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Validation of spectrophotometric assays for serum paraoxonase type-1 measurement in dogs.

OBJECTIVE
To evaluate and validate 3 spectrophotometric assays for measuring serum activity of paraoxonase type-1 (PON1), an enzyme associated with high-density lipoproteins, in dogs.


ANIMALS
22 healthy adult dogs and 10 dogs with eccentrocytosis.


PROCEDURES
2 methods were adapted for use in 96-well microplates with phenyl acetate and 5-thiobutyl butyrolactonase as substrates, and 1 was adapted for use in an automated analyzer with p-nitrophenyl acetate as substrate. Blood samples were collected from all dogs, serum was harvested, and serum PON1 activity was measured with each method.


RESULTS
Imprecision was low for all 3 methods, with the exception of interassay imprecision for 5-thiobutyl butyrolactonase, and results were linear across serial sample dilutions. The 3 methods were able to detect low PON1 activity when EDTA was used for blood sample collection, yielded lower PON1 values in sick dogs with eccentrocytosis than in healthy dogs, and yielded highly correlated results.


CONCLUSIONS AND CLINICAL RELEVANCE
The methods described here may allow a wider use of PON1 activity as a biomarker of oxidative stress in dogs in clinical and research settings. Results of each method were robust and precise (with the exception of the interassay values for the lactonase method), and the methods were easy to set up in a laboratory.