Standardization and validation of an agar gel immunodiffusion test for the diagnosis of equine infectious anemia using a recombinant p26 antigen.

This research aimed to develop and validate an agar gel immunodiffusion test using a recombinant p26 antigen for the diagnosis of equine infectious anemia. The results showed this test to be highly effective, producing consistent and accurate results. It is suggested that this testing method could be officially adopted for the diagnosis and control of equine infectious anemia in Argentina.

Development and Validation of the Test

• A new agar gel immunodiffusion test (AGID) was developed for the diagnosis of equine infectious anemia (EIA).
• The test uses the p26 protein obtained from the equine infectious anemia virus (EIAV) as an antigen. This protein was produced in an Escherichia coli expression system.
• The developed test, referred to as the rp26-AGID, was validated and showed excellent diagnostic relative sensitivity and specificity. It was accurate in identifying of EIA in horses.

Testing Methodology and Results

• The validation of rp26-AGID was conducted using 1855 field serum samples. The performance of rp26-AGID was compared to a commercial AGID test, showing a 100% match in sensitivity and specificity.
• In addition to its accuracy, the rp26-AGID displayed excellent repeatability and reproducibility, demonstrating the precision of the test.
• An analytical sensitivity trial was conducted during which positive sera displayed almost identical endpoint dilutions for both compared tests.

Analytical Specificity and Compatibility

• An important feature of rp26-AGID is its analytical specificity. No positive reactions were observed when the test was conducted with 35 serum samples that had antibodies related to other endemic agents.
• Also, no false positives were identified when severely hemolysed samples were tested.
• A 100% match was observed with blind previous results, verifying the accuracy of rp26-AGID in the detection of EIAV antibodies.

Conclusion and Recommendations

• This is reportedly the first time that a recombinant AGID test for EIA has been standardized and validated in Argentina as per international guidelines.
• The rp26-AGID, given its perfect sensitivity and specificity, is recommended as an official testing method for the diagnosis and control of EIA in Argentina.