assays to assess cancer cell growth? - (May/24/2005 )
Hi everyone,
I’m in cancer research and I have currently produced some expression microarray results and I’m now ready for further cellular/molecular studies (which I’m unfamiliar with). I have found some potential genes of interest and I’m testing to see whether they are dependent of another gene (gene A, let say) and its pathway.
So my question is what kind of assays is good to test this hypothesis? I think I will transfect cell lines w/ the potential genes of interest and perform growth/invasion assays, as well as using siRNA to downregulate either the genes of interest or gene A, and see what the effects are. Or maybe I’m not specific enough?
I would like to do more background reading or various assays so any suggestions would be appreciated. Thanks in advance.
I’m in cancer research and I have currently produced some expression microarray results and I’m now ready for further cellular/molecular studies (which I’m unfamiliar with). I have found some potential genes of interest and I’m testing to see whether they are dependent of another gene (gene A, let say) and its pathway.
So my question is what kind of assays is good to test this hypothesis? I think I will transfect cell lines w/ the potential genes of interest and perform growth/invasion assays, as well as using siRNA to downregulate either the genes of interest or gene A, and see what the effects are. Or maybe I’m not specific enough?
I would like to do more background reading or various assays so any suggestions would be appreciated. Thanks in advance.
Anchorage independent growth assays (growth in agar) and invasion assays are both quite good assays to use, and well established techniques. Try a pubmed or google search on these terms.
Also, although tedious, simple growth calculations can be informative - counting your cells each time you replate them out and calculating doubling times.
You could also check for apoptosis levels using tunnel staining if that's of relevance to your pathway.
Using siRNAs as a tool to validate your target genes found in microarray analysis is a good plan. Many research groups are taking this route and have obtained very extensive data from this.
There are many commercial cell based assays, and the thing you want to do is make sure you formulate your questions precisely before doing the assays.
Use a good positive control for the cell based assay you choose to use. Have a reagent or chemical known to affect growth/invasion. That way you can set a standard for which you are looking for changes.
You will also want to normalize for cell number when you interpret your data.
Hope this helps!
yep. What those guys said.
Thanks for all the help! I'll keep all those things in mind!! 