Synthesis of Polyclonal Antibodies against Aflatoxin B1

Wiyogo Prio Wicaksono, Deni Samsudin Permana, Romsyah Maryam, Yasuaki Einaga
2015 Makara Journal of Science  
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
more » ... y 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. Abstrak 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.
doi:10.7454/mss.v19i3.4847 fatcat:gmp4d6ji3zfvlpkms25xizznfy