Calcium Phosphate Precipitation

Andrew Doherty A.Doherty at Bris.ac.uk
Mon Feb 17 03:39:22 EST 1997


mennow at ruly46.medfac.leidenuniv.nl wrote:
> 
> Does anyone have experience with calcium phosphate precipitating DNA to macrophages (J774).
> I have been trying to get a GFP vector in the cells last weeks, for transient expression. But I never got any signal.
> While the vector works perfectly in ES-cells. Is it possible that the macrophages just eat the calcium phosphate particles with the DNA?
> I want to use the GFP as a marker to check transfection together with a macrophage specific construct.
> Does anyone have any suggestions on this, or other ways to test a macrophage specific construct in vitro.
> 
> Menno de Winther
Hi Menno,

I've never used macrophages for transient expression as we use HEK 293's
as a rule. But I foulnd that the same vector can give very different
expression levels in different cell lines. We get transfection
efficiencies of up to 40% with HEKs but only 1-2% with CHO, for
instance. So it may simply be that your macrophages don't like the
promoter on that vector. Have other vectors been used on your cell line?

HOpe it helps
Andy D
-- 
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Dr Andrew Doherty		email -  a.doherty at bris.ac.uk
Dept. Anatomy			Tel (0117)9287421
School of Medical Sciences	Fax (0117)9287402
University of Bristol
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