Aptamers: Novel Molecules as Diagnostic Markers in Bacterial and Viral Infections?

Figure 1

Schematic illustration of the SELEX methodology. The SELEX technique uses a large combinatorial oligonucleic acid library (DNA or RNA) consisting of an inner random region flanked by two constant regions. In the following, attention has solely been drawn on the development of DNA aptamers. The DNA library consisting of partially randomised DNA sequences (inner random region flanked on both sites by constant sequences) is amplified by conventional PCR. The derived double-stranded DNA is denatured and separated into single-stranded DNA by gel electrophoresis, the single-stranded DNA isolated and subsequently incubated with the respective target molecules ((a), positive selection). After incubation step, the formed target-aptamer-complex is separated from nonbinding aptamers and applied to PCR for amplification of the target-bound aptamers (grey panel). Eventually, (b) negatives cycles (blue panel) are also carried out to remove aptamers which bind unspecifically or not to the desired target molecules. As a resulting consequence, the unbound aptamers are recovered, amplified via PCR, and applied in the next (positive) selection cycle. Subsequently, from final selected library aptamer sequences are identified and aligned for the verification of consensus sequence motifs. If required post-SELEX modifications such as truncations, stabilizations, and covalent attachment of fluorescence reporters can be applied to optimize aptamers for any desired purpose.