I am using Broth micro dilution method on 96 well plates for the determination of MIC values of some of the natural compounds I have isolated. I am using varying concentrations of the compound against the bacteria and have calculated the percent inhibition values against every concentration I used, by taking their ODs. I am using MTT dye as a marker of cellular viability. Here I would like to know how can I determine the MIC value. Should I look for the concentration at which there is no visible growth, or should I look for that particular concentration of my compound at which there is a specific percentage of bacterial inhibition (say, 98% etc.).
Need expert opinion on this.
MIC (minimal inhibitory concentration) does not mean it should be calculated somehow specifically. It just means the minimal concentration of compound at which you can not see the bacterial growth. However, it is always more accurate if you may determine MIC by using a quantitative method. Scientists consider different inhibition % as MICs, usually 90% or 95%, sometimes 99%, but it should be totally fine to use any of those values (let's say 95%) if you provide your explanation of MIC calculation in the description of methods.
The determination of MIC based on the growth inhibition determined by reading the percentage reduction in bacterial density appears more accurate. Most researchers assume a 99% reduction as MIC values, while some consider the value of 95%.
MIC (minimal inhibitory concentration) does not mean it should be calculated somehow specifically. It just means the minimal concentration of compound at which you can not see the bacterial growth. However, it is always more accurate if you may determine MIC by using a quantitative method. Scientists consider different inhibition % as MICs, usually 90% or 95%, sometimes 99%, but it should be totally fine to use any of those values (let's say 95%) if you provide your explanation of MIC calculation in the description of methods.
You should use turbidity measurements, and addition of viability stains are not necessary. You could alternatively use Etest, which is rapid and convenient, and uses less lab time. Also, to comment on MIC 50, MIC 90 etc, these do not relate to individual strains (they do not mean that 50% or 90% of the bacteria in your single strain test are inhibited) but they are population studies. In other words, MIC50 mean that the MIC of an antibiotic that inhibits 50% of a panel of isolates is Xmg/L.
Hello Muhammad Amir Khan,
Adding to the answers stated above, you can actually try to inoculate and colony count the 96 wells plate of those concentration marked (by using MTT or TCC) as MBC, MIC and negative control. For such, you can actually compare where:
MBC-absent of bacerial growth
Negative Control -Maximal bacterial growth
MIC-Inhibited bacterial growth.
The colony count (CFU/ml) should be as MBC
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