iridium oksida (IrO2 NPs) sebagai kandidat label antibodinya dan dapat digunakan untuk mendeteksi sampel H IgG pada rentang 0.1 μg/mL sampai 10 μg/mL. Studi pendahuluan ini menunjukkan bahwa perangkat strip tes ini dapat digunakan untuk aplikasi konjugat sensor antibodi anti aflatoksin B1-nanopartikel iridium oksida untuk deteksi aflatoksin B1.

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ABSTRACT
Polyclonal antibody against aflatoxin B1 have been successfully produced in New Zealand White Rabbit after immunized by hapten of aflatoxin B1-CMO conjugated with Bovine Serum Albumin (BSA) as antigen. Hapten of aflatoxin B1-CMO was synthesized using carbodiimide method with afltoksin B1 as substrate and carboxymethyl hydroxylamine hemihydrochloride (CMO) as its linker. The characterization results of thin layer chromatography with Rf value of 0.395, the spectrum of UV-Visible with λ max peaks at 362, 264, 218 nm, the IR spectrum with peak at 3448.126 cm-1 (3000-3600 cm-1): OH , 1632.249 cm-1(1540-1725 cm-1): C = O, 1642.451 cm-1 (1640-1690 cm-1): C = N (oxime), and the results of mass spectrometry fragmentation (MS / MS) at m/ z of 386, 368.2, 310 proved that hapten of aflatoxin B1 -CMO successfully synthesized. Then, the hapten was conjugated to BSA to form antigen of aflatoxin B1-BSA (AFB1-BSA) before immunized to rabbits. The specificity of antigen of AFB1-BSA to its antibody and the confirmation of hapten-BSA conjugated showed positive results using dot blot immunoassay with BSA concentration in the antigen of 1.74 mg/mL. Based on Agar Gel Precipitation Test (AGPT) shown the rabbit blood serum resulted positive for polyclonal antibody against aflatoxin B1 after two weeks (day 11st) since the primary immunization of its antigen. From blood serum bleeding at harvest obtained the concentration of antibodies was 2.19 mg / mL. An Immunochromatogaphic test strip was successfully fabricated using iridium oxide nanoparticles (IrO2 NPs) as a labeled antibody candidate and can be used to detect the IgG H sample between of 0.1 μg/mL to 10 μg/mL. This preliminary study shown that the device can be used for applications of antibody against aflatoxin B1-nanoparticle iridium oxide conjugate for detection of aflatoxin B1 , Polyclonal antibody against aflatoxin B1 have been successfully produced in New Zealand White Rabbit after immunized by hapten of aflatoxin B1-CMO conjugated with Bovine Serum Albumin (BSA) as antigen. Hapten of aflatoxin B1-CMO was synthesized using carbodiimide method with afltoksin B1 as substrate and carboxymethyl hydroxylamine hemihydrochloride (CMO) as its linker. The characterization results of thin layer chromatography with Rf value of 0.395, the spectrum of UV-Visible with λ max peaks at 362, 264, 218 nm, the IR spectrum with peak at 3448.126 cm-1 (3000-3600 cm-1): OH , 1632.249 cm-1(1540-1725 cm-1): C = O, 1642.451 cm-1 (1640-1690 cm-1): C = N (oxime), and the results of mass spectrometry fragmentation (MS / MS) at m/ z of 386, 368.2, 310 proved that hapten of aflatoxin B1 -CMO successfully synthesized. Then, the hapten was conjugated to BSA to form antigen of aflatoxin B1-BSA (AFB1-BSA) before immunized to rabbits. The specificity of antigen of AFB1-BSA to its antibody and the confirmation of hapten-BSA conjugated showed positive results using dot blot immunoassay with BSA concentration in the antigen of 1.74 mg/mL. Based on Agar Gel Precipitation Test (AGPT) shown the rabbit blood serum resulted positive for polyclonal antibody against aflatoxin B1 after two weeks (day 11st) since the primary immunization of its antigen. From blood serum bleeding at harvest obtained the concentration of antibodies was 2.19 mg / mL. An Immunochromatogaphic test strip was successfully fabricated using iridium oxide nanoparticles (IrO2 NPs) as a labeled antibody candidate and can be used to detect the IgG H sample between of 0.1 μg/mL to 10 μg/mL. This preliminary study shown that the device can be used for applications of antibody against aflatoxin B1-nanoparticle iridium oxide conjugate for detection of aflatoxin B1 ]

