Hello,
I would like to ask.
I performed broth microdilution assay against E. coli and S. aureus, my extract were dissolved with 8% DMSO in dH20. The MIC for my sample was 10 mg/ml and 5 mg/ml, respectively. I use the MIC for my disk diffusion to confirm the antimicrobial activity. And the sample preparation were identical from the broth microdilution. However, after incubation, I didn't observe any inhibition zone in both E. coli and S. aureus.
Did I missed something, do the MIC from broth microdilution too low for disk diffusion assay? Do I need to replace dH20 with methanol to enhance diffusion?
d.w. is inert don't have any effect so you must do control with DMSO
Thank you for your reply Phil Geis and Anfal Izaldeen al Kateeb , I did use 8% DMSO in dH20 as control, as it was the concentration used to dissolved my extract. It used to ensure at that concentration does not have any inhibition effect on the bacteria.
I wanted to double confirm the antibacterial activity. Since I got the MIC value, I want to test the concentration in disk diffusion assay.
Phil Geis Thank you for your reply. Yes, I dissolved the extract with 8% DMSO in dH20 and transferred it to blank disk (20 ul per disk). These disk were used for disk diffusion assay. Is it possibility for it to bind to the disk? and causes no inhibition observed after incubation.
The mic for liquid broth is never the same for solid agar. (See, clinically an e test will never be the same as microdilution mic for the same sample). So you may just not be using the correct concentrations for your diffusion assay. Also double check the material your filters are made of and make sure that organic solvents will diffuse from them
No, generally, the Disk Diffusion Assay is a qualitative test which will allow you to have an idea on the presence or not of antimicrobial activity of your extract.
Broth Microdilution Assay is then used to determine the MIC.
An e test and disc diffusion rely on the same principles: solid agar diffusion. And they often result in similar mics ( though not always). E tests are just more quantitative than disc diffusion assays.. The specifics are a little different but that is moot. Regardless, my point that a microbroth mic rarely matches disc diffusion/e test still stands.