Quantitative Formats

In most applications of hybridization assays the target nucleic acid is immobilized onto a filter, where it is then allowed to react with a labeled probe. In the sandwich hybridization reaction, the target sequence is kept in solution while allowed to react with two probes, one for capture, and one for detection (Ranki et al., 1983). The two probes should recognize two essentially adjacent sequences on the target but must not recognize each other. The advantage of sandwich hybridization is that it removes the step in which the target is immobilized. This step is time-consuming but, more important, when crude samples are immobilized biological compounds other than nucleic acids may also be bound causing unspecific binding of detector probe. Furthermore the use of two probes increases the specificity of the reaction and removes the potential error caused by cloned probes recognizing the vector in addition to the actual target (Chou et al., 1983).