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How to determine minimal inhibitory concentration (MIC)


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#1 Foo. WX

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Posted 27 December 2009 - 08:48 AM

Hi Everyone,

I'm currently doing a research and I need to find minimal inhibitory concentration (MIC). My investigation concerns the inhibition of the growth of fungi. As I've not tried this before and I haven't seen anything on the web about this, I hope that you guys can help me out here. Thanks a lot! :lol:

#2 phage434

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Posted 27 December 2009 - 09:44 AM

Assuming you can culture your organism, this should be easy. Infect your standard culture medium with your organism at low concentration. Aliquot this infected medium into well of a microtiter plate. Take a small amount of culture medium, and add a known, relatively high inhibitory amount of antibiotic. Serially dilute this antibiotic containing medium with the infected medium, typically by 2x (equal volumes), mixing carefully at each stage. Grow the cultures, and note which well contains the first growth. The next well antibiotic concentration is the MIC. You may want to do this in triplicate.

#3 HomeBrew

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Posted 27 December 2009 - 04:35 PM

What are you trying to find the MIC of? Against what organism? There are standardized ways of doing this so that the numbers reported by any lab are comparable -- the MIC of a particular substance towards a particular organism under one growth condition or on a particular media may be different than that found if the conditions are different.

See a reference like this one, or consult other guidelines for the accepted way of doing this if you're going to publish the results.

#4 Mazhar Hussain

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Posted 27 December 2009 - 11:30 PM

Hi Everyone,

I'm currently doing a research and I need to find minimal inhibitory concentration (MIC). My investigation concerns the inhibition of the growth of fungi. As I've not tried this before and I haven't seen anything on the web about this, I hope that you guys can help me out here. Thanks a lot! :rolleyes:

Dear Foo. WX,
As u knw MIC, is that lowest antibiotic concentration on which Bacteria in question will stop growing. You will prepare a number of test tubes with about 5 ml of media in each (i.e. 10 tubes). After that, add antibiotic in each test tube in following pattern:
For Example
50 micgrm/ml
25 micrgrm/ml
12.5 micrgrm/ml
.
.
.
.
carry on upto last tube
Now, give 20ul of inoculum in each or any other amount... But keep in mind only Antibiotic is variable in this experiment... and every other thing is constant...
Then Incubate them for 18-24hrs at 35 C.
Then check for growth. Start from higher concentration observing...
U will see no growth on this extreme... Carry on towards less concentration... U will c a test tube with minute growth... The test tube before this can be expected to be MIC carrying concentration...
But we can nt trust it because, it may contain viable microbe with static effects... So take at least 3 test tubes which r present before test tube carrying minute growth... and spread them on plate containing media carrying no stress.
Incubate them for 24 hours
then check for plate carrying no growth...

MIC will b that tube concentration which carry less amount of antibiotic with no growth in all 3 plates...

"I knw difficult to understand," but dear i tried my best...
Mazhar Hussain
Microbiology & Molecular Genetics
Website: Microbiology On-Line
FaceBook Group: Microbiology On-Line

#5 Foo. WX

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Posted 03 January 2010 - 07:07 AM

Sorry guys, for taking such a long time to reply. Thanks for the prompt reply too, really appreciate it :P Right now, I just have certain things to clarify, hope you guys can continue helping me out. Thank you =)

Oh and a question is, which way would it be the best way to carry out the experiment? As in, by using agar plates or broths?

Assuming you can culture your organism, this should be easy. Infect your standard culture medium with your organism at low concentration. Aliquot this infected medium into well of a microtiter plate. Take a small amount of culture medium, and add a known, relatively high inhibitory amount of antibiotic. Serially dilute this antibiotic containing medium with the infected medium, typically by 2x (equal volumes), mixing carefully at each stage. Grow the cultures, and note which well contains the first growth. The next well antibiotic concentration is the MIC. You may want to do this in triplicate.


Can I ask, what do you mean by serially diluting it by 2x?

What are you trying to find the MIC of? Against what organism? There are standardized ways of doing this so that the numbers reported by any lab are comparable -- the MIC of a particular substance towards a particular organism under one growth condition or on a particular media may be different than that found if the conditions are different.

See a reference like this one, or consult other guidelines for the accepted way of doing this if you're going to publish the results.


I'm finding the MIC against Rhizopus stolonifer, a kind of fungi.

Thanks for the PDF file. I've actually read through it, but I'm unsure of the concentration that I should start off with since I'm not exactly doing any organisms related to those stated there and I'm not using antibiotics. Maybe you can help me out with that? Let's say I'm using food preservative as one of my inhibiting substance.

Dear Foo. WX,
As u knw MIC, is that lowest antibiotic concentration on which Bacteria in question will stop growing. You will prepare a number of test tubes with about 5 ml of media in each (i.e. 10 tubes). After that, add antibiotic in each test tube in following pattern:
For Example
50 micgrm/ml
25 micrgrm/ml
12.5 micrgrm/ml
.
.
.
.
carry on upto last tube
Now, give 20ul of inoculum in each or any other amount... But keep in mind only Antibiotic is variable in this experiment... and every other thing is constant...
Then Incubate them for 18-24hrs at 35 C.
Then check for growth. Start from higher concentration observing...
U will see no growth on this extreme... Carry on towards less concentration... U will c a test tube with minute growth... The test tube before this can be expected to be MIC carrying concentration...
But we can nt trust it because, it may contain viable microbe with static effects... So take at least 3 test tubes which r present before test tube carrying minute growth... and spread them on plate containing media carrying no stress.
Incubate them for 24 hours
then check for plate carrying no growth...

MIC will b that tube concentration which carry less amount of antibiotic with no growth in all 3 plates...

"I knw difficult to understand," but dear i tried my best...


Regarding the concentration that you gave, what happens if the MIC is in between concentrations like... 25 and 12.5?

Haha don't worry, I think I'm getting it.



Thank you everyone :P

#6 Mazhar Hussain

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Posted 08 January 2010 - 06:01 AM

I recommend broth method....

It is better to consider, that much concentration MIC...

But if u have extra resources to pour on ur experiment...

Then do same thing with range between 12.5 to 25 as we did with 50 - 1

In clinics we do use amount which come in first cycle... But if u want u can... No one can stop u... :) :angry:
Mazhar Hussain
Microbiology & Molecular Genetics
Website: Microbiology On-Line
FaceBook Group: Microbiology On-Line




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