SELEX: Critical factors and optimization strategies for successful aptamer selection

Within the last decade, the application range of aptamers in biochemistry and medicine has expanded rapidly. More than just a replacement for antibodies, these intrinsically structured RNA- or DNA-oligonucleotides show great potential for utilization in diagnostics, specific drug delivery, and treatment of certain medical conditions. However, what is analyzed less frequently is the process of aptamer identification known as systematic evolution of ligands by exponential enrichment (SELEX) and the functional mechanisms that lie at its core. SELEX involves numerous singular processes, each of which contributes to the success or failure of aptamer generation. In this review, critical steps during aptamer selection are discussed in-depth, and specific problems are presented along with potential solutions. The discussed aspects include the size and molecule type of the selected target, the nature and stringency of the selection process, the amplification step with its possible PCR bias, the efficient regeneration of RNA or single-stranded DNA, and the different sequencing procedures and screening assays currently available. Finally, useful quality control steps and their role within SELEX are presented. By understanding the mechanisms through which aptamer selection is influenced, the design of more efficient SELEX procedures leading to a higher success rate in aptamer identification is enabled.