Method for AmpC
Black JA, Moland ES, Thomson KS. J Clin Microbiol. 2005 Jul;43(7):3110-3. AmpC
disk test for detection of plasmid-mediated AmpC beta-lactamases in Enterobacteriaceae
lacking chromosomal AmpC beta-lactamases
The steps are:
1. Moisten Tris EDTA(TE) (catalog # 232211, BD Diagnostic Systems, Sparks,
MD) disk with 20 μl of sterile saline.
2. The TE disk should be inoculated with a heavy inoculum of the test organism
prior to placing it on the plate. We do this in an empty Petri dish and use the
wooden end of a sterile cotton swab to apply the organism. It should then be
inverted before placing it on the agar. This ensures optimal diffusion of the βlactamase into the agar.
3. Inoculate susceptibility plate with lawn of 0.5 McFarland suspension of E.coli
ATCC 25922
4. Place cefoxitin disk on the susceptibility plate.
5. Place the TE disk with test organism close to the cefoxitin disk but not touching
(1 mm separation is good).
6. Interpret test after overnight incubation. Examine zone margin for indentation or
flattening - evidence of a possible AmpC β-lactamase.
Control Strains
Strong positive control Enterobacter cloacae ATCC BAA-1143, weak positive
control Klebsiella pneumoniae ATCC BAA-1144, and negative control Escherichia
coli ATCC 25922.