Abstrak :
Strip test immunokromatografi telah dikembangkan untuk deteksi kuantitatif kandungan melamin dalam sampel susu dan produk susu. Strip test ini dibuat berdasarkan reaksi kompleks antigen-antibodi (melamin-anti melamin). Nanopartikel emas (AuNP) digunakan sebagai label terhadap antibodi membentuk kompleks nanopartikel emas-antibodi (AuNP-Ab). Studi kuantifikasinya dilakukan dengan mengukur nanopartikel emas secara elektrokimia menggunakan metode anodic stripping voltammetry (ASV). Perangkat elektroda pada strip test difabrikasi dengan menggunakan boron-doped diamond (BDD) sebagai elektroda kerja, Ag/AgCl sebagai elektroda standar, dan platina (Pt) sebagai elektroda penunjang. Limit deteksi (LOD) untuk AuNP sebesar 24.3 μM dapat dicapai menggunakan perangkat ini. Komponen strip test immunokromatografi dilakukan dengan kondisi volume sampel melamin 100μL, nanopartikel emas-antibodi (AuNP-Ab) 60 μL, antibodi penangkap 10 μL, waktu immunoreaksi 7 menit, volume pelarut HClO4 0.02 M sebesar 200 μL, dan dengan kondisi elektrokimia pada rentang potensial dari -0.5 V sampai 1.5 V, scan rate 100 mV/s, serta waktu deposisi 240 sekon. Limit deteksi (LOD) strip test immunokromatografi yang dapat dicapai adalah 0.15 mg melamin/mL PBS (phosphate buffer solution) (150 ppm). Hasil ini menunjukkan bahwa strip test immunokromatografi tersebut dapat digunakan untuk mendeteksi keberadaan melamin secara kuantitatif.

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Immunochromatographic strip test was developed for quantitative detection of melamine in sample milk and milk products. The strip test was developed based on antigen-antibody (melamine-anti melamine) complex reaction. Gold nanoparticle (AuNP) was used as a label to antibody to form antibody-labeled gold nanoparticle (AuNP-Ab). The quantification study was studied by electrochemical measurement of gold nanoparticle using anodic stripping votlammetry (ASV) method. Electrode device for the strip test was fabricated using boron-doped diamond (BDD) as the working electrode, Ag/AgCl as the reference electrode, and platinum (Pt) as the counter electrode. Limit of detection(LOD) of AuNP detectable in the strip test electrode device was 24.3 μM. Immunochromatographic strip test using sample volume of 100μL, AuNP- Ab of 60 μL, capturing antibody of 10 μL, immunoreactions time of 7 min, volume HClO4 0.02M of 200 μL performed by electrochemical condition of scan rate of 100 mV/s and deposition time of 240 s, showing an LOD of 0.15 mg melamine/mL PBS (phosphate buffer solution) (150 ppm). The results indicated that the strip test immunochromatographic could be used to quantitative detection of melamine.

Abstrak :
Polyclonal antibodies of aflatoxin B1 were successfully produced from New Zealand White female rabbits after immunization by the hapten of aflatoxin B1-carboxymethyl hydroxylamine hemihydrochloride (AFB1-CMO) conjugated with bovine serum albumin (BSA) as the antigen. The hapten was synthesized using the carbodiimide method with CMO as a linker. Absorption peaks at 362, 264, and 218 nm were observed as a result of characterization with UV-Vis spectroscopy, while IR spectroscopy showed peaks at 3448 cm-1 and 1642 cm-1 attributable to the hydroxyl and nitrile groups, respectively. Furthermore, mass spectrometry showed fragmentation at the m/z of 386, 368.2, and 310, which confirms that the hapten of AFB1-CMO was successfully synthesized. The hapten was then conjugated with BSA to serve as an antigen of AFB1 when it was injected into the rabbits. The specificity of the antigen towards its antibody and the confirmation of hapten-BSA conjugation were characterized using the dot blot immunoassay, which showed a BSA concentration of 1.74 mg/mL. Two weeks after the primary immunization by its antigen, agar gel precipitation testing showed that the rabbit blood serum had positive results for polyclonal antibodiest against AFB1 with the highest concentration of antibodiest of 2.19 mg/mL.

Sintesa of Poliklonal Antibodi Aflatoxin B1. Antibodi poliklonal aflatoksin B1 telah berhasil diproduksi pada hewan uji kelinci betina New Zealand White setelah diimunisasi dengan hapten aflatoksin B1-carboxymethyl hydroxylamine hemihydrochloride (AFB1-CMO) yang dikonjugasikan dengan bovin serum albumin (BSA) sebagai antigen. Hapten AFB1 disintesis menggunakan metode karbodiimida dengan CMO sebagai linker. Puncak absorbansi pada 362, 264, 218 nm teramati sebagai hasil karakterisasi menggunakan spektrofotometer UV-Visibel, sementara dengan spektrum IR diperoleh puncak pada 3448.126 cm-1 dan 1642.451 cm-1 yang masing-masing mengindikasikan adanya gugus hidroksil dan nitril. Hasil spektrometri massa menunjukkan fragmentasi pada m/z 386, 368.2, dan 310 yang membuktikan hapten AFB1-CMO telah berhasil disintesis. Hapten ini kemudian dikonjugasikan dengan BSA agar dapat berperan sebagai antigen AFB1 ketika diinjeksikan pada kelinci. Kekhususan antigen aflatoksin B1 terhadap antibodinya dan konfirmasi konjugat hapten-BSA menunjukkan hasil positif pada uji dot blot immunoassay dengan konsentrasi BSA sebesar 1.74 mg/mL. Dua pekan setelah imunisasi primer, agar gel precipitation test menunjukkan hasil positif terhadap antibodi poliklonal AFB1 dengan konsentrasi tertinggi sebesar 2.19 mg/mL.