I want to assess the cell migration rate through scratch wound assay for the hydrogel. I have read the literature and in some research articles it is mentioned that DMEM media was not supplemented with FBS (10%) as it supports the growth of the cells so it will effect the results. But in other studies only term culture media has been used. Kindly guide me whether FBS should be added or not into the DMEM.
Thanks in advance for your guidance and time.
Hello Noor Ul Ain
For the scratch-wound assay, you may supplement DMEM culture media with reduced serum (1%). This will not only help the cells to survive but also keep cell proliferation to a minimum.
Best.
Hi, culture your cell under low serum 1% for 48 hours. I did this test in my thesis. Check the paper link below if you'd like.
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6558512/
I've been doing what Malcolm Nobre said, and it's been working pretty well. If you are testing some kind of drug or compound, it is also important to use it at a sub-lethal concentration.
At high concentrations, you might observe migration inhibition due to cell death processes, or you may observe cells dying.
Julio Thurler I want to observe the effect of the treatment compound that it is promoting the cell migration and faster than the control. So in control as well serum concentration should be 1% ?
Noor Ul Ain Yes! The conditions in treated and untreated groups must be the same. If you want, you can make a control with 10% FBS just to observe the difference.
Hi Noor Ul Ain,
If you are incubating the cells for shorter time points (less than 12 hrs), keeping the 10%FBS is fine, but for longer time incubation for scratch close, it is recommended to keep them in low serum media. I always incubate cells at least 24 hrs in 0.2% FBS media before introducing the stratch. You can see the figure below from my publication to get a feel on how the growth curves changes with/without Serum.
Really interesting results, Anusi Sarkar ! I will definitely try this starvation before scratch initiation. Were the treatments also diluted in 0.2% FBS?
Have these results already been published? If so, could you please provide the paper?
Thank you!
Hi Julio Thurler , thanks for your interest.
yes, All the treatments were given in 0.2%FBS supplemented media. and this graph suggest, whatever, wound healing happened during the next 36hrs were mostly (if not solely) due to the cell migration and NOT by the cell duplication.
the data are published. You can check the link below:
Article Recombinant human PRG4 (rhPRG4) suppresses breast cancer cel...
This particular growth curve is in supplementary figure 1. and teh associated migration assay data are in Figure 2 in the main article. the Method section contains the details of the protocol followed.
Thanks,
Anusi Sarkar Thank you so much for your valuable advice. I will design experiments acordingly.
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