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Serological Detection of Citrus tristeza virus with Antibodies Developed to the Recombinant Coat Protein

Antibodies specific for the recombinant coat protein (rCP) of the p25 gene of Citrus tristeza virus (CTV) were developed in goats and rabbits and further evaluated as a complete kit for the detection of the virus using healthy and CTV-infected tissue. The combination of goat T1 used as primary (coating) and rabbit C3 as intermediate (detecting) rCP antibodies reacted efficiently, with optical density at 405 nm (OD₄₀₅) values between 0.250 and 2.000 with samples from an international collection of diverse CTV isolates. The CTV isolates tested cause a broad spectrum of disease syndromes in different citrus hosts. The OD₄₀₅ values for healthy tissue were less than 0.100. Likewise, the combination of goat T1 and rabbit C3 rCP antibodies gave consistent results for CTV-positive and -negative sample discrimination when directly compared with the Central California Tristeza Eradication Agency (CCTEA) antibodies used for large-scale CTV detection and a commercially available CTV serological detection kit. The combination of goat T1 and rabbit C3 rCP antibodies showed its suitability for large-scale indexing with samples collected in commercial groves as part of the CCTEA's regular monitoring program. The evaluation included 41,195 samples from 301 commercial groves from districts 1, 2, and 3. In total, 26 trees (0.063%) were found to be CTV positive using the T1/C3 rCP antibody combination. Results of this research provide evidence that rCP antibodies can be efficiently used for both capturing and detecting CTV antigens in double-antibody sandwich indirect enzyme-linked immunosorbent assay.