Figure 3
Characterization of mAb-producing stable CHO pools expressing different levels of MANII and cGNTIII genes. The expression of each gene was varied in three levels: zero (no expression, M0 and G0), medium (Mm and Gm) and high (Mh and Gh), resulting in nine combinations of different expression levels of MANII and cGNTIII genes. Two stable pools were generated independently through RMCE for each combination of MANII and cGNTIII expression levels. The NEG pools were dMCL without RMCE. Control pools expressed Rituximab LC and HC only. (A) Flow cytometry analysis of the nine stable mAb-producing cell pools after they were fully recovered from antibiotic selection. Representative data from one replicate pool of each condition was showed. (B) Relative MANII and cGNTIII transcript levels in the stable pools was analysed by quantitative real-time PCR (qRT-PCR). β-actin gene was used as internal control. Each point represents the average and standard deviation of relative transcripts from two independent stably transfected pools. (C) Western blot analysis of MANII and cGNTIII protein levels in the stable pools. GAPDH was used as loading control. Uncropped blot images are available in Supplementary Fig. S1. The relative expression of MANII and cGNTIII genes was normalized to the corresponding loading control in each sample.
