siRNA Silencing in Primary Hepatocytes

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R Bishop
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siRNA Silencing in Primary Hepatocytes

Hello silencers out there.

We are trying to use siRNA to knock down expression of genes in primary mouse hepatocytes. Has anyone successfully used siRNA in primary hepatocytes? If so what was the transfection reagent or technique used to get the siRNA into the cells?

We are pretty good at isolating the hepatocytes from the mouse and growing them in culture. Our siRNAs come pre-validated from Ambion.

Thanks

Rb[b]

R Bishop
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Still looking for help on

Still looking for help on this experiment? Can anyone help please?

AUen
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I do not have any experience

I do not have any experience transfecting primary heps.

I asked around to a few colleagues and the best starting point I found was to go to the following link at the Qiagen site.

http://www1.qiagen.com/transfectionprotocols/?r=5618

This will allow you to select the cell line, nucleic acid, and culture format to receive a custom protocol. I did not see primary heps in the drop down list but maybe one of the other cell lines would give you a good starting point.

Big E
Big E's picture
Accell siRNAshttp://www
Jon Moulton
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This is probably too late to

This is probably too late to help, Rusty.  Sorry, I don't check this forum often  However, an alternative knockdown method is Morpholinos delivered by Endo-Porter:

Liu B, Paranjpe S, Bowen WC, Bell AW, Luo JH, Yu YP, Mars WM, Michalopoulos GK. Investigation of the Role of Glypican 3 in Liver Regeneration and Hepatocyte Proliferation. Am J Pathol. 2009 Jul 2. [Epub ahead of print]

Chiang PC, Hsu JL, Yeh TC, Pan SL, Guh JH. Elucidation of susceptible factors to endoplasmic reticulum stress-mediated anticancer activity in human hepatocellular carcinoma. Naunyn Schmiedebergs Arch Pharmacol. 2008 Apr;377(2):167-77. Epub 2008 Jan 29.

Hao Q, Hong SH, Maret W.Lipid raft-dependent endocytosis of metallothionein in HepG2 cells. J Cell Physiol. 2007 Feb;210(2):428-35.

Anyway, by now your data is likely already long in the bag.

  - Jon

R Bishop
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Thanks Jon.  We are still

Thanks Jon.  We are still working on hepatocytes, so I will keep this in mind.

Rusty

Jon Moulton
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Ma always tried to deliver

Ma always tried to deliver the hepatocytes with onions, but I was uncooperative.