Chapter 5.21 : Preparation of Broth Microdilution MIC Trays

Abstract:

Broth microdilution MIC trays may be prepared in-house for performance of quantitative antimicrobial susceptibility studies. If large numbers of trays are required, dilutions of the antimicrobial agents are made in large volumes and generally dispensed in 0.1-ml amounts into microdilution trays by using a commercial dispensing apparatus. For individual test trays, dilutions of the required antimicrobial agents may be accomplished with an eight-channel micropipette. The procedure described here is based on 96-well trays containing 0.1 ml per well. Not every aspect of broth microdilution tray preparation has been standardized, and for some steps, there are multiple ways of accomplishing identical results. The method presented here conforms closely to that described by the CLSI ( 1 , 2 ) and is the procedure that has been successfully used with a Quick Spense (Sandy Springs Instrument Co.) and AutoSpense IIe (Halpro, Inc.) dispensers at the clinical microbiology laboratories of the University of California, Los Angeles for nearly three decades. It assumes that the layout of the tray in terms of positioning respective concentrations of each antimicrobial agent has been previously determined. The method is detailed for preparation of approximately 1,800 trays, but recommendations for preparing fewer trays are included.