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Titel    HEK 293 cells
Verlag    (Wikipedia)
Jahr    2007
URL    http://en.wikipedia.org/w/index.php?title=HEK_293_cells&oldid=154716929

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Fragmente    4


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[1.] Ww/Fragment 024 11 - Diskussion
Zuletzt bearbeitet: 2014-10-28 03:02:44 Hindemith
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Quelle: Wikipedia HEK 293 cells 2007
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1.4 293FT cell lines

Human Embryonic Kidney cells, also known as HEK cells, HEK 293 or just 293 cells, are a cell line originally derived, as their name indicates, from embryonic human kidney. HEK cells are very easy to grow and transfect very readily and so are widely-used in cell biology research. They are also used by biotechnology industry to produce therapeutic proteins and viruses for gene therapy.

1.4.1 Origins of 293 cells

293 cells were generated by transformation of cultures of normal human embryonic kidney cells with sheared adenovirus 5 DNA in the laboratory of Alex Van der Eb in Leiden, Holland in the early 1970s. They are called HEK for human embryonic kidney, while the number 293 roots from numbering of experiments

Human Embryonic Kidney cells, also known as HEK cells, HEK 293 or just 293 cells, are a cell line originally derived, as their name indicates, from embryonic human kidney. HEK cells are not themselves particularly interesting, but are very easy to work with, and so are a widely-used cell line in cell biology research. [...]

[...]

HEK 293 cells were generated by transformation of cultures of normal human embryonic kidney cells with sheared adenovirus 5 DNA in the laboratory of Alex Van der Eb in Leiden, Holland in the early 70s.

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[2.] Ww/Fragment 025 24 - Diskussion
Zuletzt bearbeitet: 2014-10-28 03:05:18 Hindemith
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Subsequent analysis has shown that the transformation was brought about by an insert consisting of -4.5 kilo bases from the left arm of the viral genome, which became incorporated into human chromosome 19 (Louis et al, 1997).

For many years it was assumed that HEK 293 cells were generated by transformation of either a fibroblastic, endothelial or epithelial cell all of which are abundant in kidney. However the fact that the cells originated from [cultured kidney cells does not clearly indicate the exact cellular origin of the HEK 293, as embryonic kidney cultures may contain small numbers of almost all cell types of the body.]

Subsequent analysis has shown that the transformation was brought about by an insert consisting of ~4.5 kilobases from the left arm of the viral genome, which became incorporated into human chromosome 19 (Louis 1997[2]).

For many years it was assumed that HEK 293 cells were generated by transformation of either a fibroblastic, endothelial or epithelial cell all of which are abundant in kidney. However the fact that the cells originated from cultured kidney cells does not say much about the exact cellular origin of the HEK 293, as embryonic kidney cultures may contain small numbers of almost all cell types of the body.

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[3.] Ww/Fragment 026 01 - Diskussion
Zuletzt bearbeitet: 2014-10-28 03:11:42 Hindemith
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[However the fact that the cells originated from] cultured kidney cells does not clearly indicate the exact cellular origin of the HEK 293, as embryonic kidney cultures may contain small numbers of almost all cell types of the body. In fact Graham and coworkers more recently provided evidence that HEK 293 cells and several other human cell lines generated by adenovirus transformation of human embryonic kidney cells have many properties of immature neurons, suggesting that the adenovirus was taken up and transformed a neuronal lineage cell in the original kidney culture (Shaw et al, 2002).

1.4.2 Applications of 293 cells

As an experimentally transformed cell line, HEK cells are not a particularly good model for normal cells, cancer cells, or any other kind of cell that is a fundamental object of research. However, they are extremely easy to work with, being straightforward to culture and to transfect, and so can be used in experiments in which the behaviour of the cell itself is not of interest. Typically, these experiments involve transfection in a gene (or combination of genes) of interest, and then analyzing the expressed protein; essentially, the cell is used simply as a test tube with a membrane. The widespread use of this cell line is due to its extreme transfectability by the calcium phosphate method, achieving efficiencies approaching 100% as determined by FACS using a 2 x PBS buffer. A lower efficiency might be achievable with an HBS buffer.

