The literature states that the MTS assay should be performed with a final well volume of 100uL/well with an addition of 20uL MTS reagent/well.
Does anyone think there would be a concern if using a final volume of 75uL/well with an addition of 15uL/well MTS?
I am getting sporadic results across the plate when attempting this lower volume. I will go back to 100uL/well final volume. However, I am trying to understand if the problem was due to the decreased volume in the well or another factor.
Anyone had success with lower volumes?
Thanks
I find various MTS assay questions at RG that may be helpful:
https://www.researchgate.net/topic/MTS-assay
Hello Luke Williams
You may add lower volume of MTS solution per well . But the sporadic results across the plate may be due to other factors. You can refer to the research article below for some useful information on MTS troubleshooting.
Article Inaccuracies in MTS assays: Major distorting effects of medi...
Hope this helps.
Best.
It depends on the microplate you are using for your readings. It is a matter of having a uniform solution column in each well. Note that unlike cuvettes, which have fixed pre-defined pathlength, the pathlength in microplates depends on the volume of solution in each well. Given the Lambert-Beer law, changes in pathlength lead to changes in absorbance all else being equal. Moreover, even two wells with the same volume, but different solvents/viscosity (e.g., water versus 50% ethanol), might result in different readings.
From my experience, volumes below 100 µL usually result in erratic readings for standard 96-well plates. However, lower volumes might work well for 384-well plates or half-area 96-well plates.
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