A TranscriptionallyAmplified DNA Probe Assay with Ligatable Probes and Immunochemical Detection

Transcriptionally amplified DNA probes are valuable tools in the development of sensitive nucleic acid-based diag-nostic assays. Here we describe a model assay using a novel oligonucleotide hairpin probe that encodes a 17 RNA polymerase promoter. The hairpin probe and an adjacently hybridizing biotinylated capture probe were hy-bridized to target DNA and the duplex was captured onto streptavidin-coated magnetic particles. After ligation of the immobilized probes, which served to maintain speci-ficity, the hairpin probe was transcribed by 17 RNA poly-merase. The amplified ANA product was hybridized to the capture probe and bound to the streptavidin-coated mag-netic particles. The immobilized heteroduplex was de-tected with an antibody-alkaline phosphatase conjugate specific for DNA:RNA hybrids, and the chemiluminescen