Like MTT and MTS for Cell viability (cytotoxicity). This paper (http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0068808) discusses the following:
1. Wound healing migration assay
2. Migration chamber assay
3. Aggregation and Adhesion assays
4. Matrigel invasion assay
5. Gelatin zymography
6. Nitrate/nitrite production and luciferase activity
7. Trans-endothelial migration assay
Are these reliable proxies for anti-metastatic activity? My objective is to check some plant extracts for its anti-metastatic activity
Hi Felix,
All these assays you mentioned are standard and kind of complementary to each other and one single assay may not appear conclusive for EMT and cell migration.
One recent paper published in 2010 introduced an alternative method based on the 'Spindle Index', a novel measure of spindle phenotype, derived from the ratio of maximum length to maximum width of cells. The experimental data obtained for EMT from the newly described method have been validated by conventional morphological assessment and EMT immunocytochemistry with parallel experimental cross verification for the cell migration/metastasisby traditional matrigel invasiveness studies and scratch wound assay.
I am sharing this article (Cellular Oncology 32 (2010) 67–76) for further reading.
file:///H:/Researchgate%20materials/EMT%20and%20cell%20migration/Novel%20in%20vitro%20assays%20for%20the%20characterization%20of%20EMT%20in%20Tumorigenesis.pdf
Good luck!
In case you do not have access to the full paper I am sharing the pdf file for the article that I mentioned.
These assays are reliable with varying degree of their respective limitations. Out of mentioned assay most common, simple and cost effective assay is wound healing assay. You can also further strengthen your results by respective change of in EMT markers such as E-cadherin, N-cadherin, vimentin etc in response to your plant extract treatment.
Well, metastasis and EMT are two different things, although EMT is a part of metastatic program. Any of the given methods will not give you any clue if EMT is inhibited or not. Doing expression analysis for EMT markers is the best possible way. If you want to perform an indicative method for initial screening go for spheroid invasion assay. If you are testing only few compound wound healing assay and matrigel invasion assays will be best choice.
Dear Felix,
Two tiny comments from my parts:
1) MTT and MTS assays indirectly assess "cell viablity", but certainly not "cytotoxicity" ...
2) it is a "scientific non-sense" to pretend assessing anti-metastatic effects in vitro ...
It seems to me that my current comments come too late ...
But if any one would be interested in my comments, I would be more than happy to "scientifically" develop them.
Best regards
Robert
Dear Robert,
I am interested in in vitro assay of EMT as an early indication of metastasis and the inhibtion of the same. I would value your thoughts on the "scientific non-sense" to do in vitro studies on this topic.
Best regards,
Martin
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