Magnetic nanoparticle based purification and enzyme-linked immunosorbent assay using monoclonal antibody against enrofloxacin

Kim, Nam-Gun; Kim, Myeong-Ae; Park, Young-Il; Jung, Tae-Sung; Son, Seong-Wan; So, ByungJae; Kang, Hwan-Goo

A total of 3 mg of an amine-functionalized MNP suspension (100 L) was washed three times using a magnet in coupling buffer (0.01 M pyridine, pH 6.0), after which a 1 mL solution of 5% glutaraldehyde was added. Following incubation for 30 min at room temperature while shaking, the particles were washed with coupling buffer by magnetic separation. Next, mAb coupling with the MNPs was conducted by dissolving 130 g of ENR mAb in 500 L of coupling buffer. The mixture was subsequently rotated at 94 × g overnight at room temperature, after which the MNP-mAb conjugates were washed with coupling buffer (500 L). The reaction was quenched by the addition of a solution of glycine and then washed three times with PBS buffer and stored in PBS buffer at 4 o C before use.