Aptamer Binding on Microarrays
Aptamers are nucleic acid affinity reagents that have diagnostic, therapeutic, and basic research applications. They are short single stranded DNA or RNA molecules that can be selected for their binding ability to target molecules such as proteins. Aptamers offer advantages over many other molecular recognition tools in that they are chemically synthesizable, stable over a wide range of temperatures, can be used in vitro, and can be selected to carry out specific functions.
The characterization of aptamer candidate binding affinity and specificity is a critical but challenging step in the discovery process, as traditional tools are typically low in throughput. However, characterization can be carried out in a high throughput parallel fashion by performing these assays on microarrays.
Using the ArrayIt SpotBot3 microarray printer system now located in the BNL at CNSI, aptamers can be spotted onto the surface of a substrate at known locations. Following the conjugation of nucleic acids to the surface of a microarray slide, binding assays can be performed to evaluate the affinity and specificity of the selected aptamer candidates.
In the image, the left side shows a microarray substrate that has been printed with negative and positive control DNA sequences, prior to carrying out a binding assay. After incubating the slide with a fluorophore labeled protein target, the slide was scanned again, and the results are shown on the right side of the image. These results show that the negative control sequence does not bind to the target protein, whereas the aptamer selected for the target protein and the aptamer selected for the target protein with a 10T spacer both show affinity.
Microarray experiments are extremely valuable in allowing the rapid identification of sequences for further testing in downstream applications.