Antimicrobial Synergy Testing

Checkerboard Assay Services

As an expert in providing solutions to assist virology and microbiology research, Creative Diagnostics has recently announced the launch of its new Checkerboard Assay services to support researchers in evaluating the interaction of two antimicrobial test compounds, including Standard Bacteria Culturing, Antimicrobial Synergy Testing, and Statistics Analysis of experimental results.

The checkerboard test is an experimental method used to evaluate the interaction of two antimicrobial test compounds. In this test, the MIC and MBC values of the test compounds are used individually and combined with the MIC and MBC values of each bacterial strain being evaluated to calculate accumulation.

Synergy measurements by checkerboard analysis can be utilized to determine changes in the antibacterial potency of an antibiotic combination relative to its activity. Comparative results are then calculated using the Fractional Inhibitory Concentration (FIC) index value, which shows the greatest change in antibiotic combinations compared to the MIC of a single antibiotic.

Creative Diagnostics now offers a series of Checkerboard Assay services to the research communities, including Standard Bacteria Culturing, Antimicrobial Synergy Testing, and Statistics Analysis of experimental results. Creative Diagnostics works with scientists in industry and academia to support the early stages of antimicrobial, vaccine and diagnostic discovery and to advance research in bacterial infections.

Before testing, a stock solution and two serial dilutions of at least twice the MIC should be prepared for each drug. A total of 50 μL of Müller-Hinton broth is then injected into each well of the microdilution plate. Next, the first antibiotic in the combination is sequentially diluted along the ordinate and the second drug is diluted along the abscissa. An inoculum equivalent to 0.5 McFarland turbidity standard is then prepared from each isolated Pseudomonas aeruginosa and placed in Müller-Hinton broth.

Finally, 100 μl of 5 x 105 CFU/ml bacterial inoculum is inoculated into each microwell and incubated for 48 hours at 35°C under aerobic conditions. The resulting checkerboard grid contains each combination of the two antibiotics, with the diagonal representing the tube with the highest concentration of each antibiotic. To quantify the interaction between the tested antibiotics (the FIC index), researchers can calculate using the following formula: A /MIC A + B/MIC B = FIC A + FIC B= FIC Index. When the FIC value is <0.5, the two antibiotics are synergistic; when the FIC is >4, the two antibiotics are antagonistic; and when the FIC is 0.5-4, the two are either additive or non-additive.

Creative Diagnostics offers assays for the major bacterial families and can help customers select the appropriate assay based on the compound and mechanism of action. The company can also develop custom assays based on customers’ specific processes. For more information, please visit

About Creative Diagnostics

Headquartered in New York, Creative Diagnostics is a consulting and experimental service provider specializing in virology and microbiology. The company provides comprehensive solutions to conquer obstacles in virology and microbiology research, from high-security infrastructure provision, and biosafety regulation elucidation, to expert viral system assistance.

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