To begin, prepare two milligrams per milliliter of zinc oxide nanoparticle solution using DPBS. Perform twofold serial dilution to make different concentrations. Add 100 microliters of each tested zinc oxide nanoparticles concentration into a 96-well plate.
Dilute the bacterial culture to one million CFU per milliliter with tryptic soy broth or TSB media. Add 100 microliters to each well containing different concentrations of zinc oxide nanoparticle solution. Incubate the plate at 37 degrees Celsius for 24 hours.
Pipette 100 microliters of zinc oxide nanoparticles with bacterial solution from each well and prepare various tenfold cereal dilutions until 10 to the negative six. Transfer 50 microliters from four dilutions to tryptic soy agar TSA media plates. Incubate the agar plates at 37 degrees Celsius for 24 hours.
After selecting a dilution factor that is countable for each group, mark all the colonies in the countable dilution plate and recalculate so that the concentration becomes number of CFU per milliliter. Use the obtained data to represent the percentage of live bacteria relative to those in the negative control. The antibacterial properties of zinc oxide nanoparticles were tested on 0.5 million CFU per milliliter Pseudomonas aeruginosa strain.
The antimicrobial activity of the nanoparticles increased in a concentration-dependent manner. However, a visible decrease in the number of bacterial colonies from the starting undiluted bacterial culture was not evident. When tested against a 0.5 million CFU per milliliter MRSA strain, these nanoparticles exhibited increased antibacterial activity, leading to a significant reduction in bacterial colony formation.
