CHO Cell Transfection: Reagents, Methods, Protocols
CHO Cells
CHO cells (Chinese Hamster Ovary cells) are a laboratory-cultured cell line derived from cells of the ovaries of Chinese hamsters. Chinese hamsters are a popular laboratory mammal, partially due to their small size and low chromosome number, which makes them a good model for tissue culture and radiation studies. CHO cells are a commonly-used mammalian cell model used in biology, medical and pharmaceutical research. In addition, they are frequently used for commercial purposes to manufacture therapeutic recombinant proteins. Growing demand for therapeutic proteins has led to the development of technologies that promote high quality and productivity in CHO expression systems. The CHO cultivation process is well established and the cost of using CHO cells for therapeutic protein production is equivalent to that of microbial culture.
CHO Cell Line Origin
CHO is an acronym for Chinese Hamster Ovary, which alludes to the origin of the CHO cell line, a line that has become a hugely popular research tool in the molecular biology community. The Chinese Hamster was brought to the US in 1948. In 1957, T. Puck obtained a Chinese hamster from a lab at the Boston Cancer Research Foundation, where he used it to derive the original CHO cell line. Since then, CHO cells have been found useful for many research purposes.
Characteristics of CHO Cells
Over the 60 year history of CHO cell studies, multiple different lines with different attributes have been derived from the original cell line. A common CHO derivative is CHO-K1, which has less DNA than the original CHO cells. A later mutation of CHO-K1 generated CHO-DXB11 (also known as CHO-DUKX), which lacks dihydrofolate reductase (DHFR) activity. Another offshoot of the original CHO cell line was CHO-pro3, which is proline-dependent. CHO-pro3 was then later mutated to create CHO-DG44, which also is DHFR-deficient (link).
CHO cells are epithelial-like cells that can be grown as adherent cells or in suspension. This cell line has a chromosome frequency distribution of 50 cells: 2n=22 and its stemline number is hypodiploid. CHO cells are widely used and well-characterized in methods for cell transfection, gene amplification, and clone selection. CHO has been well-established for regulatory approval for recombinant protein expression, in turn improving therapeutic efficacy, protein longevity, and reducing safety concerns by performing human-compatible post-translational modifications. Antibody glycosylation and protein sialylation are examples of strategies that have been developed to enhance the post-translational modifications of CHO cells.
The CHO cell line now plays a dominant role in bioprocessing research and the development of therapeutic biopharmaceuticals. CHO cells are important in research for their stability due to a low rate of spontaneous transformation. Also, the biomanufactured recombinant proteins generated by CHO cells are functionally and structurally similar to native proteins. These include CD4 cell surface antigen, interferon, factor VIII and prorenin.
DHFR-deficient CHO Cell Lines and Stable Transfection
The DHFR-deficient strains require supplementation with glycine, hypoxanthine and thymidine. These strains were used to demonstrate that an exogenous DHFR gene could be stably transfected and selected using glycine/hypoxanthine/thymidine deficient (i.e. GHT-minus) media into cells that would otherwise be DHFR-deficient. This selection method has become a standard method for the establishment of stable transfection in CHO cell lines intended for production of therapeutic proteins. A gene that expresses the protein of interest and the DHFR gene are either combined in one mammalian expression vector or put into separate vectors. The plasmid or plasmids then are transfected into the CHO cells and the cells are grown in GHT-minus media to provide a selective environment. As a result, all surviving cells have copies of the DHFR gene and the gene of interest integrated into their genome.
Links
Chinese Hamster Ovary (Wikipedia)
CHO Transfection Kit – Commercially available from Altogen Biosystems
CHO Cell Line – Commercially available from ATCC
CHO Xenograft Studies – In Vivo Laboratory Contract Research by Xenograft Labs
Stable CHO RNAi Cell Line Generation – Gene silencing in CHO cells
In Vivo Transfection Products – Commercially available from Altogen Biosystems
CHO Cells I Cell Culture I CHO Transfection Protocols, Kits, Methods I Get CHO Transfection Reagent