I prefer well diffusion as some natural products will adhere to the discs preferentially. Once you've narrowed your scope, I would recommend doing growth curves as well.

Additionally, be careful to control for your solvent. Some bacteria are fastidious and can survive quite high concentrations of solvents like methanol and DMSO, but some are intolerant to even small changes in their salt concentration. It's always a battle to find a way to get your natural products into solution and avoid adverse effects on your bacteria. Good luck!

I prefer well diffusion as some natural products will adhere to the discs preferentially. Once you've narrowed your scope, I would recommend doing growth curves as well.

Additionally, be careful to control for your solvent. Some bacteria are fastidious and can survive quite high concentrations of solvents like methanol and DMSO, but some are intolerant to even small changes in their salt concentration. It's always a battle to find a way to get your natural products into solution and avoid adverse effects on your bacteria. Good luck!

If you have access to a multi-well plate reader, I would suggest using a high-throughput assay in liquid media. For example, if you have clear 96-well plates, then you can pipet fixed bacteria number into each well and test against more samples and controls than would be feasibly possible between your other alternatives. Furthermore, in addition to providing raw anti-bacterial data, you can calculate IC50 and kinetic more readily.

I agree with Charles Jones.

Microtitre broth dilution method is the best way to identify the antibacterial natural product with lowest concentrations.

Follow CLSI guidelines for the specific pathogen you are going to do. You can get the valid information from there.

thanks to all. I dont have acces to a multi-well plate reader, so i will work on agar.

One method I'm working with is similar to what Dr. Mann suggests - apply your extract in spots on an agar plate and allow time for the solvent to evaporate (I use a laminar hood with the plate open/cracked). Applying a lawn of bacteria or spores (I work with fungi) is then easily done via TLC sprayer, allowing you to get a fine coverage and look for clearing zones or weakened growth around your extracts.

The best method is the agar well diffusion method.ht gives you the oppurtunity to mearure the zones of inhibition on t he plates and the clearity of the extract diffusing out of the plate

Sebastian, do take care of the solvent in which you have extracted your material. A non-polar solvent extract like ethyl acetate or hexane would create diffusion problems in water laden agar plates. For such extracts, disc diffusion could be prefered.

Microbroth dilution is an easy method and ensure good mixing of the natural product and the test organism. With no reader available, you can add a solution of tetrazolium chloride which is a redox indicator and a change in color will indicate growth. No change in color indicates inhibition

I would like to suggest several publications: A new effective assay to detect antimicrobial activity of filamentous fungi (Pereiraa et al., Microbiological Research 168 (2013) 1– 5); Optimal methods for evaluating antimicrobial activities from plant extracts (Othman et al. J Microbiol Meth 2011;84:161–6).

Well, there are number of good, better and best methods and people ahve already suggested lots. I would like to add most simple and time saving method which every microbiologist usually do but not the best one. Just grow your bacterial culture in presence and absence of your interest of compound/extract and just take OD after 20 or 40 minutes. It should tell you whether you should go ahead for better assays.