I applied a MTT assay for A549 cell lines treated with different concentrations of Aluminium Oxide, I got the data from the plate reader, but I can't decide if my results are good or not ? And is there any specific methods or protocols that I can use to analyze my results
Hopefully this ideas are too basic! One way is to perform a standard curve using increasing cell numbers to see whether you are indeed measuring a linear increase in signal with increasing cell numbers. You could also have a negative control (such as hydrogen peroxide treatment) to simulate a "sick" cell. Is there a particular reason for choosing MTT (vs a soluble alternative like AlamarBlue, or CellTiter, PrestoBlue, etc which are not end-point assays and therefore allow you to use the same cells for something else)? Another potential advantage of these soluble redox assays is that and depending on the size of your Aluminum oxide, the protocol can be easily modified to ensure that the assay is not affected by the presence of the particles. Finally, incase you haven't already done this, you should also run "no cell controls" that have the same concentrations you are using in your cells to make sure that the aluminum oxide doesn't interfere (possibly by blocking absorbance). If I can help further you can message me. We have found, particularly when working with particles, that they interfere with the assays fairly frequently and controls (even if they seem very simple) are important to accurate interpretation of the data.
I could give you the protocol that used with different cell types,and it always workerd. The important thing is to set up controls in the std (wells with all the reagents, no cells). I'm sending you the protocol that I used.
I aaplied MTT assay for Molt-4 cell lines treated with different concentrations of EPA & DHA , I got the data from the plate reader, but I can't decide if my results are good or not ? And is there any specific methods or protocols that I can use to analyze my results so thank you
In mtt assay , you have to start the test by standard curve to set up the cell no. , in this assay OD between .4-.6 is ok in next step you have to consider some controls in your assay, 1. Media without cells as an indicator of ontamination and you have to get OD 0+-.1 otherwise you have contamination and cant trust on your data , you can also use a positive control in your assay to validate your data
You can also check the probable background of your reagent if it has interefernce at mtt wavelength by considering media +drug in your test in mtt if you consider all the control be sure the data is real data
Hope these basic comments help you and message me if you have furthure question
Actually I have a column with dead cells ( treated with Triton ), 1 column with only cells and media ( no Aluminium oxide added), and 2 rows are only cells+diff. concentrations of Aluminium oxide without MTT ! I also have 1 column with only media ( cell free ) ! I tried fitting the data to a linear curve but it looks really bad ! I guess I need to repeat the whole thing as my results do not really fit !
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