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The Oregon Department of Agriculture's official virus testing program for Vaccinium has grown 915% since its inception. To address this increased workload, we evaluated a high-throughput ELISA protocol that uses a 96-well format for sap extraction. To verify the efficacy of the protocol, it was used to test leaves from known infected and healthy plants (positive and negative controls) and from 940 Vaccinium plants grown in a nursery with Blueberry shock virus (BlShV) problems. Each leaf collected was cut in half; one-half was macerated in an extraction buffer using mesh sample bags and a roller press (standard protocol) and the other half was placed into 10 × 96 collection microtubes and macerated in the extraction buffer using a Mixer Mill MM300 (high-throughput protocol). All sample sap extractions were then tested with a commercial ELISA kit. In experiments with the BlShV positive and negative control plants, the high-throughput protocol had high sensitivity (100%) and specificity (100%), matching the standard protocol. In experiments with the blueberry plants of unknown BlShV infection status, the high-throughput protocol had a sensitivity of 94.85% and a specificity of 96.70%, compared to the standard protocol. It is concluded the high-throughput protocol can be used successfully with ELISA to detect BlShV in Accepted for publication 17 May 2009. Published 10 July 2009. Accepted for publication 17 May 2009. Published 10 July 2009. |
