Hybrid Capture

Hybrid capture (HC) is a signal amplification system based on antibody binding of RNA:DNA hybrids. The first step of HC is the hybridization of unlabeled RNA probes with denatured target DNA. The resulting RNA:DNA hybrids are captured to the surface of microplate wells by an immobilized antibody that recognizes RNA:DNA hybrids. A second anti-RNA:DNA monoclonal antibody conjugated to alkaline phosphatase then is added. Many secondary antibodies bind to each RNA:DNA hybrid. Dioxetane-based substrate is added, which is cleaved by the alkaline phos-phatase,producing a chemiluminescent signal. The signal is measured using a microplate luminometer and is proportional to the amount of target DNA in the specimen.73 The sensitivity of HC can be increased by using a precipitation buffer and high-speed centrifugation.

Examples of Applications of HC

1. Human papilloma virus detection

2. CMV quantitation74

3. Chlamydia trachomatis and Neisseria gonorrhoeae detection75

4. HBV quantitation76

5. HSV detection77

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