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  • Fluorometric detection of active alkaline phosphatase and gamma-glutamyl transferase in fluid dairy products from multiple species.

Fluorometric detection of active alkaline phosphatase and gamma-glutamyl transferase in fluid dairy products from multiple species.

Journal of food protection (2013-05-07)
George C Ziobro, Kevin M McElroy
ABSTRACT

Over the past 80 years, a variety of methods have been developed to detect underpasteurized or improperly pasteurized milks used in dairy products. Existing methods are hampered by duration of analysis, poor reproducibility, and in some cases the use of hazardous chemicals. To overcome these issues, two new methods have been developed using fluorogenic substrates for two marker enzymes, alkaline phosphatase and gamma-glutamyl transferase. In 30 min, up to 18 samples can be analyzed in triplicate by both methods on two separate 96-well plates. Sample preparation is not necessary for liquid milks when using these methods. The relative standard deviation for each assay is less than 9%, and the correlation coefficient for results of the two methods is greater than 0.98. Using the new methods, milks from four species and nine commercially available liquid milk products were tested. The new methods were also tested directly against an existing phosphatase method (Fluorophos) in spiked whole milk samples.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
γ-Glutamyltranspeptidase from equine kidney, Type VI, 5-12 units/mg solid
Sigma-Aldrich
γ-Glutamyltranspeptidase from porcine kidney, Type IV, 2.0-6.0 units/mg solid
Sigma-Aldrich
Phosphatase, Alkaline bovine, recombinant, expressed in Pichia pastoris, ≥4000 units/mg protein
Sigma-Aldrich
Phosphatase, Alkaline from Escherichia coli, buffered aqueous glycerol solution, 20-50 units/mg protein (in glycine buffer)
Sigma-Aldrich
Phosphatase, Alkaline from Escherichia coli, ammonium sulfate suspension, 30-90 units/mg protein (modified Warburg-Christian, in glycine buffer)
Sigma-Aldrich
Phosphatase, Alkaline from Escherichia coli, lyophilized powder, 30-60 units/mg protein (in glycine buffer)
Sigma-Aldrich
Phosphatase, Alkaline from porcine kidney, lyophilized powder, ≥100 DEA units/mg protein
Sigma-Aldrich
Phosphatase, Alkaline from bovine intestinal mucosa, buffered aqueous solution, ≥2,000 DEA units/mg protein
Sigma-Aldrich
Phosphatase, Alkaline from bovine intestinal mucosa, lyophilized powder, ≥10 DEA units/mg solid
Sigma-Aldrich
Phosphatase, Alkaline from bovine intestinal mucosa, buffered aqueous glycerol solution, ≥4,000 DEA units/mg protein
Sigma-Aldrich
Phosphatase, Alkaline from bovine intestinal mucosa, ≥5,500 DEA units/mg protein
Sigma-Aldrich
Phosphatase, Alkaline shrimp, ≥900 DEA units/mL, buffered aqueous glycerol solution, recombinant, expressed in proprietary host
Sigma-Aldrich
Phosphatase, Alkaline from bovine intestinal mucosa, BioUltra, ≥5,700 DEA units/mg protein