Development of an ELISA using a universal method of enzyme-labelling drug-specific antibodies. Part I: Detection of dexamethasone in equine urine.

The research discusses the creation and testing of an ELISA (Enzyme-Linked Immunosorbent Assay) for the detection of the drug dexamethasone in horse urine. This test was validated and used to identify drug residues in unextracted diluted horse urine after a single dexamethasone dose.

Development of the ELISA

• Researchers developed an Enzyme-Linked Immunosorbent Assay (ELISA) specifically for identifying the presence of the drug dexamethasone in equine urine. The ELISA creates a competitive environment where drug-specific antiserum battles against sample or standard drug and an immobilized drug-protein conjugate in the microtitre plate wells.
• The drug-protein conjugate is essentially the drug dexamethasone linked to a protein and immobilized or trapped in the wells of a microtitre plate, which is a flat plate with multiple “wells” that are used as small test tubes.

Antiserum and Detection

• The proportion of antiserum binding to the immobilised drug-protein conjugate is detected using a biotinylated protein G/extravidin-alkaline phosphatase complex in situ.
• This complex reacts with a substrate to create a measurable product, which indicates the level of interaction between the antiserum and the immobilised drug-protein conjugate, and hence the concentration of drug in the sample.

Application and Validation

• The developed ELISA was used to identify presence of drug-derived material in unextracted diluted urine from a horse that was administered with a single intra-muscular dose of dexamethasone at approximately 0.04 mg/kg.
• To ensure accuracy and reliability, the method was validated against a radioimmunoassay and gas chromatography/mass spectrometry (GC/MS) analysis, which are established methods of drug detection.

Overall, the research presents a successful development of a reliable and sensitive method for identifying dexamethasone in equine urine, which could be useful in sporting events for horses and veterinary medicine.