Integrated Microfluidic Selex Using Free Solution Electrokinetics

Abstract

Systematic evolution of ligands by exponential enrichment (SELEX) offers a powerful method to isolate affinity oligonucleotides known as aptamers, which can then be used in a wide range of applications from drug delivery to biosensing. However, conventional SELEX methods rely on labor intensive and time consuming benchtop operations. A simplified microfluidic approach is presented which allows integration of the affinity selection and amplification stages of SELEX for the isolation of target-binding oligonucleotides by combining bead-based biochemical reactions with free solution electrokinetic oligonucleotide transfer. Free solution electrokinetics allows coupling of affinity selection and amplification for closed loop oligonucleotide enrichment without the need for offline processes, flow handling components or gel components, while bead based selection and amplification allow efficient manipulation of reagents and reaction products thereby realizing on-chip loop closure and integration of the entire SELEX process. Thus the approach is capable of multi-round enrichment of oligonucleotides using simple transfer processes while maintaining a high level of device integration, as demonstrated by the isolation of an aptamer pool against a protein target (IgA) with significantly higher binding affinity than the starting library in approximately 4 hours of processing time.

Recent Microdevice-Based Aptamer Sensors

“…Otherwise, some external molecule may be used as an optical label. Fluorescent materials [ 13 , 17 , 19 , 21 , 28 , 37 , 38 , 42 , 45 , 46 , 47 , 48 , 50 , 53 , 54 , 55 , 56 , 60 , 61 , 68 , 69 , 90 , 91 , 92 , 93 , 94 , 95 , 96 ], and dyes (colorimetry) [ 25 , 64 , 74 , 77 , 78 , 82 , 83 , 84 , 97 , 98 ] are commonly used as labels in microdevices based on aptasensors.…”

“…The entire SELEX process is shortened and the possibility to produce the aptamer as a biorecognition element is increased [ 13 ]. Integration of the affinity selection and amplification steps in SELEX by combining bead-based biochemical reactions has been demonstrated [ 14 , 15 , 16 , 17 , 18 ]. A simple microfluidic SELEX device was developed by Olsen et al [ 17 ], this device was fabricated using single layer soft lithography ( Figure 2 ).…”

“…Integration of the affinity selection and amplification steps in SELEX by combining bead-based biochemical reactions has been demonstrated [ 14 , 15 , 16 , 17 , 18 ]. A simple microfluidic SELEX device was developed by Olsen et al [ 17 ], this device was fabricated using single layer soft lithography ( Figure 2 ). In this work, an electrokinetic microfluidic device for aptamer enrichment was demonstrated as an integrated microfluidic device without requiring an offline process.…”

“…In recent years, microfluidic chips have aroused increasing interest for various application because of their desirable features, such as the smaller sample amount needed and lowered reagent consumption. The substrate materials of microfluidic chips such as polymers (e.g., PDMS, PMMA, PS) [ 26 , 27 , 28 , 29 , 30 , 31 , 32 , 33 , 34 ], ceramics (e.g., glass) [ 6 , 13 , 14 , 16 , 17 , 18 , 19 , 21 , 25 , 35 , 36 , 37 , 38 , 39 , 40 , 41 , 42 , 43 , 44 , 45 , 46 , 47 , 48 , 49 , 50 , 51 , 52 , 53 , 54 , 55 , 56 , 57 , 58 , 59 , 60 , 61 , 62 , 63 ], and semiconduct...…”

“…( a ) Polydimethylsiloxane (PDMS) channel on glass substrate; ( b ) Top view with detailed features; ( c ) Selection and amplification microchamber connected by a single serpentine shaped microchannel. Reproduced with permission from reference [ 17 ]. Copyright 2017 Electrochemical Society.…”