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- Author or Editor: Asta Tvarijonaviciute x
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Abstract
Objective—To evaluate 5 commercially available ELISAs for determination of leptin concentrations in serum samples from dogs.
Sample Population—Serum samples from overweight-obese and thin–ideal weight clientowned dogs.
Procedures—Serum samples with high and low leptin concentrations (n = 7 samples each) were used for validation of the assays. Intra- and interassay precision, linearity under dilution, spiking recovery, and limit of quantification were determined. In addition, leptin concentrations in thin–ideal weight (n = 8) and overweight-obese (37) dogs were quantified.
Results—Use of 2 of the 5 ELISAs (A and B) revealed reactivity with canine leptin. Intra-and interassay coefficients of variation were < 6.1% and 76%, respectively, for assay A and 14.0% and 13.7%, respectively, for assay B. In assays A and B, dilutions of canine serum pools were used to determine linear regression equations. Recoveries were 77% to 101% for assay A and 67% to 125% for assay B. Significant differences in leptin concentrations between thin–ideal weight and overweight-obese dogs were detected only when analyzed with assay A.
Conclusions and Clinical Relevance—Among 5 leptin ELISAs evaluated, a canine-specific leptin ELISA had adequate precision, linearity, and ability to discriminate between high and low leptin concentrations corresponding to overweight-obese and thin–ideal weight dogs, respectively.
Abstract
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.
