Improved Sensitivity of the Bioluminescent Determination of Numbers of Bacteria in Milk Samples.

A procedure was developed for the sensitive measurement of bacterial ATP in raw milk samples using the firefly luciferase assay. A sensitivity of 10 bacteria/ml was achieved in the laboratory. Treatment of milk samples with a detergent (Lubrol PX) and a proteolytic enzyme (trypsin) allowed filtration of 50 ml of milk through a 0.4-μm Nucleopore filter; a 50-ml sample of milk was required to provide sufficient ATP for measurement. The bacterial ATP was released from the bacteria on the filter using the commercial product Nucleotide Releasing Reagent for Bacteria (NRB from Lumac). The procedure allows determination of 10 bacteria/ml of raw milk in 20 min.