An important variant of this cell line is the 293T cell line that contains, in addition, the SV40 large T antigen, that allows for episomal replication of transfected plasmids containing the SV40 origin of replication. This allows for amplification of transfected plasmids and extended temporal expression of the desired gene products. Note that any similarly domesticated cell line can be used for this fort of work; Hela, COS and Chinese Hamster Ovary cell are common alternatives. Examples of such experiments include: A study of the effects of drug on sodium channels; testing of an inducible RNAi system; testing of an isoform-selective protein kinase C agonist; investigation of the interaction between two proteins; analysis of a nuclear export signal in a [protein (He et al, 1998).]

However the fact that the cells originated from cultured kidney cells does not say much about the exact cellular origin of the HEK 293, as embryonic kidney cultures may contain small numbers of almost all cell types of the body. In fact Graham and coworkers more recently provided evidence that HEK 293 cells and several other human cell lines generated by adenovirus transformation of human embryonic kidney cells have many properties of immature neurons, suggesting that the adenovirus was taken up and transformed a neuronal lineage cell in the original kidney culture (Shaw et al. 2002[3]).

Uses of HEK 293 Cells

As an experimentally transformed cell line, HEK cells are not a particularly good model for normal cells, cancer cells, or any other kind of cell that is a fundamental object of research. However, they are extremely easy to work with, being straightforward to culture and to transfect, and so can be used in experiments in which the behaviour of the cell itself is not of interest. Typically, these experiments involve transfecting in a gene (or combination of genes) of interest, and then analysing the expressed protein; essentially, the cell is used simply as a test tube with a membrane. The widespread use of this cell line is due to its extreme transfectability by the calcium phosphate method, achieving efficiencies approaching 100% as determined by FACS using a 2XPBS buffer. A lower efficiency might be achievable with an HBS buffer.

An important variant of this cell line is the 293T cell line that contains, in addition, the SV40 large T antigen, that allows for episomal replication of transfected plasmids containing the SV40 origin of replication. This allows for amplification of transfected plasmids and extended temporal expression of the desired gene products. Note that any similarly domesticated cell line can be used for this sort of work; HeLa, COS and Chinese Hamster Ovary cell are common alternatives.

Examples of such experiments include:

  • A study of the effects of a drug on sodium channels [4]
  • Testing of an inducible RNA interference system [5]
  • Testing of an isoform-selective protein kinase C agonist [6]
  • Investigation of the interaction between two proteins [7]
  • Analysis of a nuclear export signal in a protein [8]
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[4.] Ww/Fragment 027 01 - Diskussion
Zuletzt bearbeitet: 2016-01-20 18:15:23 Schumann
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A more specific use of HEK cells is in the propagation of adenoviral vectors.

Viruses offer an extremely efficient means of delivering genes into cells, since this is what they have evolved to do, and are thus of great use as experimental tools. However, as pathogens, they also present a degree of danger to the experimenter. This danger can be avoided by the use of viruses which lack key genes, and which are thus unable to replicate after entering a cell. In order to propagate such viral vectors, a cell line that expresses the missing genes is required. Since HEK cells express a number of adenoviral genes, they can be used to propagate adenoviral vectors in which these genes (typically, E1 and E3) are deleted, such as AdEasy. Another application of 293, especially 293T, cells is commonly used for the production of lentiviral and retroviral vectors. Various retroviral and lentiviral packaging cell lines are based on these cells.

A more specific use of HEK cells is in the propagation of adenoviral vectors. Viruses offer an extremely efficient means of delivering genes into cells, since this is what they have evolved to do, and are thus of great use as experimental tools. However, as pathogens, they also present a degree of danger to the experimenter. This danger can be avoided by the use of viruses which lack key genes, and which are thus unable to replicate after entering a cell. In order to propagate such viral vectors, a cell line that expresses the missing genes is required. Since HEK cells express a number of adenoviral genes, they can be used to propagate adenoviral vectors in which these genes (typically, E1 and E3) are deleted, such as AdEasy (He 1998).

293, and especially 293T, cells are commonly used for the production of lentiviral and retroviral vectors. Various retroviral and lentiviral packaging cell lines are based on these cells.

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