Screening and Subcloning of High Producer Transfectomas Using Semisolid Media and Automated Colony Picker

Abstract

Generation of high-producing clones is a perquisite for achieving recombinant protein yields suitable for biopharmaceutical production. However, in many industrially important cell lines used to produce recombinant proteins such as Chinese hamster ovary, mouse myeloma line (NS0), and hybridomas, only a minority of clones show significantly above-average productivity. Thus, in order to have a reasonable probability of finding rare high-producing clones, a large number of clones need to be screened. Limiting dilution cloning is the most commonly used method, owing to its relative simplicity and low cost. However the use of liquid media in this method makes the selection of monoclonal hybridoma and transfectoma colonies to be labor intensive and time consuming, thus significantly limiting the number of clones that can be feasibly screened. Hence, we describe the use of semisolid media to immobilize clones and a high-throughput, automated colony picker (ClonePix FL) to efficiently isolate monoclonal high-producing clones secreting monoclonal antibodies.

Optimization of cell line development in the GS-CHO expression system using a high-throughput, single cell-based clone selection system

“…Optimization of the semi-solid medium formulation for single cell-based clone selection There are several reports of single cell-based clone selection using semi-solid media (27,31,32). However, they do not describe the medium formulation for single cell-based selection.…”

Deletion of a telomeric region on chromosome 8 correlates with higher productivity and stability of CHO cell lines

“…Screening of hundreds of clones is usually required to identify and select the ones with favorable characteristics, as the productivity of many clones is rather low. To reduce the manual workload, various automated systems have been established for cell cultivation and screening (Browne and Al-Rubeai, 2007;Dharshanan and Hung, 2014;Yoshimoto and Kuroda, 2014). The proportion of high producing cells can be enlarged via the stringency of selection and several fluorescence-activated cell sorting (FACS)-based methods (Browne and Al-Rubeai, 2007;Carroll and Al-Rubeai, 2004;Lai et al, 2013).…”

Integration of high‐throughput analytics and cell imaging enables direct early productivity and product quality assessment during Chinese Hamster ovary cell line development for a complex multi‐subunit vaccine antigen

“…In addition to the approach described here, other high‐throughput early clone characterization strategies have also been developed. ClonePix (Molecular Devices, Sunnyvale, CA) was able to automatically image, select, and pick mammalian cell colonies expanded in semi‐solid media, based on a number of parameters such as colony size and yield of a recombinant secreted product . However, the ClonePix readouts are not quantitative.…”

“…ClonePix (Molecular Devices, Sunnyvale, CA) was able to automatically image, select, and pick mammalian cell colonies expanded in semi-solid media, based on a number of parameters such as colony size and yield of a recombinant secreted product [22][23][24]. However, the ClonePix readouts are not quantitative.…”