Scalable, Serum-Free, High Efficiency Transfection of CHO Cells Using an Economical DNA Transfer Vehicle

2005 ◽  
pp. 629-632
Author(s):  
P. Girard ◽  
M. Derouazi ◽  
F. Van Tilborgh ◽  
F.M. Wurm
Keyword(s):  
Cho Cells ◽  
High Efficiency ◽  
Dna Transfer ◽  
Serum Free

scholarly journals Establishment of fast-growing serum-free immortalised cells from Chinese hamster lung tissues for biopharmaceutical production

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Noriko Yamano-Adachi ◽  
Rintaro Arishima ◽  
Sukwattananipaat Puriwat ◽  
Takeshi Omasa
Keyword(s):  
Cell Line ◽  
Cho Cells ◽  
Mammalian Cells ◽  
High Efficiency ◽  
Chinese Hamster ◽  
Serum Free ◽  
Sterility Testing ◽  
Fast Growing ◽  
Defined Culture

Abstract Chinese hamster (Cricetulus griseus) ovary-derived Chinese hamster ovary (CHO) cells are the most commonly used mammalian hosts for the industrial production of recombinant therapeutics because of their ability to fold, assemble, and perform post-translational modifications, such as glycosylation, on proteins. They are also valuable for their ability to grow in serum-free suspension cultures. In this study, we established a cell line derived from lung tissue of Chinese hamsters, named Chinese hamster lung (CHL)-YN cells. The biosafety of CHL-YN cells was confirmed by in vitro sterility testing, mycoplasma detection, and reverse transcriptase assays. One of the key characteristics of CHL-YN cells was their doubling time of 8.1 h in chemically defined culture medium; thus, they proliferate much faster than conventional CHO cells and general mammalian cells. Transgenes could be introduced into CHL-YN cells with high efficiency. Finally, between 50% to > 100% of the amount of glycosylated immunoglobulin G (IgG)1 produced by CHO-K1 cells was produced by CHL-YN cells over a shorter period of time. In summary, fast-growing CHL-YN cells are a unique cell line for producing recombinant proteins.

Sodium Chloride Modified Silica Nanoparticles as a Non-Viral Vector with a High Efficiency of DNA Transfer into Cells

2003 ◽  
Vol 3 (3) ◽  
pp. 273-279 ◽  
Author(s):  
Yuxiang Chen ◽  
Zhigang Xue ◽  
Duo Zheng ◽  
Kun Xia ◽  
Yanzhong Zhao ◽  
...  
Keyword(s):  
Sodium Chloride ◽  
Silica Nanoparticles ◽  
High Efficiency ◽  
Viral Vector ◽  
Dna Transfer ◽  
Modified Silica ◽  
Modified Silica Nanoparticles

Serum-free large-scale transient transfection of CHO cells

2004 ◽  
Vol 87 (4) ◽  
pp. 537-545 ◽  
Author(s):  
Madiha Derouazi ◽  
Philippe Girard ◽  
Fr�d�ric Van Tilborgh ◽  
Keyvan Iglesias ◽  
Natalie Muller ◽  
...  
Keyword(s):  
Cho Cells ◽  
Large Scale ◽  
Transient Transfection ◽  
Serum Free

scholarly journals Efficient differentiation of beige adipocytes from adult human adipose-derived stem/stromal cells

2020 ◽  
Author(s):  
Amar M. Singh ◽  
Liang Zhang ◽  
John Avery ◽  
Stephen Dalton
Keyword(s):  
Stem Cells ◽  
Cell Therapy ◽  
Drug Screening ◽  
Stromal Cells ◽  
High Efficiency ◽  
Disease Modelling ◽  
Serum Free ◽  
Beige Adipocytes ◽  
Free Media ◽  
Stromal Stem Cells

Abstract Beige adipocytes (also known as brite adipocytes) have significant utility for numerous applications, such as drug screening, cell therapy, and disease modelling. However, a high-efficiency protocol from human adult adipose-derived stem/stromal stem cells (ADSC) has not been described. The protocol described here achieves beige adipocyte purities of >92% in a fully-defined, serum-free media cocktail, which enables these downstream applications. This method provides a significant leap forward over previously described, serum-based protocols that were inconsistent and inefficient.

High efficiency recombinant factor VIII production and secretion in CHO cells

2015 ◽  
Author(s):  
Abdulmecit Gokce ◽  
Kaya Molo ◽  
Elif Gelmez ◽  
Gursel Turgut ◽  
Gulay Bulut
Keyword(s):  
Factor Viii ◽  
Cho Cells ◽  
High Efficiency ◽  
Recombinant Factor Viii

Serum-Free Transfection of CHO Cells with Chemically Defined Transfection Systems to Generate Transient and Stable Cell Lines

2010 ◽  
pp. 475-478
Author(s):  
Reisinger Hannes ◽  
Vorauer-Uhl Karola ◽  
Steinfellner Willibald ◽  
Wagner Andreas ◽  
Katinger Hermann ◽  
...  
Keyword(s):  
Cell Lines ◽  
Cho Cells ◽  
Serum Free ◽  
Stable Cell Lines

PRODUCTION AND CHARACTERIZATION OF A RECOMBINANT FACTOR VIII EXPRESSED BY CHO CELLS CULTURED IN SERUM SUPPLEMENTED OR IN SERUM FREE MEDIA

1992 ◽  
pp. 152-154
Author(s):  
G. MIGNOT ◽  
N. BIHOREAU ◽  
P. PAOLANTONACCI ◽  
A. SAUGER ◽  
V. TOULLY ◽  
...  
Keyword(s):  
Factor Viii ◽  
Cho Cells ◽  
Recombinant Factor Viii ◽  
Serum Free ◽  
Free Media ◽  
Cells Cultured

scholarly journals Polyamine starvation prolongs the S and G2 phases of polyamine-dependent (arginase-deficient) CHO cells.

