All depends of what point you want to make. and you are right about 48hrs  in same cases everything is back to normal. However if you want to show let's say that cells A have an excessive and prolonged response over cells B, then you can go up to 72hrs (if look at my publication you see that is the case)

If looking for gene/protein expression and you have no idea I would use an early time point 30min (TNF for example is the early cytokine to be secreted around 15min) and at late time point 24hrs. 

If working with primary microglia and you don't have luxuray to do multiple points, then  the last thing you can do is do 6hrs (I am currently working with primary retinal microglia and 6hrs seems to be good time for what we are after).

good luck. aicha

Thank you Aicha. That Indeed was helpful.

Since my question doesnt involve comparative expression over time, I shall start with 24 hours.

Much appreciated. Thanks again

It depends of cytokine you want to study. There are early, intermediate and late response cytokines. Can you do a time course for your cytokine? 48 hours seems too late for me.

I agree with Mikhail. If you are not  lacking any of your resources, try  30 minutes, 6 hr, 12 ,24 and 48 h.

What cytokine(s) you are interested? I estimated expression kinetics for many cytokines. What is (are) your stimuli?

I do agree with Valsamma and Mikhail. I gathered a lot of data on pro-and anti-inflammatory cytokine expression in different cell types and systems.  Depending also on your stimuli and the cytokine of interest, you will find a difference in kinetics of the cytokine expression. However running a time course like Valsamma suggested will prevent from missing any relevant data. 

Thank you very much Mikhail, Valsamma and Marietta,

I am studying IL6, TNFalpha, IFN gamma and IL10 currently. Both protein as well as mRNA expression, in Astrocytes and microglia. 48 hrs  does give some results in protein expression, but i doubt that might not be optimal. IL6 TNF and IFN mostly fall in the immediate cytokine category. Again, what could be the ideal time point for studying m RNA expression for these early cytokines ?

Mikhail,

Our stimulus is the CSF from patients suffering from neurodegeneration (ALS). Previously we have shown the effect in neurons post 48 hrs. However, for cytokines in glia,i realise that is not the right interval. an extended time point analysis is not feasible presently. Please suggest.

Thanks.

The kinetics for mRNA and protein detection will be different and will vary by cytokine. A time course to determine the kinetics will be the best way to determine the ideal time to measure. Taking the time to do the time course will give you (and reviewers) increased confidence in the results of the study.

Well, as all people here suggested, time course is the best solution. I would advise you at least two time points, 4-6 and 18-24 hours. You may see upregulation of TNF as early as 30 minutes post stimulation, peaking at 1-2 hours and decreasing expression starting 4 hours post stimulation. If you don't see TNF upresulation at early time point, I would not exprect to see it at later time points.  IL6 and IFNg we see withing 2 hour of stimulation peaking at 6-8 hours, then decreasing as well. IL10 expression starts slightly after IL6 but well detected 2 hours post stimulation. Of course, proteins release will be shifted for 2-4 hours. If you unable to do a time course, I would favour an early time point for genes expression, 6-8 hours post stimulation.

Instead of doing one time course (48hrs), you can have a time-point kinetics, starting from 6hrs to 72 hrs. So it will cover all your early and late cytokines depending upon the type of stimulation and also give you a brief overview which cytokine is imp at what time point which is different from cell to cell. 

Thanks all!

Your suggestions have been helpful. I have started the analysis for 6 hrs (m-RNA) expression, and 24 hrs(protein expression) to begin with. will surely do a time point analysis, (at least at the two time points suggested). Thanks much indeed.