This study investigates the antibacterial efficacy of antibiotic-loaded polymeric strips using a bacterial culture model. The methodology involves measuring luminescence to assess bacterial growth inhibition.
Take syringes containing antibiotic-free and antibiotic-loaded polymeric strips.
Fill the syringes with a metabolically active culture of pathogenic bacteria.
Incubate the syringes with shaking to ensure uniform bacterial exposure to the medium.
During incubation, antibiotics from the loaded strip diffuse into the media and inhibit bacterial growth.
In the syringe with the antibiotic-free strip, the bacteria proliferate.
Collect the bacterial suspension from each syringe into microcentrifuge tubes.
Transfer the aliquots into a multiwell plate.
Add a reagent mixture containing luciferin, luciferase enzyme, and a lysis agent.
Incubate with shaking to enable bacterial lysis, which releases cellular components, including ATP. ATP drives the luciferase-mediated luminescent reaction.
Using a microplate reader, record the luminescence.
Bacterial culture exposed to the antibiotic-loaded strip exhibits reduced luminescence compared to the culture unexposed to antibiotics.
This reduced luminescence confirms the antibacterial efficacy of the antibiotic-loaded strip.
Place the virgin and drug loaded UHMWPE strips within a three-milliliter syringe. Then draw MHB-containing bacterial suspension into the syringe through the attached needle up to the 1.5 milliliter mark.
Place the syringe setup on a shaking incubator until the indicated time points of six hours, and then every day until day seven. After each indicated time point, take out the syringe setup and dispense the media in a two-milliliter tube. Perform a real-time microbial viability assay as described in the manuscript using 100 microliters of bacterial suspension.
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