Development of a mouse-feline chimeric antibody against feline tumor necrosis factor-alpha

DOKI, Tomoyoshi; TAKANO, Tomomi; HOHDATSU, Tsutomu

Western immunoblotting assay of mouse mAb 2-4 and chimeric mAb 2-4: FO cells harboring the expression vector were cultured, and chimeric mAb 2-4 in the culture supernatant was collected. Chimeric mAb 2-4 and mouse mAb 2-4 were individually purified using a protein A or protein G column. The purified mAbs were subjected to SDS-PAGE, and their purities were confirmed by CBB staining. No protein band other than those of mAbs was detected by SDS-PAGE. The purified chimeric mAb 2-4 was western blotted with mouse mAb 2-4. When non-reduced mouse mAb 2-4 and chimeric mAb 2-4 were electrophoresed, smear-like bands were detected at 135-kDa and higher (Fig. 1A) . When the blot was reacted with anti-mouse IgG antibody, bands were detected in the lane applied with mouse mAb 2-4, but no band was detected in the lane applied with chimeric mAb 2-4. When the blot was reacted with anti-feline IgG antibody, bands were detected in the lane applied with chimeric mAb 2-4, but not in the lane with mouse mAb 2-4.