Study 5 Serial Dilution method flashcards from Mandy S. On StudyBlue. Advantages of serial dilution-agar plate; 1. Only viable cells counted. Disadvantages of. The dilutions cover the range from 1/2 to 1/100 unevenly. In fact, the 1/2 vs. 1/3 dilutions differ by only 1.5-fold in concentration, while the 1/10 vs. 1/100 dilutions differ by ten-fold. If you are going to measure results for four dilutions, it is a waste of time and materials to make two of them almost the same.
The antibiotic to be tested is diluted with water to produce a series of concentrations. An appropriate volume is then combined with melted agar to produce plates in which the final antibiotic concentrations represent a 2-fold dilution series. After this, bacteria prepared to a standard concentration are added as a spot to each plate, with 10^4 colony forming units (CFU) per spot. This technique allows for replicate spots of one bacterial type to be tested or spots of different bacteria so that the MIC of the antibiotic against multiple types of bacteria can be tested. Necessary controls include a control plate that does not receive any antibiotics and bacterial spread plates demonstrating that the bacterial inocula are in the correct concentration range.
The dilution plates are then incubated at a temperature of 37 degrees. The plates are then incubated for sixteen to eighteen hours, although incubation time may be less for bacteria populations that divide quickly.: 374 After incubation, the plates are examined to determine if bacterial growth has occurred in the inoculated spots. The lowest concentration of antibiotics that prevented bacterial growth is considered to be the minimum inhibitory concentration of that antibiotic against that bacterium. ^ Lorian, Victor (2005).
Lippincott Williams & Wilkins. Wiegand, Irith; Hilpert, Kai; Hancock, Robert E. (17 January 2008). 'Agar and broth dilution methods to determine the minimal inhibitory concentration (MIC) of antimicrobial substances'. Nature Protocols.
European Committee for Antimicrobial Susceptibility Testing of the European Society of Clinical Microbiology and Infectious Disease (September 2000). 'Determination of minimum inhibitory concentrations (MICs) of antibacterial agents by agar dilution'. Clinical Microbiology and Infection. 6 (9): 509–515. ^ Parija (2009). Elsevier India.
Lee, Mary (2013). Retrieved 16 November 2014.
Demonstration video showing how to perform a serial dilution on a liquid food sample (in this case raw unpasteurised milk). The raw milk sample is diluted down to 1/1000 in a sterile diluent (Ringers solution). 1ml samples of each dilution are then used to prepare 2 sets of pour plates. A 0.1ml sample from the 1/10 and the 1/100 dilutions is used to generate spread plates.
For this method you require the following: Sterile 1ml pipettes A pipette pump Sterile agar plates (for pour plates) Poured agar plates (for spread plates) A spreader (glass or metal) A dish of lab ethanol Bunsen burner Glass Universal tubes with 9 ml of sterile diluent Molten agar medium (20ml plate count agar per Universal tube - kept at 48C until ready to pour).