Improved CHO Cell Line Stability and Recombinant Protein Expression During Long-Term Culture

Therapeutic proteins require proper folding and posttranslational modifications to be effective and biologically active. Chinese hamster ovary (CHO) cells are by far the most frequently used host for commercial production of therapeutic proteins. However, an unpredictable decrease in protein productivity during the time required for scale up impairs process yields, time, finance, and regulatory approval for the desired product. Therefore, it is important to assess cell lines at stages throughout the period of long-term culture in terms of productivity and various molecular parameters including plasmid and mRNA copy numbers and location of the plasmid on the host cell chromosome. Here, we describe methods, which are frequently used to analyze stability of the recombinant CHO cells over long-term culture. These procedures include the following; western blotting, ELISA to evaluate protein production, real-time PCR to analyze plasmid and mRNA copy numbers, and fluorescent in situ hybridization (FISH) to assess the location of the inserted plasmid on host cell chromosomes.