SCREENING METHODS: Broth dilution methods(2)

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Broth dilution methods(2)

For organisms sensitive to oxygen tension in the media this can present an additional problem as the oil can inhibit gaseous exchange. Tween or ethanol may be used to enhance incorporation into the aqueous media, however as previously discussed these compounds may interfere with the assay results. The broth assay was used in both the macro-and microdilution versions. The microtube version allows an increase in productivity through the use of microtiter plates. Critical control points or standardization of the assay may be achieved at the following stages, medium type, pH, stock solutions, concentration range, inoculums density, and incubation conditions.

Micro broth methods have also been developed, which utilize microtiter plates, thus reducing the volume of extract needed, and have endpoints that can be determined spectrophotmetrically, either a measure of turbidity or use of a cell viability indicator (e.g. resazurin, methylthiazoldiphenyltetrazolium (MTT)) (Mann and Markham, 1998; Panda et al., 2010c, 2011a) (plate-4). They also propose that the cell viability indicator is the best method of endpoint determination for experiments with essential oils interface with turbidity measures.
plate4Screening Methods in the Study-7
Plate 4 : Photograph of broth dilution method i. model of 96 well microtiter plate;
ii. MIC result of plant extract by microdilution method using TTC (A-G: decreasing concentration of extract, H-control)

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