Efficient selection of high-producing subclones during gene amplification of recombinant Chinese hamster ovary cells by flow cytometry and cell sorting

2001 ◽  
Vol 77 (1) ◽  
pp. 118-118
Author(s):  
Nicole Borth ◽  
Max Zeyda ◽  
Renate Kunert ◽  
Hermann Katinger
Keyword(s):  
Flow Cytometry ◽  
Gene Amplification ◽  
Chinese Hamster Ovary ◽  
Cell Sorting ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Ovary Cells ◽  
Efficient Selection ◽  
Selection Of

Efficient enrichment of high-producing recombinant Chinese hamster ovary cells for monoclonal antibody by flow cytometry

2015 ◽  
Vol 120 (3) ◽  
pp. 340-346 ◽  
Author(s):  
Takeshi Okumura ◽  
Kenji Masuda ◽  
Kazuhiko Watanabe ◽  
Kenji Miyadai ◽  
Koichi Nonaka ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Flow Cytometry ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Ovary Cells

scholarly journals Characterization by flow cytometry of fluorescein-methotrexate transport in chinese hamster ovary cells

Cytometry ◽  
1989 ◽  
Vol 10 (1) ◽  
pp. 50-55 ◽  
Author(s):  
Yehuda G. Assaraf ◽  
Larry C. Seamer ◽  
Robert T. Schimke
Keyword(s):  
Flow Cytometry ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Ovary Cells

scholarly journals Selection of mutant Chinese hamster ovary cells altered glycoproteins by means of tritiated fucose suicide.

1981 ◽  
Vol 1 (10) ◽  
pp. 902-909 ◽  
Author(s):  
C B Hirschberg ◽  
R M Baker ◽  
M Perez ◽  
L A Spencer ◽  
D Watson
Keyword(s):  
Chinese Hamster Ovary ◽  
Gel Electrophoresis ◽  
Trichloroacetic Acid ◽  
Chinese Hamster Ovary Cells ◽  
Sodium Dodecyl ◽  
Chinese Hamster ◽  
Wild Type ◽  
Ovary Cells ◽  
Replica Plating ◽  
Selection Of

Mutant Chinese hamster ovary cells altered in glycoproteins have been isolated by selecting for ability to survive exposure to [6-3H]fucose. Mutagenized wild-type cells were permitted to incorporate [3H]fucose to approximately 1 cpm of trichloroacetic acid-insoluble radioactivity per cell and then frozen for several days to accumulate radiation damage. The overall viability of the population was reduced by 5- to 50-fold. Four consecutive selection cycles were carried out. The surviving cells were screened by replica plating-fluorography for clones showing decreased incorporation of fucose into trichloroacetic acid-insoluble macromolecules. Considerable enrichment for cells deficient in fucose uptake or incorporation into proteins (or both) was found in populations surviving the later selection cycles. Two mutant clones isolated after the fourth selection cycle had the same doubling time as the wild type, but contained only 30 to 40% as much fucose bound to proteins as the wild type. Sialic acid contents of the mutants and the wild type were similar. The mutants differed quantitatively and qualitatively from the wild type and from each other with respect to total glycoprotein profiles as visualized by sodium dodecyl sulfate gel electrophoresis. Differences were also found in resistances to cytotoxicity of lectins such as concanavalin A and wheat germ agglutinin.

Selection of mitotic Chinese hamster ovary cells from microcarriers

1980 ◽  
Vol 109 (2) ◽  
pp. 231-238 ◽  
Author(s):  
John J.Y. Ng ◽  
Charles L. Crespi ◽  
William G. Thilly
Keyword(s):  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Ovary Cells ◽  
Selection Of

Amplification and expression of human alpha-globin genes in Chinese hamster ovary cells

1984 ◽  
Vol 4 (8) ◽  
pp. 1469-1475
Author(s):  
Y F Lau ◽  
C C Lin ◽  
Y W Kan
Keyword(s):  
Gene Expression ◽  
Gene Amplification ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Globin Gene ◽  
Chinese Hamster ◽  
Globin Genes ◽  
Ovary Cells ◽  
Alpha Globin ◽  
Globin Gene Expression

We studied the effects of gene amplification on human globin gene expression in Chinese hamster ovary cells. The normal human alpha-globin gene (N alpha 2) and a hybrid gene (M alpha G) containing the 5' promoter-regulator region of the mouse metallothionein gene and the human structural alpha 2-globin gene were linked to a modular SV2-cDNA dihydrofolate reductase (DHFR) gene. The recombinant DNA molecules were introduced into Chinese hamster ovary cells by calcium phosphate precipitation. After initial selection to retain the DHFR and linked sequences, the cells were cultured in increasing concentrations of methotrexate up to 0.2 mM. Southern blot analysis of total cellular DNA showed an approximately 500- to 1,000-fold increase in the number of copies of DHFR and human alpha-globin genes. The transcription of the alpha-globin and DHFR genes increased as their copy number within the cells increased. The transcription of the amplified hybrid M alpha G gene was also inducible with cadmium treatments. Both mature mRNA and "read-through" transcripts were observed. DHFR constituted approximately 10% of pulse-labeled cellular proteins in these cells, but no human alpha-globin was detected. In vitro translation of polyadenylated RNA from these cells showed that alpha-globin mRNA transcribed from the amplified alpha-globin genes was functional and directed alpha-globin chain synthesis. In situ hybridization of 3H-labeled alpha-globin and DHFR DNA probes in chromosome preparations from the two cell lines indicated that both genes were coamplified in the same chromosomal locations in each cell type. These results indicate that gene amplification enhances human globin gene expression in cultured Chinese hamster ovary cells.

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