Scientist Solutions - culture of primary mature adipocyte?

laurence Posted 7/1/2009 8:27:18 PM	Dear collegues, I would like to do a coculture macrophage/mature adipocyte to measure changes in the insulin induced glucose uptake in the adipocytes. In most of the articles I saw, scientists used either 3T3L1 or adipocytes differentiated from the stromal vascular fraction. I was just wondering if some of you had ever tried the culture of freshly isolated mature "floating" adipocytes. If yes how long do they survive in culture? Is there a special way to culture them?and finally, do they keep their functional properties? Thanks a lot in advance, Laurence
cqrogers Posted 9/30/2009 10:06:14 AM	Hi Laurence. Did anyone ever get back with you on this topic? I am also having problems culturing the "floating" adipocytes. Thanks. CQ
R Bishop Posted 9/30/2009 10:22:54 AM	CQ, Are you adding adenosine to suppress lipolysis? That is a critical addition to both the Adipocyte Incubation Solution and media. Laurence, Im sorry we missed your post. The answer is yes you can culture primary adipocytes in "ceiling cultures". This takes advantage of the "floating" adipocyte by allowing it to attach to the top of the tissue culture flask by filling the flask with media. They do keep many functional properties including lipolysis. There are several papers on this technique, you might try this one . http://joe.endocrinology-journals.org/cgi/reprint/164/2/119 Bishop
Pippuri Posted 9/30/2009 10:28:41 AM	Hi there, In my hands, fully differentiated adipocytes from 3T3-L1 look a bit like floating in the culture dish (imagine you are looking down from the sky and see a ballon floating in the air which is attached by a string to the ground)... Thus, my question is, when you mentioned 'floating' adipocytes, do you mean that they won't attached at all? As far as I understand, most of the procedure to isolate pre-adipocytes and differentiate them into mature adipocytes in the culture dish. It is very difficult to directly culture mature adipocytes as they tend to burst during the handling process.... If you are working with human primary adipocytes, there is a well-established protocol from this company, which states that the cells will stay good for 4 weeks after post-differentiation. But it may depends on whether your assay is sensitive to the age of the cells. Good luck.
laurence Posted 9/30/2009 5:43:48 PM	Hi all, thanks for all the reply. I finally chose the 3T3L1 differentiation option instead of the mature adipocyte. Laurence