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A study was conducted to design a more sensitive and flexible technique for the assay of neomycin in milk. Sterile Antibiotic Assay medium (Difco; 15 ml) was poured into glass petri dishes with aluminum tops equipped with absorbent discs. Seed agar (4 ml; containing 1% sodium chloride) inoculated with standardized amounts of the test organism (Staphylococcus epidermidis ATCC 12228) was laid over the hardened base agar, and stainless-steel cylinders were placed equal distances apart in the agar. The plates were refrigerated for 30 min and the cups were removed in an atmosphere of minimal contamination. The resultant holes were sealed with melted agar. After preparation, the holes were used for assay procedures. Samples to be assayed were pipetted into the holes and the plates were refrigerated for 90 min at 4 C. Plates were incubated for 18 to 20 hr at 32 C. The zones of inhibition were recorded and compared with a standard curve. The approximate sensitivity of this method was 0.05 μg/ml.

Increase in Sensitivity for the Assay of Neomycin in Milk