New Bioluminescent Alkaline Phosphatase Test for Verification of Milk Pasteurization

Introduction: This study introduces a new technology from Hygiena for the monitoring of Alkaline Phosphatase verification from pasteurised milk samples. The technology uses new bioluminescent substrates in a simple self-contained device using a new luminometer to measure and interpret results from samples

Purpose: To demonstrate the performance of ZymoSnap ALP as a rapid and efficient method for pasteurization verification.  

Methods: Pasteurized milk from 10 supermarkets across USA covering all fat contents from 4% down to 0.1% were evaluated in 2 assays for various Hygiena luminometers. Both assays produce equivalent results to the gold standard Fluorophos system. Each milk sample was spiked with 4 levels of bovine alkaline phosphatase to the following levels 0 mU/L, 100mU/L 350mU/L and 1000mU/L. Each milk was assayed using 5 replicates with means compared and charted for correlation.

Results: The assay demonstrated excellent linearity at all milk fat contents and demonstrated an inversely proportional relationship between fat content and light output. At 4%, 2%, 1% and 0.1% the conversion of mU/L alkaline phosphatase to RLU (relative light unit) was as follows – 4% RLU= 3xmU/L, at 2% RLU=1.75mU/L, at 1% RLU=1.5mU/L and at 0.1% RLU=1.0mU/L. The correlation coefficients for the 10 milk types at n=5 was as follows 4% R²=09561, at 2% R²=09561, at 1% R²=09561 and at 0.1% R²=09561. Other flavoured milks were also tested and followed a similar trend with fat content of 4% having the largest influence on signal output and not color of milk. Strawberry milk had a conversion to RLU of 1.2 and chocolate milk a conversion of 1.1 RLU per mU/L.

Significance: The easy-to-use assay gives dairy processors of all sizes the ability to run a quick and inexpensive alkaline phosphatase assay on milks of any fat content or flavorsfor rapid pasteurisation verification.