Surface display vectors for selective detection and isolation of high level antibody producing cells.

Lang S1, Drewello D1, Wichter J2, Nommay A1, Wilms B1, Knopf HP1, Jostock T3.
Source: Biotechnol Bioeng
Publication Date: (2016)
Issue: 113: 11
Research Area:
Gene Expression
Cells used in publication:
Species: hamster
Tissue Origin: ovarian
CHO [suspension]
Species: hamster
Tissue Origin: ovarian
4D-Nucleofector® 96-well Systems
4D-Nucleofector® X-Unit

Nucleofection of CHO suspension cells in serum free, chemically defined, animal component free mdeium during Nucleofection, cloning and clone screening. Transfection efficiency with the Nucleofector technology about 60% GFP positive cells, followed by FACS sorting, selection of high producing cell clones. The set up increased the average of productivity and reduced the time for clone screening and shortened the development process.


Cell line generation for production of biopharmaceuticals in mammalian cells usually involves intensive screening of clones to identify the rare high producers. In order to facilitate efficient and selective fluorescence activated cell sorting (FACS) based enrichment and cloning of antibody producing CHO cells, we developed a special vector setup by inserting a leaky translation termination signal between the heavy chain of an IgG antibody and an IgG transmembrane domain. Partial read-through during translation of the antibody heavy chain leads to display of a subset of the produced antibody on the surface of the expressing cell. We could show that the level of surface expression correlates well with the productivity. By applying FACS, high producing cells can be selectively enriched and cloned. Two sequential FACS enrichment cycles were performed which led to more than eightfold increased productivities of transfected and selected cell populations without cloning. The combination of selective FACS enrichment and FACS cloning with the new vector setup led to a sevenfold higher average productivity of the resulting clones as compared to a reference vector. Productivity and production stability assessment of clones generated with the new vector showed no negative impact of the co-expression of transmembrane antibody. Clone productivities of 4?g/L in a generic shake flask fed-batch model were achieved. Thus, this new vector setup facilitates fast and selective isolation of high producing production cell lines and allows significant reduction of clone screening efforts during cell line development for production cell lines. Additionally, the high productivity of FACS-enriched but non-clonal cell populations supports rapid, high yield, and cost efficient material production in early project phases.