Peripheral Blood Macrophages

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qiancheng
qiancheng's picture
Peripheral Blood Macrophages

I am working with macrophages.  We get  them from peripheral blood from water buffalo but the yield is frequently inadequate.

We want to use macrophage colony-stimulating factor (M-CSF) to stimulate generation of  macrophages from mononuclear cells but I do not know the concentration of M-CSFto use and the working concentration of M-CSF to use with PBS or cell culture medium.

I am a student, thank you

marcus muench
marcus muench's picture
 I generally use M-CSF at

 I generally use M-CSF at 20ng/ml, which is probably in excess for many applications.  The cell density, length of culture all effect how much you should add and how frequently because the cells will consume the M-CSF over time.  In a short term culture, you may just do fine with 2-5ng/ml.  the best way to answer your question is to perform a titration in your specific application.

I dilute the cytokine in medium+some protein (2% BSA in my case) when I aliquot and freeze (-80C).  I make stock solutions of 100X.  I would recommend using whatever medium and FBS, if that is what your using in your cultures, to dilute and store your cytokine.  

heehawmcduff
heehawmcduff's picture
Hi,

Hi,

I've never used it myself but I just spoke to a colleague who also used it at 20ng/ml in RPMI + 2% HI FBS for a 5 day incubation before washing.  Also seen papers that have used as high as 100ng/ml but that's probably too excessive.

Is it human M-CSF that you are using and does it have a good homology with water buffalo M-CSF?

qiancheng
qiancheng's picture
 thank you for your message!

 thank you for your message! I will try it and choose .

qiancheng
qiancheng's picture
It is murine M-CSF.

It is murine M-CSF.

I do not know whether it have a good homology or not ! 

marcus muench
marcus muench's picture
 Given the questionable

 Given the questionable species cross-reactivity, I really think your best plan of action is to perform a dose titration.