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How to concentrate conditioned medium? - (Sep/22/2006 )

Dear all,

is anybody experienced in concentrating conditioned medium for cell culture and could give me some general hints about it, I´m quite unexperienced in that, so every hint will be helpful!

Thanks in advance.
All the best and a nice weekend

-Bomber-

if you have the opportunity of large-scale lyophilasation I would try; you may look for precipitates

-The Bearer-

bomber, we have used that method to detect differences in expression of low-prevalence secreted peptides. we have done it two different ways:

1. those little centricon MW cutoff filters; you pick the correct cutoff and spin the medium until it concentrates

2. we have also used a vacuum-driven slot-blot technique, for Westerns. we add a whole bunch of conditioned medium to the wells and it all concentrates into one spot on the membrane for further blotting

I suppose the most important question, what is your application?

-aimikins-

Thanks for input everybody.

Aimkins, I would need to concentrate conditioned medium, and use this medium for further cell culture.
I think the cut-off-filters are suited. I think about using, let´s say 10kDa cut-off or even smaller. I tried to look up some company where I could get these filters (e.g Filtron MacroSep or similar) but without success so far.
Do you have any recommendations maybe, or could spare any hints for my purpose?

Thanks everybody for sharing ideas,
all the best
Cheers

-Bomber-

I do not know what sort of volumes you are talking, so this may or may not help you...

we use microcons, made by Amicon (which is really made by Millipore, I think). you can buy them from Fisher, and I'm sure there are other suppliers if you want to find the cheapest.

we use model YM-3, which has a 3kD cutoff range. we used it for 3.5kD peptides with pretty good success. here is a link to the product spiel page

-aimikins-

Hi Aimkins,

thanks for your help!
I took a look at the Millipores´ website, seems quite suitable. For my puspose I think I will try to get conditioned medium of some ml and store it.

All the best,
Cheers

-Bomber-

Hi, Bomber.

The centriprep (millipore) can be a good option if you have a volume until 15 mL. The maximum concentration capacity is close to 25X. There are different cut-off membrane: 3, 10, 30, 100 KDa. In the millipore site you can find a pdf manual with the instructions and with recomendations about compatible centrifuges.

Good Luck,
Patricia.

-patfidelis-

Hi patfidelis,

sorry for getting back late; thanks for your input.
I will have a look at the recommended sites; 15 ml sounds very good to me... .

All the best
Cheers

-Bomber-

QUOTE (Bomber @ Sep 22 2006, 05:36 PM)
Dear all,

is anybody experienced in concentrating conditioned medium for cell culture and could give me some general hints about it, I´m quite unexperienced in that, so every hint will be helpful!

Thanks in advance.
All the best and a nice weekend



you can use the cut -off column for concentrating the conditioned medium , and then lyophilise it

-Amit nayak-