Difference between revisions of "Plate bioassay for antimicrobial activity"
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== Protocol == | == Protocol == | ||
| + | -Inoculate culture to be tested on NBY agar plates and incubate overnight at 30 C | ||
| + | |||
| + | -Use a sterile toothpick to stab inoculate another NBY agar plate with a single colony from the culture | ||
| + | |||
| + | -Incubate at 30 C overnight | ||
| + | |||
| + | -Incubate the plates at 65 C for one hour to kill the bacteria, leaving just the antimicrobial compound it produced behind | ||
| + | |||
| + | -Mix a colony of ''Geotrichum candidum'' into 750 microliters of water | ||
| + | |||
| + | -Mix the water-bacteria mixture with 25-30 mL of molten NBY Top Agar. Make sure the molten agar is not too hot for the fungus, or too cold that it solidifies early | ||
| + | |||
| + | -Pour the top agar solution onto the NBY plates that were stab inoculated, insuring the entire plate is covered with agar | ||
| + | |||
| + | -Let the top agar harden and incubate at 30 C overnight | ||
| + | |||
| + | -Measure the diameter of the clearing of the bacterial lawn around each stab, this is called the zone of inhibition | ||
== Notes == | == Notes == | ||
Revision as of 12:38, 14 July 2015
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Contents
Purpose
This assay can be used to test for production of or sensitivity to antimicrobial agents by measuring a zone of inhibition around an antimicrobial compound.
Materials Needed
(For testing four plates)
8 NBY Agar plates
25-30 mL NBY Top Agar
750 mL water
Sterile toothpicks
Millimeter measuring stick
Indicator Fungus-Geotrichum candidum
30 C incubator
65 C incubator
Protocol
-Inoculate culture to be tested on NBY agar plates and incubate overnight at 30 C
-Use a sterile toothpick to stab inoculate another NBY agar plate with a single colony from the culture
-Incubate at 30 C overnight
-Incubate the plates at 65 C for one hour to kill the bacteria, leaving just the antimicrobial compound it produced behind
-Mix a colony of Geotrichum candidum into 750 microliters of water
-Mix the water-bacteria mixture with 25-30 mL of molten NBY Top Agar. Make sure the molten agar is not too hot for the fungus, or too cold that it solidifies early
-Pour the top agar solution onto the NBY plates that were stab inoculated, insuring the entire plate is covered with agar
-Let the top agar harden and incubate at 30 C overnight
-Measure the diameter of the clearing of the bacterial lawn around each stab, this is called the zone of inhibition
Notes
Papers where this or a similar method has been used
Related Ontology Terms
References
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