Sandwich hybridization assays

DNA probing can be done not only on a membrane, but also on solid supports. An example is the classical sandwich hybridization assay, which uses a capture... 

Amplification of nucleic acids also include RNA amplifications. For instance, the heat-shock mRNA (103-mer) in viable Cryptosporidium parvum in water treatment plants was detected after amplification by nucleic acid sequence-based amplification (NASBA). After amplification, a sandwich hybridization assay was subsequently conducted within a PDMS channel [955]. 

Figure 20. Schematic representation of the analytical protocol of the biosensor (a) Dual hybridization event of the sandwich hybridization assay, leading to the capture of the CdS-loaded CNT tags in the microwell (b) dissolution of the CdS tracer (c) stripping voltammetric detection of cadmium at a mercury-coated glassy carbon electrode. PI, DNA probe 1 T, DNA target P2, DNA probe 2. From reference 114.

Fig. 5. Sandwich-hybridization assay (A) A biotinylated DNA oligonucleotide, which is complementary to one portion of the target sequence, is mixed with streptavidin and applied to form the capture zone 1.5 cm from the base of the membrane. An unmodified DNA oligonucleotide, which is complementary to the reporter probe sequence on the liposomes, is applied to form the control zone 2.5cm from the base of the membrane. (B) In the presence of target, a sandwich hybridization complex forms with dye-encapsulating liposomes resulting in a magenta-colored band at the capture zone. (C) In the absence of target, only the control band is visible as no sandwich complex has formed.

Corstjens, P. L., Zuiderwijk, M., Nilsson, M., Feindt, H., Sam Niedbala, R., and Tanke, H. J. (2003) Lateral-flow and up-converting phosphor reporters to detect single-stranded nucleic acids in a sandwich-hybridization assay. AnalBiochem. 312, 191-200... 

Rule, G. S., Montagna, R. A., and Durst, R. A. (1997) Characteristics of DNA-tagged liposomes allowing their use in capillary-migration, sandwich-hybridization assays. Anal Biochem. 244, 260-269... 

Sandwich hybridization assay that employs a covalently labeled reporter or signaling probe (RP) and involves two tDNA recognition steps (CP-tDNA and tDNA-RP) [32]. The RPs are designed to hybridize with the tDNA at a site next to the sequence recognized by the capture probe to confer efficient electronic communication between the label and the electrode. 

Figure 9.4. Schematic representation of a impedimetric genosensor [sandwich hybridization assay]. Unmodified PGR products [b] were captured at the sensor interface [a] via sandwich hybridization with the surface-tethered probe and a biotinylated signaling probe. The biotinylated hybrid [c] was then coupled with a streptavidin-alkaline phosphatase conjugate [d] and finally exposed to the substrate solution [e]. The biocatalyzed precipitation of an insulating product [f] blocked the electrical communication between the gold surface and the [Fe(CN]6] redox probe [published by Elsevier in Ref. 50].

Pioch, D., JUrgen, B., Evers, S., Maurer, K. H., Hecker, M., Schweder, T. (2008). Improved sandwich-hybridization assay for an electrical DNA-chip-based monitoring of bioprocess-relevant marker genes. Applied Microbiology and Biotechnology, 78(A), 719-728. http //dx.doi. org/10.1007/s00253-008-1347-z. 

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