Silver nanoparticles may impede the MTT assay output by either adsorbing the tetrazolium salt or by modifying the catalytic activity of mitochondrial reductases in cells showed that AgNPs interacted with the MTT reagent, generating incorrect optical density values.
Waht the alternative approaches with better protocol steps?
Is MTT assay really the best choice for determine the effect of silver nanoparticles on the cell number ?
Hello, One can make a series of dilutions and then plate it on agar plates( for bacterial cells), incubate at 37C for 24-48h and count bacteria cells(CFU). This is the most reliable assay.
H Ibrahim Alghoraibi I have probably misunderstood your question.
Please correct me if I am wrong, but Ag nanoparticles do not have any effect on mitochondria in the presence of free sulfhudryls.
1)http://www.nanospainconf.org/2014/Abstracts/Nanospain2014_Pugliara_Alessandro_NanoSpain2014_Pugliara_Alessandro.pdf
2) https://www.semanticscholar.org/paper/Biotic-and-abiotic-interactions-in-aquatic-fate-and-Unrine-Colman/84ed22d001352531dfed676dc3cf839bd65493d2
I think the main probleme in MTT Process is that it occurs an interferance in spectrophotometric measurement between the absorbance of silver nanoparticles and the absorbance of formazan crystalline at 570nm. where the amount of absorbance correlates with viable cell number. As a result, there is an error in the estimating the number of viable cells
MTT assay is not very reliable for too low concentrations of cells. There should not be a problem with overlapping of absorbance with plasmon peak of AgNPs which is usually at 400nm. To overcome possible effect of overlapping you need to run exactly the same volume and concentration of AgNPs as a blank and then Absorbance (MTT+AgNPs) -Abs(AgNPs)
One can make a series of dilutions and then plate it on agar plates(for bacteria ) incubate at 37C for 24-48h and count bacteria cells(CFU). This method is more reliable. otherwise for mammalian cells use staining and then Flow cytometry
Dear Dmitriy Berillo thanks for your answer
In fact, we found by experiment that there is overlapping of absorbance MTT and AgNPs, So We look for another process such as Flow cytometry or alamar blue ............
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