Reference materials for water microbiology were prepared by spray-drying milk, artificially contaminated with a known test strain. The resulting highly contaminated milk powder was mixed with sterile milk powder to a contamination level of 2000-3000 cfu/g. Gelatin capsules were filled with the mixture (0.2 g/capsule) to produce the reference materials. Test strains used were: WR1 Escherichia coli, WR3 Enterobacter cloacae, WR63 Enterococcus faecium and WR51 Staphylococcus spec. Optimalization of the mixing procedure and aging of the highly contaminated milk powder resulted in relatively homogeneous and stable reference materials. Short-time challenge tests at “high” temperatures (30 and 37 °C) may be predictive for long-term stability at “low” temperatures (4-6 °C). Reference materials with strain WR3 were stable for one week at 30 °C and for at least 6 months at 4-6 °C.

Possible applications of the reference materials are: quality control of routine measurements, comparison of the efficiency of different culturing methods and as a standardized sample in collaborative studies.

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