Equine Infectious Anemia Virus (EIAV), a lentivirus marked by considerable genetic variability, poses significant diagnostic challenges. Existing diagnostic tools encompass the Agar Gel Immunodiffusion Assay (AGID), enzyme-linked immunosorbent assay (ELISA), and Western blotting (WB). ELISA and AGID mainly utilize the p26 capsid protein, often sourced from the Wyoming reference strain. To broaden the range of viral proteins and strains employed in these immunoassays, we previously developed a novel p26/double-strain gp45 indirect ELISA. In this study, we evaluated the performance of this ELISA in comparison to two commercial EIAV ELISAs using Cohen’s Kappa test and Bayesian Latent Class Analysis (BLCA), a statistical method that estimates test performance without requiring a perfect reference standard. A comparison with the official classification of the sera by the Italian Veterinary Service was also performed. A total of 372 serum samples, including 96 that were positives by all three tests, were analyzed. Results from both Cohen’s Kappa test and BLCA, alongside comparison with official classifications, affirm the diagnostic reliability of the two commercial ELISAs and suggest that the novel ELISA, with its enhanced antigenic diversity, could offer an accurate and reliable diagnostic option for EIAV. This novel assay enhances existing commercial ELISAs and has the potential to strengthen routine diagnostic workflows.