1984 ◽  
Vol 4 (5) ◽  
pp. 915-922 ◽  
Author(s):  
S Anehus ◽  
P Pohjanpelto ◽  
B Baldetorp ◽  
E Långström ◽  
O Heby
Keyword(s):  
Cell Cycle ◽  
Ornithine Decarboxylase ◽  
Cho Cells ◽  
Decarboxylase Activity ◽  
Serum Free Medium ◽  
Free Medium ◽  
Cho Cell ◽  
Irreversible Inhibitor ◽  
Serum Free ◽  
Cell Variant

This study analyzes the effects of polyamine starvation on cell cycle traverse of an arginase-deficient CHO cell variant (CHO-A7). These cells grow well in serum-free medium, provided that it contains ornithine or polyamines or both. In the absence of ornithine or polyamines or both, the CHO-A7 cells develop severe polyamine deficiency and, as a consequence, grow more slowly. When grown to a stationary phase in the presence of ornithine or putrescine or both, the CHO-A7 cells became arrested in G0/early G1. However, when starved for ornithine and polyamines, they accumulated in the S and G2 phases. Ornithine and polyamine starvation of CHO-A7 cells causes an increase in ornithine decarboxylase activity. When this increase was prevented by treatment with DL-alpha-difluoromethylornithine, an enzyme-activated irreversible inhibitor of ornithine decarboxylase, growth was further suppressed, and a greater fraction of cells were found in the S and G2 phases of the cell cycle.

Expression of PRV-1 Decreases Dependency of Cells on Growth Factors for Proliferation.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 655-655
Author(s):  
Vahid Afshar-Kharghan ◽  
Jun Li ◽  
Zakar Mnjoyan
Keyword(s):  
Growth Factors ◽  
Cho Cells ◽  
Growth Media ◽  
Maximal Effect ◽  
Serum Free Medium ◽  
Free Medium ◽  
Transfected Cells ◽  
Serum Free ◽  
Cell Lysate ◽  
Cpg Dinucleotide

Abstract Polycythemia rubra vera-1 (PRV-1) is a GPI-linked protein expressed on surface of neutrophils. Polycythemia vera and essential thrombocythemia were found to be associated with an increase in the level of PRV-1 mRNA; however, the function of PRV-1 remains unknown. We have studied the functional effect of expression of PRV-1, both in heterologous cell line and in neutrophils. cDNA encoding PRV-1 was subcloned from a leukocyte cDNA library and expressed in Chinese hamster ovary cells (CHO). After seeding equal numbers of CHO cells stably transfected with PRV-1 or empty plasmid, cell count was conducted at different time intervals and under different concentration of fetal bovine serum (FBS) in growth media. The numbers of PRV-1 transfected cells were higher than sham-transfected cells at 24, 48, 96 and 144 hrs after seeding, and this difference was enhanced with cells growing in a medium with a reduced concentration of serum with the maximal effect seen in serum-free medium. The percentage of apoptotic cells was similar between PRV-1 and sham-transfected cells. These results are suggestive of that CHO cells expressing PRV-1 proliferate faster than sham-transfected cells. The difference in proliferation rate was more significant at lower concentrations of serum and was the highest in serum-free medium. An immunoblot analysis with anti-phosphotyrosine antibody on the whole cell lysate demonstrated that the presence of PRV-1 alters cell signaling. A 60kDa protein band that disappeared after removal of serum in growth medium of sham-transfected cells continued to be present in the PRV-1 transfected cell lysate regardless of the presence or absence of FBS in the media. In order to understand the function of PRV-1 in neutrophils, we used the fact that in 85% of individuals PRV-1 is expressed only by a subgroup of neutrophils. We separated PRV-1 expressing neutrophils of an individual from those lacking this protein in the same donor, using anti-PRV-1 monoclonal antibody. We then compared the gene expression profile of the two groups of neutrophil using DNA microarray technique. Expression of PRV-1 was associated with several folds increase in expression of growth factors (fracture callus-1, X 14.9 times; Insulin-like 3, X 6.7; neural proliferation differentiation, and control-1 or NPDC1, X 3.4 times) and post-translational modifying enzyme of cell surface protein (N-acetylase N sulfotransferase-1, X 18.9 times). Under physiologic conditions, transcriptional regulation of the PRV-1 gene limits its expression to only a certain percentage of neutrophils. In order to investigate the role of epigenetic factors in regulation of transcription of the PRV-1 gene, we analyzed methylation of CpG dinucleotide in promoter of the PRV-1 gene in the genomic DNA obtained from neutrophils expressing PRV-1 to those that don’t express this protein in the same individual. Expression of PRV-1 was associated with a decrease in methylation of the CpG dinucleotide located 2937 bp before initiation codon (45% methylated in PRV-1 negative neutrophils compared to 30% methylated in PRV positive neutrophils). We are currently studying the methylation pattern of CpG islands inside the PRV-1 gene. Studying the effect of DNA methylation on expression of PRV-1 in physiologic conditions, may shed light to the mechanism of overexpression of PRV-1 in MPD.

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