The Application of Microfluidic Technologies in Aptamer Selection

Abstract

Aptamers are sequences of single-strand oligonucleotides (DNA or RNA) with potential binding capability to specific target molecules, which are increasingly used as agents for analysis, diagnosis, and medical treatment. Aptamers are generated by a selection method named systematic evolution of ligands by exponential enrichment (SELEX). Numerous SELEX methods have been developed for aptamer selections. However, the conventional SELEX methods still suffer from high labor intensity, low operation efficiency, and low success rate. Thus, the applications of aptamer with desired properties are limited. With their advantages of low cost, high speed, and upgraded extent of automation, microfluidic technologies have become promising tools for rapid and high throughput aptamer selection. This paper reviews current progresses of such microfluidic systems for aptamer selection. Comparisons of selection performances with discussions on principles, structure, operations, as well as advantages and limitations of various microfluidic-based aptamer selection methods are provided.

Keywords: SELEX; aptamer; microarray; microfluidics; oligonucleotide.

FIGURE 1

Scheme of aptamer selection mechanism. The aptamer selection (SELEX) processes usually consist of four major steps: preparing a library of randomly generated oligonucleotide sequences, incubating with the target ligand, eluting bound sequences, amplifying the bound oligonucleotides by PCR, and obtaining ssDNA ready to undergo next SELEX cycle. Sometimes, negative selections are added to remove the non-specific binding sequences from the pool. With microfluidic-based SELEX, small amounts of fluids can be precisely manipulated inside the microfluidic devices, which can reduce the cost, improve the speed, increase the resolving power, and upgrade the extent of automation compared to the conventional SELEX processes.

FIGURE 2

Microarray chips for aptamer selection. (A) Schematic diagram of the Ag10-NPs-library SPRI-microfluidic-SELEX method. The biochip consists of eight parallel channels on an SAM-gold substrate sealed in a customized flow cell. (B) Principle of microarray-SELEX for aptamer screening. (C) Magnetism-array selection chip. Reprinted with permission from Hong et al. (2017). Copyright 2018 American Chemical Society. (D) Schematic of the PS–SG in vitro selection aptamer chip, where small target molecules are entrapped in a spotted sol–gel microarray, which is then incubated with aptamers.

FIGURE 3

Force field driven microfluidic chips. (A) Dielectrophoresis and electrophoresis combined microfluidic chip for performing cell-SELEX procedure. (B) The schematic overview of the acoustophoretic device and SELEX process. (C) Design of the CMACS microfluidic SELEX device. (D) Myoglobin-aptamer selection microfluidic chip. The positive and negative selection unites are integrated in one micro-channel. Reprinted with permission from Wang et al. (2014). Copyright 2014 American Chemical Society. (E) Inertial microfluidic SELEX (I-SELEX) utilizing Dean vortices to select aptamers for red blood cells (RBCs).

FIGURE 4

Integrated full-SELEX microfluidic systems. (A) Schematic diagram of the integrated microfluidic SELEX chip. It consisted of two PDMS layers and one glass substrate. (B) Integrated microfluidic SELEX. It includes a selection microchamber and an amplification microchamber interconnected by a gel-filled microchannel: top and cross-sectional schematics. A micrograph of the microchip with microbeads retained in the microchambers and agarose gel (dyed blue for visualization) filling the interconnection channel. Inset: Micrograph of the weir-like flow constriction region with retained beads.