DNA hybridization assays

Collins, M. L., et al. (1995). Preparation and characterization of RNA standards for use in quantitative branched-DNA hybridization assays. Anal. Biochem. 226,120-129. 

Lanthanides also have potential as DEFRET energy donors. Selvin et al. have reported the use of carbostyril-124 complexes (53) with europium and terbium as sensitizers for cyanine dyes (e.g., (54)) in a variety of immunoassays and DNA hybridization assays.138-140 The advantage of this is that the long lifetime of the lanthanide excited state means than it can transfer its excitation energy to the acceptor over a long distance (up to 100 A) sensitized emission from the acceptor, which occurs at a wavelength where there is minimal interference from residual lanthanide emission, is then measured. 

A similar type of biotin-dendritic multimer also was used to boost sensitivity in DNA microarray detection by 100-fold over that obtainable using traditional avidin-biotin reagent systems (Stears, 2000 Striebel et al., 2004). With this system, a polyvalent biotin dendrimer is able to bind many labeled avidin or streptavidin molecules, which may carry enzymes or fluorescent probes for assay detection. In addition, if the biotinylated dendrimer and the streptavidin detection agent is added at the same time, then at the site of a captured analyte, the biotin-dendrimer conjugates can form huge multi-dendrimer complexes wherein avidin or streptavidin detection reagents bridge between more than one dendrimer. Thus, the use of multivalent biotin-dendrimers can become universal enhancers of DNA hybridization assays or immunoassay procedures. 

Diamandis, E.P., and Christopoulos, T.K. (1990) Europium chelate labels in time-resolved fluorescence immunoassays and DNA hybridization assays (Review). Anal. Chem. 62, 1149-1157. 

Z.P. Aguilar and I. Fritsch, Immobilized enzyme-linked DNA-hybridization assay with electrochemical detection for Cryptosporidium parvum hsp70 mRNA. Anal. Chem. 75, 3890-3897 (2003). 

M. Urban, R. Moller, and W. Fritzsche, A paralleled readout system for an electrical DNA-hybridization assay based on a microstructured electrode array. Rev. Sci. Instrum. 74, 1077-1081 (2003). 

In 1987, CL started to be applied in DNA hybridization assays as an alternative to the use of radioactive tags. These assays are based on the specificity of a binding process that of DNA strands for each other. An unknown DNA can be identified with the Southern blot method in which the strands of the analyte are separated and allowed to interact with labeled probe DNA strands on nitrocellulose filter paper. If the label on the probe is detected, the DNA can be identified and, in some cases, quantitated. Conventionally, radioactive tags were used be-... 

All DNA hybridization assays are subject to cross-hybridization, in which an oligonucleotide that is not a perfect sequence match hybridizes with the capture probe. The cross-hybridization in the OFRR was investigated using samples of oligonucleotides with either 0-, 1-, 2-, 5-, or 25-base mismatches when compared with the 25 base-pair biorecognition capture probe. The resulting resonant mode spectral shifts for the respective mismatch are plotted in Fig. 14.7b. The measurements show a difference of 1.3 pm and 2.8 pm for one and two base-pair... 

PPEs in water and prevent aggregation and self-quenching [53]. Heeger and co-workers paired anionic PPV with a nonconjugated cationic polymer to form a charge-neutral complex and saw both reduction of nonspecific effects and some loss in sensitivity in protein detection [54]. Polyelectrolytes may be best used in assay schemes, such as DNA hybridization assays or drug discovery screens, where the assay conditions are well controlled and nonspecific interactions reduced or avoided. 

Leclerc, Ho, and co-workers have developed many sensing systems for different targets using cationic PT and exploiting the differences in emission and color between the extended conjugation form and the coiled form. The basis of these assays is that the polymer microstructure and conformation change as it interacts with the target. For example, in DNA hybridization assays, imidazolium-PT (see... 

The purpose of this study was to prepare a series of random copolymers of NIPAAM with predictable and well defined temperatures of precipitation covering the range of 0 to 45 C, as well as some that precipitate above 55 C under the conditions of high salt used in DNA hybridization assays. The N-substituted acrylfiunides offer the greatest chemical similarity to NIPAAM and therefore should copolymerize randomly with the latter ( ). Thus, copolymers of NIPAAM with AAM, NMAAM, NEAAM, NNBAAM and NTBAAM were prepared at selected monomer ratios and their aqueous solution behavior was evaluated. 

The electrochemical DNA sensor shows impressive sensitivity and selectivity. The detection limit, estimated to be 5 pM, is competitive with previously reported HCMV DNA hybridization assays based on an enzyme label.52 Introduction of a noncomplementary DNA sequence (human ETS2 DNA gene) records an electrochemical signal consistent with background noise. The electrochemical DNA sensor is therefore selective for the HCMV DNA sequence. 

E. P. Diamandis and T. K. Christopoulos, Europium Chelate Labels in Time-Resolved Fluorescence Immunoassays and DNA Hybridization Assays, Anal. Chem. 1990, 62, 1149A. 

Schultz E, Galland R, Du Bouetiez D et al (2008) A novel fluorescence-based array biosensor principle and application to DNA hybridization assays. Biosens Bioelectron 23 987-994... 

Sueda, S., Yuan, J.L., and Matsumoto, K. (2000) Homogeneous DNA hybridization assay by using europium luminescence energy transfer. Bioconjugate Chemistry, 11, 827-831. 

Luminescent RE + chelates have been successfully developed as labels and probes for highly sensitive and selective bioassays in the past two decades. Time-resolved Inmines-cence detection has been widely applied in fluoroimmunoassay, DNA hybridization assay, enzyme assay, cell activity assay and fluorescence imaging microscopy. ... 

In this review chapter, the reader will find information on the following a general descriptive section on the MEF phenomenon, a section on the effects of microwave heating on the components of the MAMEF-based assays, and several applications of the MAMEF technique in ultra fast protein detection assays, immunoassays and DNA hybridization assays. 

Figure 7.9 (A) Model DNA hybridization assay (B) Emission spectra of fluorescein-oligo (3 DM) after MAMEF-based and room temperature hybridization on SiFs, Insert photographs, both before and after hybridization is completed. Adapted from reference 50.

Malicka, J., Gryczynski, 1., and Lakowicz, J. R. (2003). DNA hybridization assays using metal-enhanced fluorescence Biochemical and biophysical research communications 306 213-218. 

Aslan, K., Malyn, S. N., and Geddes, C. D. (2006). Fast and sensitive DNA hybridization assays using microwave-accelerated metal-enhanced fluorescence Biochemical and biophysical research communications 348 612-617. 

The use of fluorophores and nano-structured metal surfaces or adsorbed colloidal particles led to the formation of metal-enhanced planar immunoassay, such as sandwich assay or DNA hybridization assay as illustrated in Scheme 8.9. 

---